Tokushima University / Educator and Researcher Directory --- Tsuji, Akihiko

Personal Web Page

https://researchmap.jp/tsujiakihiko (researchmap)

Field of Study

○	Biochemistry, Enzyme Chemistry

Subject of Study

○	Molecular basis of regulatory mechanism for cell growth and differentiation by proprotein convertases, Structure and function of plant proteases, Development of animal cellulases by protein engineering (protease, プロセシング, 翻訳後修飾, 血清タンパク, 酵素補充療法)

Book / Paper

Book:

1.	Akihiko Tsuji : 科学と生物実験ライン48 タンパク質と核酸の分離精製基礎と実験(編集:寺田弘), 株式会社廣川書店, 東京都, Jun. 2001.
2.	石田寿昌, 石野良純, Hideaki Nagamune, 岡本敬の介, 梶本和昭, 加藤郁之進, 金保安則, 河野通明, 小池晴彦, 佐藤真悟, 鮫島勇次, Yasuo Shinohara, Akihiko Tsuji, Hiroshi Terada, 富田基郎, 友尾幸司, 野沢義則, 野元裕, 濱崎直孝, 日野真美, 平松宏一, 藤井敏, 藤井儀夫, 洪英満, Yoshiko Matsuda, 森秀治, 山崎尚志, 山本格 and 若原章男 : タンパク質と核酸の分離精製, 株式会社廣川書店, Tokyo, Jun. 2001.
3.	Yoshiko Matsuda, Akihiko Tsuji and Hideaki Nagamune : タンパク質と核酸の分離精製·基礎と実験(共著), 株式会社廣川書店, Tokyo, Jan. 2001. (Summary) 本著作はタンパク質と核酸の精製方法を解析したもので，松田·辻は細胞分画，蛋白質抽出法と精製法および核酸，DNA, plasmid, mRNAの分離，精製法について解説した．また長宗は，微量の生理活性タンパク質を調製して使用する際に必須である濃縮技術の中でも特に利用頻度の高い7種類の方法:塩析法，有機溶媒沈殿法，等電点沈殿法，脱水法，限外濾過法，凍結乾燥法，遠心真空蒸発法を例にとり，その原理や具体的な操作方法について解説を行った．
4.	Hideaki Nagamune, Kazuaki Muramatsu, Masakatsu Higashine, Tetsuya Akamatsu, Yasuhiro Tamai, Yasuo Yonetomi, Akihiko Tsuji, Keisuke Izumi, Takemasa Sakaguchi, Tetsuya Yoshida and Yoshiko Matsuda : Cyclic AMP-inducible procalcitonin processing in thyroidal parafollicular cells is regulated by the Kexin family protease, PC1., IOS Press, Amsterdam, Mar. 1997. (Summary) カルシウム調節ホルモンであるカルシトニンは甲状腺傍濾胞細胞より分泌されるが, 不活性な前駆体として合成された後, プロセシングを受けて初めて活性化される. 本研究ではcAMP刺激により著増するカルシトニンのプロセシングがKexin family proteaseに属する酵素PC1により調節されることを初めて明らかにした. (Keyword) Calcitonin / Processing / PC1
5.	Akihiko Tsuji and Yoshiko Matsuda : 膜酵素基礎と実験II, 株式会社廣川書店, 東京都, May 1990. (Summary) 本書は，生体膜，膜結合酵素の生理学的機能研究法について詳細に解説したものである．
6.	Akihiko Tsuji and Yoshiko Matsuda : 膜酵素基礎と実験I, 株式会社廣川書店, 東京都, May 1990. (Summary) 本書は，生体膜分離法，及び膜結合酵素の全般的な研究法について詳細に解説したものである．
7.	阿久津秀雄, 伊藤明夫, Hideaki Nagamune, 久野健夫, 小南英紀, 田代裕, 田中啓二, Akihiko Tsuji, Hiroshi Terada, 中野明彦, 中安博司, 能見貴人, 日野幸伸, 二井将光, 前田正知, Yoshiko Matsuda, 三原勝芳, 克子山下 and 山本章嗣 : 膜酵素I –基礎と実験-，膜酵素II –基礎と実験-, 株式会社廣川書店, Tokyo, May 1990. (Summary) 本著作は膜酵素の精製や解析法を詳細に解説したものである．第7章では，膜酵素の膜結合様式，界面活性剤の性質，膜酵素の可溶化，精製，純度検定法について解説した．また，第8章(pp.208-239)においては，生物の神経伝達や原尿の再吸収において重要な役割を果たす細胞膜酵素Na,K-ATPaseの触媒サブユニットの構造解析を具体例に取り，蛋白質一次構造をコードするcDNA構造の解析手法を分かりやすく解説した．その際に必要となるDNAやRNAの取り扱い技術，クローニング技術なども，初心者にも分かりやすい表形式にして巻末(第二巻末: pp455-491)に紹介している．
8.	Akihiko Tsuji and Yoshiko Matsuda : 電気泳動法基礎と実験, 株式会社廣川書店, 東京都, Apr. 1989. (Summary) 本書は，最新の電気泳動の理論及び実験法を解説したものである．
9.	市川哲生, Fusao Ota, Hideaki Nagamune, 久保兼信, 桜林郁之介, 佐藤真悟, Yasuo Shinohara, 高木俊夫, 竹尾和典, 田中経彦, Akihiko Tsuji, Hiroshi Terada, 中尾真, 中村和行, 藤田清貴, Yoshiko Matsuda and 真鍋敬 : 電気泳動法 –基礎と実験-, 株式会社廣川書店, Tokyo, Apr. 1989. (Summary) 本著作は，タンパク質の電気泳動法の代表例を挙げてその原理や具体的実施法を詳細に解説している．辻らは等電点電気泳動の原理，方法，応用例を解説した．長宗らは，第7章(pp.255-305)においてゲル電気泳動後にゲル中で蛋白質を検出したり，泳動後の蛋白質を転写した膜上で特定の分子だけを検出する場合に汎用される方法の原理と具体的操作法を分かりやすく解説し，第9章(pp.329-344)では，泳動後のゲルの保存方法やゲル中からの蛋白質の回収方法を詳しく述べた．また第11章(pp.371-410)においては，電気泳動に用いられる各種試薬や標準蛋白質の性状について詳述するともに蛋白質の検出法や染色法の各論について，分かりやすい表形式で紹介した．
10.	Kichiko Koike, Akihiko Tsuji, Yoshishige Urata, Matsuko Moriyasu and Masahiko Koike : Cell-free biosynthesis of lipoamide dehydrogenase, Walter de Gruyter & Co., Germany, Jan. 1984. (Summary) 本研究は，豚肝臓よりポリゾーム免疫沈降法よりリポアミド脱水素酵素mRNAを精製し，その精製mRNAをウサギ網状赤血球ライゼートにて翻訳させ，SDS-ゲル電気泳動法によりその分子量を求めたものである．
11.	Nobuhiko Katunuma, Yoshiko Matsuda, Akihiko Tsuji and Tsutomu Miura : Release and Modulation of Membrane-bound γ-Glutamyltranspeptidase by Limited Proteolysis, Springer-Verlag, Berlin, Jan. 1981. (Summary) 本研究は，γ-グルタミルトランスペプチダーゼの膜結合様式，可溶化様式，疾病との関連について調べた．

Academic Paper (Judged Full Paper):

1.	Mika Nishida, Kenji Miyamoto, Shogo Abe, Maki Shimada, Yuki Shimizu, Akihiko Tsuji and Keizo Yuasa : Natriuretic peptide receptor-C releases and activates guanine nucleotide-exchange factor H1 in a ligand-dependent manner., Biochemical and Biophysical Research Communications, Vol.552, 9-16, 2021. (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2021.03.028 (Link to Search Site for Scientific Articles) ● Summary page in Scopus @ Elsevier: 2-s2.0-85102492627 (DOI: 10.1016/j.bbrc.2021.03.028, Elsevier: Scopus)
2.	Xiaomei Sun, Yuxin Ye, Naofumi Sakurai, Koji Kato, Keizo Yuasa, Akihiko Tsuji and Min Yao : Crystallographic analysis of Eisenia hydrolysis-enhancing protein using a long wavelength for native-SAD phasing., Acta Crystallographica. Section F, Structural Biology Communications, Vol.76, No.1, 20-24, 2020. (Link to Publication Site) ● Publication site (DOI): 10.1107/S2053230X19016716 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 31929182 ● Search Scopus @ Elsevier (PMID): 31929182 ● Search Scopus @ Elsevier (DOI): 10.1107/S2053230X19016716 (DOI: 10.1107/S2053230X19016716, PubMed: 31929182)
3.	Akihiko Tsuji and Keizo Yuasa : Identification and enzymatic characterization of clip domain serine protease in the digestive fluid of the sea hare, Aplysia kurodai., Comparative Biochemistry and Physiology. Part B, Biochemistry & Molecular Biology, Vol.237, 110322, 2019. (Summary) Clip domain serine proteases (CDSPs) participate in the extracellular signaling cascades of various biological processes such as innate immune responses in invertebrates. CDSP genes have been isolated from numerous invertebrates. Nevertheless, the enzymatic properties of mollusk CDSPs are poorly understood. In the present study, we demonstrated that the amino acid sequences of the trypsin-like serine protease purified from the digestive fluid of the sea hare, Aplysia kurodai resemble those of the unidentified CDSP-type protein (TPS3) of Aplysia californica predicted by genome analysis. The purified enzyme produced single 34 and 26.5 kDa bands on SDS-PAGE under non-reducing and reducing conditions, respectively. The 34-kDa band generated two amino-terminal sequences that were similar to the deduced sequences of the clip and catalytic domains of TPS3. The single amino-terminal sequence of the 26.5 kDa band showed a single sequence homologous to the catalytic domain. Thus, the purified enzyme consists of clip and catalytic domains bridged by disulfide linkage(s). The subsite specificity and inhibitor sensitivity of the purified enzyme were clearly distinct from those of horseshoe crab and silkworm CDSPs. A good substrate for the sea hare enzyme was pyroglutamyl-Arg-Thr-Lys-Arg-4-methyl-7-coumarylamide. The enzyme activity was strongly inhibited by aprotinin but not leupeptin. The physiological function of the enzyme in the digestive fluid remains to be determined. (Keyword) Animals / Aplysia (Link to Publication Site) ● Publication site (DOI): 10.1016/j.cbpb.2019.110322 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 31454681 ● Summary page in Scopus @ Elsevier: 2-s2.0-85071372814 (DOI: 10.1016/j.cbpb.2019.110322, PubMed: 31454681, Elsevier: Scopus)
4.	Akihiko Tsuji, Keizo Yuasa and Chikako Asada : Cellulose-binding activity of a 21-kDa endo-ß-1,4-glucanase lacking cellulose-binding domain and its synergy with other cellulases in the digestive fluid of Aplysia kurodai, PLoS ONE, Vol.13, No.11, e0205915, 2018. (Summary) Endo-ß-1,4-glucanase AkEG21 belonging to glycosyl hydrolase family 45 (GHF45) is the most abundant cellulase in the digestive fluid of sea hare (Aplysia kurodai). The specific activity of this 21-kDa enzyme is considerably lower than those of other endo ß-1,4-glucanases in the digestive fluid of A. kurodai, therefore its role in whole cellulose hydrolysis by sea hare is still uncertain. Although AkEG21 has a catalytic domain without a cellulose binding domain, it demonstrated stable binding to cellulose fibers, similar to that of fungal cellobiohydrolase (CBH) 1 and CBH 2, which is strongly inhibited by cellohexaose, suggesting the involvement of the catalytic site in cellulose binding. Cellulose-bound AkEG21 hydrolyzed cellulose to cellobiose, cellotriose and cellotetraose, but could not digest an external substrate, azo-carboxymethyl cellulose. Cellulose hydrolysis was considerably stimulated by the synergistic action of cellulose-bound AkEG21 and AkEG45, another ß-1,4-endoglucanase present in the digestive fluid of sea hare; however no synergy in carboxymethylcellulose hydrolysis was observed. When AkEG21 was removed from the digestive fluid by immunoprecipitation, the cellulose hydrolyzing activity of the fluid was significantly reduced, indicating a critical role of AkEG21 in cellulose hydrolysis by A. kurodai. These findings suggest that AkEG21 is a processive endoglucanase functionally equivalent to the CBH, which provides a CBH-independent mechanism for the mollusk to digest seaweed cellulose to glucose. (Keyword) Animals / Aplysia / Catalytic Domain / Cellobiose / Cellulase / Cellulose / Digestion / Glucose / Hydrolysis / Kinetics / Oligosaccharides / Protein Binding / Protein Domains / Tetroses (Tokushima University Institutional Repository) ● Metadata: 112918 (Link to Publication Site) ● Publication site (DOI): 10.1371/journal.pone.0205915 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 30412581 ● Summary page in Scopus @ Elsevier: 2-s2.0-85056302997 (Tokushima University Institutional Repository: 112918, DOI: 10.1371/journal.pone.0205915, PubMed: 30412581, Elsevier: Scopus)
5.	Shogo Abe, Saki Hirose, Mami Nishitani, Ichiro Yoshida, Masao Tsukayama, Akihiko Tsuji and Keizo Yuasa : Citrus peel polymethoxyflavones, sudachitin and nobiletin, induce distinct cellular responses in human keratinocyte HaCaT cells, Bioscience, Biotechnology, and Biochemistry, Vol.82, No.12, 2064-2071, 2018. (Link to Publication Site) ● Publication site (DOI): 10.1080/09168451.2018.1514246 (Link to Search Site for Scientific Articles) ● Search Scopus @ Elsevier (DOI): 10.1080/09168451.2018.1514246 (DOI: 10.1080/09168451.2018.1514246)
6.	Yuna Oue, Sara Murakami, Kinuka Isshiki, Akihiko Tsuji and Keizo Yuasa : Intracellular localization and binding partners of death associated protein kinase-related apoptosis-inducing protein kinase 1., Biochemical and Biophysical Research Communications, Vol.496, No.4, 1222-1228, 2018. (Summary) Death associated protein kinase (DAPK)-related apoptosis-inducing protein kinase (DRAK)-1 is a positive apoptosis regulator. However, the molecular mechanisms underlying the DRAK1-mediated apoptotic pathway remain unclear. In this study, we demonstrated the intracellular localization and binding partners of DRAK1. In human osteosarcoma cell line U2OS cells, DRAK1 was mainly localized in the nucleus and translocated outside the nucleus through Ser phosphorylation by protein kinase C. In the nucleus, DRAK1 associated with tumor suppressor p53 and positively regulated p53 transcriptional activity in response to DNA-damaging agent cisplatin. On the other hand, DRAK1 interacted with the mitochondrial inner-membrane protein, adenine nucleotide translocase (ANT)-2, an anti-apoptotic oncoprotein, outside the nucleus. These findings suggest that DRAK1 translocates in response to stimuli and induces apoptosis through its interaction with specific binding partners, p53 and/or ANT2. (Keyword) Adenine Nucleotide Translocator 2 / Apoptosis / Apoptosis Regulatory Proteins / Cell Line, Tumor / Humans / Osteosarcoma / Protein Binding / Protein-Serine-Threonine Kinases / Subcellular Fractions / Tissue Distribution / Tumor Suppressor Protein p53 (Tokushima University Institutional Repository) ● Metadata: 111888 (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2018.01.175 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 29397938 ● Summary page in Scopus @ Elsevier: 2-s2.0-85041573302 (Tokushima University Institutional Repository: 111888, DOI: 10.1016/j.bbrc.2018.01.175, PubMed: 29397938, Elsevier: Scopus)
7.	Shinya Matsuda, Kohei Kawamoto, Kenji Miyamoto, Akihiko Tsuji and Keizo Yuasa : PCTK3/CDK18 regulates cell migration and adhesion by negatively modulating FAK activity, Scientific Reports, Vol.7, 45545, 2017. (Summary) PCTAIRE kinase 3 (PCTK3) is a member of the cyclin dependent kinase family, but its physiologicalfunction remains unknown. We previously reported that PCTK3-knockdown HEK293T cells showedactin accumulation at the leading edge, suggesting that PCTK3 is involved in the regulation of actinreorganization. In this study, we investigated the physiological function and downstream signaltransduction molecules of PCTK3. PCTK3 knockdown in HEK293T cells increased cell motility and RhoA/Rho-associated kinase activity as compared with control cells. We also found that phosphorylationat residue Tyr-397 in focal adhesion kinase (FAK) was increased in PCTK3-knockdown cells. FAKphosphorylation at Tyr-397 was increased in response to fibronectin stimulation, whereas itsphosphorylation was suppressed by PCTK3. In addition, excessive expression of PCTK3 led to theformation of filopodia during the early stages of cell adhesion in HeLa cells. These results indicate thatPCTK3 controls actin cytoskeleton dynamics by negatively regulating the FAK/Rho signaling pathway. (Tokushima University Institutional Repository) ● Metadata: 111881 (Link to Publication Site) ● Publication site (DOI): 10.1038/srep45545 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 28361970 ● Search Scopus @ Elsevier (PMID): 28361970 ● Search Scopus @ Elsevier (DOI): 10.1038/srep45545 (Tokushima University Institutional Repository: 111881, DOI: 10.1038/srep45545, PubMed: 28361970)
8.	Norio Kamemura, Sara Murakami, Hiroaki Komatsu, Masahiro Sawanoi, Kenji Miyamoto, Kazumi Ishidoh, Koji Kishimoto, Akihiko Tsuji and Keizo Yuasa : Type II cGMP-dependent protein kinase negatively regulates fibroblast growth factor signaling by phosphorylating Raf-1 at serine 43 in rat chondrosarcoma cells., Biochemical and Biophysical Research Communications, Vol.483, No.1, 82-87, 2017. (Summary) Although type II cGMP-dependent protein kinase (PKGII) is a major downstream effector of cGMP in chondrocytes and attenuates the FGF receptor 3/ERK signaling pathway, its direct target proteins have not been fully explored. In the present study, we attempted to identify PKGII-targeted proteins, which are associated with the inhibition of FGF-induced MAPK activation. Although FGF2 stimulation induced the phosphorylation of ERK1/2, MEK1/2, and Raf-1 at Ser-338 in rat chondrosarcoma cells, pretreatment with a cell-permeable cGMP analog strongly inhibited their phosphorylation. On the other hand, Ser-43 of Raf-1 was phosphorylated by cGMP in a dose-dependent manner. Therefore, we examined the direct phosphorylation of Raf-1 by PKGII. Wild-type PKGII phosphorylated Raf-1 at Ser-43 in a cGMP-dependent manner, but a PKGII D412A/R415A mutant, which has a low affinity for cGMP, did not. Finally, we found that a phospho-mimic mutant, Raf-1 S43D, suppressed FGF2-induced MAPK pathway. These results suggest that PKGII counters FGF-induced MEK/ERK activation through the phosphorylation of Raf-1 at Ser-43 in chondrocytes. (Tokushima University Institutional Repository) ● Metadata: 111887 (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2017.01.001 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 28057484 ● Summary page in Scopus @ Elsevier: 2-s2.0-85008689720 (Tokushima University Institutional Repository: 111887, DOI: 10.1016/j.bbrc.2017.01.001, PubMed: 28057484, Elsevier: Scopus)
9.	Akihiko Tsuji, Shuji Kuwamura, Akihiro Shirai and Keizo Yuasa : Identification and Characterization of a 25 kDa Protein That Is Indispensable for the Efficient Saccharification of Eisenia bicyclis in the Digestive Fluid of Aplysia kurodai, PLoS ONE, Vol.12, No.1, e0170669, 2017. (Summary) The digestive fluid of the sea hare Aplysia kurodai can liberate approximately 2.5 mg of glucose from 10 mg of dried Eisenia bicyclis powder. Although laminaran, a major storage polysaccharide in E. bicyclis, is easily digested to glucose by the synergistic action of the 110 and 210 kDa A. kurodai -glucosidases (BGLs), glucose is not liberated from E. bicyclis by direct incubation with these BGLs. To clarify this discrepancy, we searched for an Eisenia hydrolysis enhancing protein (EHEP) in the digestive fluid of A. kurodai. A novel 25 kDa protein that enhances E. bicyclis saccharification by -glucosidases was purified to a homogeneous state from the digestive fluid of A. kurodai, and its cDNA was cloned from total cDNAs reverse-transcribed from hepatopancreas total RNA. The E. bicyclis extract strongly inhibited BGLs, suggesting some compound within this brown alga functioned as a feeding deterrent. However, when E. bicyclis was incubated with BGLs in the presence of EHEP, glucose production was markedly increased. As E. bicyclis is rich in phlorotannin, which are only found in brown algae, our study suggested that these compounds are the main BGL inhibitors in E. bicyclis extract. EHEP protects BGLs from phlorotannin inhibition by binding to phlorotannins and forming an insoluble complex with phloroglucinol and phlorotannins. These findings indicated that EHEP plays a key role in the saccharification of brown seaweeds containing phlorotannins in the digestive fluid of A. kurodai. This is the first report of EHEP as a phlorotannin-binding protein that protects BGLs from inhibition. (Tokushima University Institutional Repository) ● Metadata: 111878 (Link to Publication Site) ● Publication site (DOI): 10.1371/journal.pone.0170669 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 28129373 ● Summary page in Scopus @ Elsevier: 2-s2.0-85010878231 (Tokushima University Institutional Repository: 111878, DOI: 10.1371/journal.pone.0170669, PubMed: 28129373, Elsevier: Scopus)
10.	Ichiro Yoshida, Chihiro Ito, Shinya Matsuda, Akihiko Tsuji, Noriyuki Yanaka and Keizo Yuasa : Alisol B, a triterpene from Alismatis rhizoma (dried rhizome of Alisma orientale), inhibits melanin production in murine B16 melanoma cells, Bioscience, Biotechnology, and Biochemistry, Vol.81, No.3, 534-540, 2017. (Summary) To develop new whitening agents from natural products, we screened 80 compounds derived from crude drugs in Kampo medicine in a melanin synthesis inhibition assay using murine B16 melanoma cells. The screen revealed that treatment with alisol B, a triterpene from Alismatis rhizoma, significantly decreased both melanin content and cellular tyrosinase activity in B16 cells. However, alisol B did not directly inhibit mushroom tyrosinase activity in vitro. Therefore, we investigated the mechanism underlying the inhibitory effect of alisol B on melanogenesis. Alisol B suppressed mRNA induction of tyrosinase and its transcription factor, microphthalmia-associated transcription factor (MITF). Furthermore, alisol B reduced the phosphorylation of CREB and maintained the activation of ERK1/2. These results suggest that the reduction in melanin production by alisol B is due to the downregulation of MITF through the suppression of CREB and activation of ERK and that alisol B may be useful as a new whitening agent. (Keyword) Alisma / Animals / Cell Line, Tumor / Cholestenones / Cyclic AMP Response Element-Binding Protein / Drug Evaluation, Preclinical / Melanins / Melanoma, Experimental / Mice / Microphthalmia-Associated Transcription Factor / Monophenol Monooxygenase / Phosphorylation / Rhizome / Signal Transduction / Skin Lightening Preparations (Tokushima University Institutional Repository) ● Metadata: 111886 (Link to Publication Site) ● Publication site (DOI): 10.1080/09168451.2016.1268042 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 28051915 ● Summary page in Scopus @ Elsevier: 2-s2.0-85012247112 (Tokushima University Institutional Repository: 111886, DOI: 10.1080/09168451.2016.1268042, PubMed: 28051915, Elsevier: Scopus)
11.	Kohichi Kuwahara, Hiroshi Hirata, Kengo Ohbuchi, Kentaro Nishi, Akira Maeda, Akihiko Kuniyasu, Daisuke Yamada, Takehiko Maeda, Akihiko Tsuji, Makoto Sawada and Hitoshi Nakayama : The novel monoclonal antibody 9F5 reveals expression of a fragment of GPNMB/osteoactivin processed by furin-like protease(s) in a subpopulation of microglia in neonatal rat brain, Glia, Vol.64, No.11, 1938-1961, 2016. (Tokushima University Institutional Repository) ● Metadata: 111877 (Link to Publication Site) ● Publication site (DOI): 10.1002/glia.23034 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 27464357 ● Summary page in Scopus @ Elsevier: 2-s2.0-84990849595 (Tokushima University Institutional Repository: 111877, DOI: 10.1002/glia.23034, PubMed: 27464357, Elsevier: Scopus)
12.	Kinuka Isshiki, Taishi Hirase, Shinya Matsuda, Kenji Miyamoto, Akihiko Tsuji and Keizo Yuasa : Death-associated protein kinase 2 mediates nocodazole-induced apoptosis through interaction with tubulin, Biochemical and Biophysical Research Communications, Vol.468, No.1-2, 113-118, 2015. (Summary) Death-associated protein kinase 2 (DAPK2) is a positive regulator of apoptosis. Although we recently reported that 14-3-3 proteins inhibit DAPK2 activity and its subsequent apoptotic effects via binding to DAPK2, the molecular mechanisms underlying the DAPK2-mediated apoptotic pathway remain unclear. Therefore, we attempted to further identify DAPK2-interacting proteins using pull-down assays and mass spectrometry. The microtubule β-tubulin was identified as a novel DAPK2-binding protein in HeLa cells. Pull-down assays revealed that DAPK2 interacted with the α/β-tubulin heterodimer, and that the C-terminal region of DAPK2, which differs from that of other DAPK family members, was sufficient for the association with β-tubulin. Although the microtubule-depolymerizing agent nocodazole induced apoptosis in HeLa cells, the level of apoptosis was significantly decreased in the DAPK2 knockdown cells. Furthermore, we found that treatment with nocodazole resulted in an increased binding of DAPK2 to β-tubulin. These findings indicate that DAPK2 mediates nocodazole-induced apoptosis via binding to tubulin. (Keyword) apoptosis / phosphorylation (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2015.10.151 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 26529546 ● Search Scopus @ Elsevier (PMID): 26529546 ● Search Scopus @ Elsevier (DOI): 10.1016/j.bbrc.2015.10.151 (DOI: 10.1016/j.bbrc.2015.10.151, PubMed: 26529546)
13.	Yuki Yoshikatsu, Yo-ichi Ishida, Haruka Sudo, Keizo Yuasa, Akihiko Tsuji and Masami Nagahama : NVL2, a nucleolar AAA-ATPase, is associated with the nuclear exosome and is involved in pre-rRNA processing, Biochemical and Biophysical Research Communications, Vol.464, No.3, 780-786, 2015. (Summary) Nuclear VCP-like 2 (NVL2) is a member of the chaperone-like AAA-ATPase family and is involved in the biosynthesis of 60S ribosomal subunits in mammalian cells. We previously showed the interaction of NVL2 with a DExD/H-box RNA helicase MTR4/DOB1, which is a known cofactor for an exoribonuclease complex, the exosome. This finding implicated NVL2 in RNA metabolic processes during ribosome biogenesis. In the present study, we found that a series of mutations within the ATPase domain of NVL2 causes a defect in pre-rRNA processing into mature 28S and 5.8S rRNAs. Co-immunoprecipitation analysis showed that NVL2 was associated with the nuclear exosome complex, which includes RRP6 as a nucleus-specific catalytic subunit. This interaction was prevented by depleting either MTR4 or RRP6, indicating their essential role in mediating this interaction with NVL2. Additionally, knockdown of MPP6, another cofactor for the nuclear exosome, also prevented the interaction by causing MTR4 to dissociate from the nuclear exosome. These results suggest that NVL2 is involved in pre-rRNA processing by associating with the nuclear exosome complex and that MPP6 is required for maintaining the integrity of this rRNA processing complex. (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2015.07.032 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 26166824 ● Summary page in Scopus @ Elsevier: 2-s2.0-84938740493 (DOI: 10.1016/j.bbrc.2015.07.032, PubMed: 26166824, Elsevier: Scopus)
14.	Keizo Yuasa, Reina Ota, Matsuda Shinya, Kinuka Isshiki, Masahiro Inoue and Akihiko Tsuji : Suppression of death-associated protein kinase 2 by interaction with 14-3-3 proteins, Biochemical and Biophysical Research Communications, Vol.464, No.1, 70-75, 2015. (Summary) Death-associated protein kinase 2 (DAPK2), a Ca(2+)/calmodulin-regulated serine/threonine kinase, induces apoptosis. However, the signaling mechanisms involved in this process are unknown. Using a proteomic approach, we identified 14-3-3 proteins as novel DAPK2-interacting proteins. The 14-3-3 family has the ability to bind to phosphorylated proteins via recognition of three conserved amino acid motifs (mode 1-3 motifs), and DAPK2 contains the mode 3 motif ((pS/pT)X1-2-COOH). The interaction of 14-3-3 proteins with DAPK2 was dependent on the phosphorylation of Thr(369), and effectively suppressed DAPK2 kinase activity and DAPK2-induced apoptosis. Furthermore, we revealed that the 14-3-3 binding site Thr(369) of DAPK2 was phosphorylated by the survival kinase Akt. Our findings suggest that DAPK2-induced apoptosis is negatively regulated by Akt and 14-3-3 proteins. (Keyword) apoptosis / phosphorylation (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2015.05.105 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 26047703 ● Summary page in Scopus @ Elsevier: 2-s2.0-84937632159 (DOI: 10.1016/j.bbrc.2015.05.105, PubMed: 26047703, Elsevier: Scopus)
15.	Kim Hyejin, Atsushi Tabata, Toshifumi Tomoyasu, Ueno Tomomi, Uchiyama Shigeto, Keizo Yuasa, Akihiko Tsuji and Hideaki Nagamune : Estrogen stimuli promote osteoblastic differentiation via the subtilisin-like proprotein convertase PACE4 in MC3T3-E1 cells., Journal of Bone and Mineral Metabolism, Vol.33, No.1, 30-39, 2015. (Summary) Estrogenic compounds include endogenous estrogens such as estradiol as well as soybean isoflavones, such as daidzein and its metabolite equol, which are known phytoestrogens that prevent osteoporosis in postmenopausal women. Indeed, mineralization of MC3T3-E1 cells, a murine osteoblastic cell line, was significantly decreased in medium containing fetal bovine serum treated with charcoal-dextran to deplete endogenous estrogens, but estradiol and these soybean isoflavones dose-dependently restored the differentiation of MC3T3-E1 cells; equal was tenfold more effective than daidzein. These differentiation promoting effects were inhibited by the addition of fulvestrant, which is a selective downregulator of estrogen receptors. Analysis of the expression pattern of bone-related genes by reverse transcription PCR (RT-PCR)/quantitative real-time PCR (qRT-PCR), which focused on responsiveness to the estrogen stimuli, revealed that the transcription of PACE4, a subtilisin-like proprotein convertase, was tightly linked with the differentiation of MC3T3-E1 cells induced by estrogen stimuli. Moreover, treatment with RNAi of PACE4 in MC3T3-E1 cells resulted in a drastic decrease of mineralization in the presence of estrogen stimuli. These results strongly suggest that PACE4 participates in bone formation at least in osteoblast differentiation, and estrogen receptor-mediated stimuli induce osteoblast differentiation through the upregulation of PACE4 expression. (Keyword) osteoblast / PACE4 / Estrogen / Soybean / isoflavone (Link to Publication Site) ● Publication site (DOI): 10.1007/s00774-014-0567-9 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 24557631 ● Summary page in Scopus @ Elsevier: 2-s2.0-84893948353 (DOI: 10.1007/s00774-014-0567-9, PubMed: 24557631, Elsevier: Scopus)
16.	Shinya Matsuda, Kyohei Kominato, Shizuyo Koide-Yoshida, Kenji Miyamoto, Kinuka Isshiki, Akihiko Tsuji and Keizo Yuasa : PCTAIRE Kinase 3/Cyclin-dependent Kinase 18 Is Activated through Association with Cyclin A and/or Phosphorylation by Protein Kinase A, The Journal of Biological Chemistry, Vol.289, No.26, 18387-18400, 2014. (Summary) PCTAIRE kinase 3 (PCTK3)/cyclin-dependent kinase 18 (CDK18) is an uncharacterized member of the CDK family because its activator(s) remains unidentified. Here we describe the mechanisms of catalytic activation of PCTK3 by cyclin A2 and cAMP-dependent protein kinase (PKA). Using a pulldown experiment with HEK293T cells, cyclin A2 and cyclin E1 were identified as proteins that interacted with PCTK3. An in vitro kinase assay using retinoblastoma protein as the substrate showed that PCTK3 was specifically activated by cyclin A2 but not by cyclin E1, although its activity was lower than that of CDK2. Furthermore, immunocytochemistry analysis showed that PCTK3 colocalized with cyclin A2 in the cytoplasm and regulated cyclin A2 stability. Amino acid sequence analysis revealed that PCTK3 contained four putative PKA phosphorylation sites. In vitro and in vivo kinase assays showed that PCTK3 was phosphorylated by PKA at Ser(12), Ser(66), and Ser(109) and that PCTK3 activity significantly increased via phosphorylation at Ser(12) by PKA even in the absence of cyclin A2. In the presence of cyclin A2, PCTK3 activity was comparable to CDK2 activity. We also found that PCTK3 knockdown in HEK293T cells induced polymerized actin accumulation in peripheral areas and cofilin phosphorylation. Taken together, our results provide the first evidence for the mechanisms of catalytic activation of PCTK3 by cyclin A2 and PKA and a physiological function of PCTK3. (Keyword) Animals / Cell Line / Cyclic AMP-Dependent Protein Kinase Catalytic Subunits / Cyclin A2 / Cyclin-Dependent Kinases / Enzyme Activation / Gene Expression Regulation, Enzymologic / Humans / Mice / Phosphorylation / Protein Binding (Link to Publication Site) ● Publication site (DOI): 10.1074/jbc.M113.542936 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 24831015 ● Search Scopus @ Elsevier (PMID): 24831015 ● Search Scopus @ Elsevier (DOI): 10.1074/jbc.M113.542936 (DOI: 10.1074/jbc.M113.542936, PubMed: 24831015)
17.	Akihiko Tsuji, Nami Nishiyama, Miki Ohshima, Saori Maniwa, Shuji Kuwamura, Masataka Shiraishi and Keizo Yuasa : Comprehensive enzymatic analysis of the amylolytic system in the digestive fluid of the sea hare, Aplysia kurodai: Unique properties of two α-amylases and two α-glucosidases, FEBS Open Bio, Vol.4, 560-570, 2014. (Summary) Sea lettuce (Ulva pertusa) is a nuisance species of green algae that is found all over the world. East-Asian species of the marine gastropod, the sea hare Aplysia kurodai, shows a clear feeding preference for sea lettuce. Compared with cellulose, sea lettuce contains a higher amount of starch as a storage polysaccharide. However, the entire amylolytic system in the digestive fluid of A. kurodai has not been studied in detail. We purified α-amylases and α-glucosidases from the digestive fluid of A. kurodai and investigated the synergistic action of these enzymes on sea lettuce. A. kurodai contain two α-amylases (59 and 80 kDa) and two α-glucosidases (74 and 86 kDa). The 59-kDa α-amylase, but not the 80-kDa α-amylase, was markedly activated by Ca(2+) or Cl(-). Both α-amylases degraded starch and maltoheptaose, producing maltotriose, maltose, and glucose. Glucose production from starch was higher with 80-kDa α-amylase than with 59-kDa α-amylase. Kinetic analysis indicated that 74-kDa α-glucosidase prefers short α-1,4-linked oligosaccharide, whereas 86-kDa α-glucosidase prefers large α-1,6 and α-1,4-linked polysaccharides such as glycogen. When sea lettuce was used as a substrate, a 2-fold greater amount of glucose was released by treatment with 59-kDa α-amylase and 74-kDa α-glucosidase than by treatment with 45-kDa cellulase and 210-kDa β-glucosidase of A. kurodai. Unlike mammals, sea hares efficiently digest sea lettuce to glucose by a combination of two α-amylases and two α-glucosidases in the digestive fluids without membrane-bound maltase-glucoamylase and sucrase-isomaltase complexes. (Link to Publication Site) ● Publication site (DOI): 10.1016/j.fob.2014.06.002 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 25161866 ● Search Scopus @ Elsevier (PMID): 25161866 ● Search Scopus @ Elsevier (DOI): 10.1016/j.fob.2014.06.002 (DOI: 10.1016/j.fob.2014.06.002, PubMed: 25161866)
18.	Junki Fukumoto, Mohd Nor Ismaliza Ismail, Masaki Kubo, Keita Kinoshita, Masahiro Inoue, Keizo Yuasa, Makoto Nishimoto, Hitoshi Matsuki and Akihiko Tsuji : Possible role of inter-domain salt bridges in oligopeptidase B from Trypanosoma brucei: critical role of Glu172 of non-catalytic -propeller domain in catalytic activity and Glu490 of catalytic domain in stability of OPB., The Journal of Biochemistry, Vol.154, No.5, 465-473, 2013. (Summary) Oligopeptidase B (OPB) is a member of the prolyl oligopeptidase (POP) family of serine proteases. OPB in trypanosomes is an important virulence factor and potential pharmaceutical target. Characteristic structural features of POP family members include lack of a propeptide and presence of a -propeller domain (PD), although the role of the -PD has yet to be fully understood. In this work, residues Glu(172), Glu(490), Glu(524) and Arg(689) in Trypanosoma brucei OPB (Tb OPB), which are predicted to form inter-domain salt bridges, were substituted for Gln and Ala, respectively. These mutants were evaluated in terms of catalytic properties and stability. A negative effect on kcat/Km was obtained following mutation of Glu(172) or Arg(689). In contrast, the E490Q mutant exhibited markedly decreased thermal stability, although this mutation had less effect on catalytic properties compared to the E172Q and R689A mutants. Trypsin digestion showed that the boundary regions between the -PD and catalytic domains (CDs) of the E490Q mutant are unfolded with heat treatment. These results indicated that Glu(490) in the CD plays a role in stabilization of Tb OPB, whereas Glu(172) in the -PD is critical for the catalytic activity of Tb OPB. (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvt077 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 23946505 ● Search Scopus @ Elsevier (PMID): 23946505 ● Search Scopus @ Elsevier (DOI): 10.1093/jb/mvt077 (DOI: 10.1093/jb/mvt077, PubMed: 23946505)
19.	Akihiko Tsuji, Keiko Tominaga, Nami Nishiyama and Keizo Yuasa : Comprehensive enzymatic analysis of the cellulolytic system in digestive fluid of the sea hare aplysia kurodai. efficient glucose release from sea lettuce by synergistic action of 45 kDa endoglucanase and 210 kDa ß-glucosidase., PLoS ONE, Vol.8, No.6, 2013. (Summary) Although many endo-ß-1,4-glucanases have been isolated in invertebrates, their cellulolytic systems are not fully understood. In particular, gastropod feeding on seaweed is considered an excellent model system for production of bioethanol and renewable bioenergy from third-generation feedstocks (microalgae and seaweeds). In this study, enzymes involved in the conversion of cellulose and other polysaccharides to glucose in digestive fluids of the sea hare (Aplysia kurodai) were screened and characterized to determine how the sea hare obtains glucose from sea lettuce (Ulva pertusa). Four endo-ß-1,4-glucanases (21K, 45K, 65K, and 95K cellulase) and 2 ß-glucosidases (110K and 210K) were purified to a homogeneous state, and the synergistic action of these enzymes during cellulose digestion was analyzed. All cellulases exhibited cellulase and lichenase activities and showed distinct cleavage specificities against cellooligosaccharides and filter paper. Filter paper was digested to cellobiose, cellotriose, and cellotetraose by 21K cellulase, whereas 45K and 65K enzymes hydrolyzed the filter paper to cellobiose and glucose. 210K ß-glucosidase showed unique substrate specificity against synthetic and natural substrates, and 4-methylumbelliferyl (4MU)-ß-glucoside, 4MU-ß-galactoside, cello-oligosaccharides, laminarin, and lichenan were suitable substrates. Furthermore, 210K ß-glucosidase possesses lactase activity. Although ß-glucosidase and cellulase are necessary for efficient hydrolysis of carboxymethylcellulose to glucose, laminarin is hydrolyzed to glucose only by 210K ß-glucosidase. Kinetic analysis of the inhibition of 210K ß-glucosidase by D-glucono-1,5-lactone suggested the presence of 2 active sites similar to those of mammalian lactase-phlorizin hydrolase. Saccharification of sea lettuce was considerably stimulated by the synergistic action of 45K cellulase and 210K ß-glucosidase. Our results indicate that 45K cellulase and 210K ß-glucosidase are the core components of the sea hare digestive system for efficient production of glucose from sea lettuce. These findings contribute important new insights into the development of biofuel processing biotechnologies from seaweed. (Link to Publication Site) ● Publication site (DOI): 10.1371/journal.pone.0065418 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 23762366 ● Summary page in Scopus @ Elsevier: 2-s2.0-84878807011 (DOI: 10.1371/journal.pone.0065418, PubMed: 23762366, Elsevier: Scopus)
20.	Masahiro Inoue, Kouichi Yasuda, Haruki Uemura, Natsumi Yasaka, Achim Schnaufer, Mihiro Yano, Hiroshi Kido, Daisuke Kohda, Hirofumi Doi, Toshihide Fukuma, Akihiko Tsuji and Nobuo Horikoshi : Trypanosoma brucei 14-3-3I and II proteins predominantly form a heterodimer structure that acts as a potent cell cycle regulator in vivo., The Journal of Biochemistry, Vol.153, No.5, 431-439, 2013. (Summary) Hetero- and homodimerization of 14-3-3 proteins demonstrate distinctive functions in mammals and plants. Trypanosoma brucei 14-3-3I and II (Tb14-3-3I and II) play pivotal roles in motility, cytokinesis and the cell cycle; however, the significance and the mechanism of Tb14-3-3 dimerization are remained to be elucidated. We found that ectopically expressed epitope-tagged Tb14-3-3I and II proteins formed hetero- and homodimers with endogenous Tb14-3-3I and II proteins. However, we also found the ability to form hetero- or homodimers between Tb14-3-3I and II proteins was clearly affected by the sequence and location of the epitope tag used. We found a blue native polyacrylamide gel electrophoresis system followed by western blotting may distinguish monomer from dimer structure, and stable from unstable conformation of Tb14-3-3. Combined with co-immunoprecipitation results, we revealed that Tb14-3-3 proteins mainly existed as heterodimeric form. Furthermore, co-overexpression of Tb14-3-3I and II proteins in T. brucei induced aberrant numbers of organelles in cells, but overexpression of either isoform alone rarely produced such morphology. These results suggest that heterodimers play more significant roles than homodimers not only in the maintenance of steady-state levels of the 14-3-3 proteins but also in the regulation of cytokinesis. (Keyword) 14-3-3 Proteins / cell cycle / cytokinesis / Humans / Polymerase Chain Reaction / Protozoan Proteins / Trypanosoma brucei brucei (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvt016 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 23457405 ● Summary page in Scopus @ Elsevier: 2-s2.0-84877278778 (DOI: 10.1093/jb/mvt016, PubMed: 23457405, Elsevier: Scopus)
21.	Koichiro Hada, Kinuka Isshiki, Shinya Matsuda, Keizo Yuasa and Akihiko Tsuji : Engineering of α1-antitrypsin variants with improved specificity for the proprotein convertase furin using site-directed random mutagenesis, Protein Engineering, Design & Selection, Vol.26, No.2, 123-131, 2013. (Summary) Furin, PACE4, PC5/6 and PC7 are members of the subtilisin-like proprotein convertase (SPC) family. Although these enzymes are known to play critical roles in various physiological and pathological events including cell differentiation, tumor growth, virus replication and the activation of bacterial toxins, their distinct functions are yet to be fully delineated. α1-PDX is an engineered α1-antitrypsin variant carrying the RXXR consensus motif for furin within its reactive site loop. However, α1-PDX inhibits other SPCs in addition to furin. In this work, we prepared various rat α1-antitrypsin variants containing Arg at the P1 site within the reactive site loop, and examined their respective selectivity. The novel α1-antitrypsin variant AVNR (AVPM352/AVNR) was identified as a highly selective inhibitor of furin. This variant formed a sodium dodecyl sulfate- and heat-stable furin/α1-antitrypsin complex and inhibited furin activity ex vivo and in vitro. Other SPC members including PACE4, PC5/6 and PC7 were not inhibited by the AVNR variant. Furin-mediated maturation of bone morphogenetic protein-4 was completely inhibited by ectopic expression of the AVNR variant. The AVNR variant should prove to be a useful inhibitor in identifying the specific role of furin. (Keyword) Amino Acid Motifs / Amino Acid Substitution / Animals / Antithrombin III / Bone Morphogenetic Protein 4 / COS Cells / Catalytic Domain / Cercopithecus aethiops / Furin / HEK293 Cells / Humans / Mutagenesis, Site-Directed / Proprotein Convertase 5 / Proprotein Convertases / Protein Binding / Protein Precursors / Protein Processing, Post-Translational / Protein Stability / Proteolysis / Rats / Serine Endopeptidases / Substrate Specificity / alpha 1-Antitrypsin (Link to Publication Site) ● Publication site (DOI): 10.1093/protein/gzs091 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 23155057 ● Search Scopus @ Elsevier (PMID): 23155057 ● Search Scopus @ Elsevier (DOI): 10.1093/protein/gzs091 (DOI: 10.1093/protein/gzs091, PubMed: 23155057)
22.	Akihiko Tsuji, Kana Tsukamoto, Keiko Iwamoto, Yuka Ito and Keizo Yuasa : Enzymatic characterization of germination-specific cysteine protease-1 expressed transiently in cotyledons during the early phase of germination., The Journal of Biochemistry, Vol.153, No.1, 73-83, 2013. (Summary) Papain-like cysteine protease activity that shows a unique transient expression profile in cotyledons of daikon radish during germination was detected. The enzyme showed a distinct elution pattern on DEAE-cellulose compared with cathepsin B-like and Responsive to dessication-21 cysteine protease. Although this activity was not detected in seed prior to imbibition, the activity increased markedly and reached a maximum at 2 days after imbibition and then decreased rapidly and completely disappeared after 5 days. Using cystatin-Sepharose, the 26 kDa cysteine protease (DRCP26) was isolated from cotyledons at 2 days after imbibition. The deduced amino acid sequence from the cDNA nucleotide sequence indicated that DRCP26 is an orthologue of Arabidopsis unidentified protein, germination-specific cysteine protease-1, belonging to the C1 family of cysteine protease predicted from genetic information. In an effort to characterize the enzymatic properties of DRCP26, the enzyme was purified to homogeneity from cotyledons at 48 h after imbibition. The best synthetic substrate for the enzyme was carbobenzoxy-Phe-Arg-4-methylcoumaryl-7-amide. All model peptides were digested to small peptides by the enzyme, suggesting that DRCP26 possesses broad cleavage specificity. These results indicated that DRCP26 plays a role in the mobilization of storage proteins in the early phase of seed germination. (Keyword) Amino Acid Sequence / Catalytic Domain / Cotyledon / Coumarins / Cysteine / Cysteine Endopeptidases / Cystine / Dipeptides / Gene Expression Regulation, Plant / Germination / Indicators and Reagents / Japan / Molecular Sequence Data / molecular weight / Peptide Fragments / Plant Proteins / Proteolysis / Raphanus / Recombinant Proteins / Sequence Alignment / Sequence Homology, Amino Acid / Substrate Specificity (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvs125 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 23112094 ● Search Scopus @ Elsevier (PMID): 23112094 ● Search Scopus @ Elsevier (DOI): 10.1093/jb/mvs125 (DOI: 10.1093/jb/mvs125, PubMed: 23112094)
23.	Akihiko Tsuji, Shiori Sato, Ayumi Kondo, Keiko Tominaga and Keizo Yuasa : Purification and characterization of cellulase from North Pacific krill (Euphausia pacifica). Analysis of cleavage specificity of the enzyme., Comparative Biochemistry and Physiology. Part B, Biochemistry & Molecular Biology, Vol.163, No.3-4, 324-333, 2012. (Summary) Krill are filter feeders that consume algae, plankton and detritus, indicating that krill possess an adequate cellulose digesting system. However, less is known about the enzymatic properties of crustacean cellulases compared to termite cellulases. In the present study, 48 kDa-cellulase was highly purified from krill (Euphausia pacifica) in an effort to determine the cleavage specificity of the enzyme. The most notable characteristic of the enzyme was its high activity against both lichenan and carboxymethyl cellulose. The enzyme hydrolyzed internal β-1,4 glycosidic bonds within lichenan as well as carboxymethyl cellulose to release oligosaccharides and glucose. The effects of pH and temperature on the activity and stability of both enzyme activities were almost identical. Cello-oligosaccharides with a degree of polymerization of 4-6 were hydrolyzed by the enzyme, and the same endo-products, cellotriose, cellobiose and glucose, were produced from these oligosaccharides. Neither cellotriose nor cellobiose was hydrolyzed by the enzyme. The enzyme digested filter paper and sea lettuce to produce cellobiose, cellotriose and glucose as major products. Although amino acid sequence homology of the enzyme with termite cellulases and the presence of oligosaccharides in the enzyme suggested that the enzyme is produced by krill itself, further analysis is necessary. (Link to Publication Site) ● Publication site (DOI): 10.1016/j.cbpb.2012.08.005 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 22981467 ● Search Scopus @ Elsevier (PMID): 22981467 ● Search Scopus @ Elsevier (DOI): 10.1016/j.cbpb.2012.08.005 (DOI: 10.1016/j.cbpb.2012.08.005, PubMed: 22981467)
24.	Keizo Yuasa, Go Futamatsu, Tsuyoshi Kawano, Masaki Muroshita, Yoko Kageyama, Hiromi Taichi, Hiroshi Ishikawa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Subtilisin-like proprotein convertase paired basic amino acid-cleaving enzyme 4 is required for chondrogenic differentiation in ATDC5 cells, The FEBS Journal, Vol.279, No.21, 3997-4009, 2012. (Summary) Bone morphogenetic proteins (BMPs) have been implicated in the regulation of multiple stages of endochondral bone development. BMPs are synthesized as inactive precursors, and activated by removal of the propeptide. The subtilisin-like proprotein convertase (SPC) family comprises seven members [furin/SPC1, PC2/SPC2, PC1/PC3/SPC3, paired basic amino acid-cleaving enzyme 4 (PACE4)/SPC4, PC4/SPC5, PC6/PC5/SPC6, and PC8/PC7/LPC/SPC7], and activates various signaling molecules, including BMPs. In this study, we analyzed the role of this family in chondrogenic differentiation by using the mouse embryonal carcinoma-derived clonal cell line ATDC5. Both SPC-specific inhibitors, decanoyl-Arg-Val-Lys-Arg-chloromethylketone and α1-antitrypsin Portland variant, suppressed chondrogenic differentiation. RT-PCR analysis revealed that PACE4 mRNA levels increased markedly during chondrogenic differentiation, whereas furin expression remained unchanged. Knockdown of PACE4 expression significantly reduced chondrogenic differentiation. Furthermore, proBMP6, which shows an expression pattern similar to that of PACE4, was efficiently processed into its mature form by PACE4, whereas furin could not process proBMP6. These results suggest that PACE4 may regulate the rate of hypertrophic conversion of ATDC5 cells through activation of proBMP6. (Keyword) Amino Acids, Basic / Animals / Blotting, Western / Bone Morphogenetic Protein 6 / Carcinoma, Embryonal / Cell Differentiation / Chondrocytes / Furin / Humans / Mice / Proprotein Convertases / RNA, Messenger / RNA, Small Interfering / Rats / Real-Time Polymerase Chain Reaction / Reverse Transcriptase Polymerase Chain Reaction / Tumor Cells, Cultured / alpha 1-Antitrypsin (Link to Publication Site) ● Publication site (DOI): 10.1111/j.1742-4658.2012.08758.x (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 22925071 ● Search Scopus @ Elsevier (PMID): 22925071 ● Search Scopus @ Elsevier (DOI): 10.1111/j.1742-4658.2012.08758.x (DOI: 10.1111/j.1742-4658.2012.08758.x, PubMed: 22925071)
25.	Kimihiko Mizutani, Sae Tsuchiya, Mayuko Toyoda, Yuko Nanbu, Keiko Tominaga, Keizo Yuasa, Nobuyuki Takahashi, Akihiko Tsuji and Bunzo Mikamia : Structure of β-1,4-mannanase from the common sea hare Aplysia kurodai at 1.05 Å resolution., Acta Crystallographica. Section F, Structural Biology and Crystallization Communications, Vol.68, No.10, 1164-1168, 2012. (Summary) β-1,4-Mannanase (EC 3.2.1.78) catalyzes the hydrolysis of β-1,4-glycosidic bonds within mannan, a major constituent group of the hemicelluloses. Bivalves and gastropods possess β-1,4-mannanase and may degrade mannan in seaweed and/or phytoplankton to obtain carbon and energy using the secreted enzymes in their digestive systems. In the present study, the crystal structure of AkMan, a gastropod β-1,4-mannanase prepared from the common sea hare Aplysia kurodai, was determined at 1.05 Å resolution. This is the first report of the three-dimensional structure of a gastropod β-1,4-mannanase. The structure was compared with bivalve β-1,4-mannanase and the roles of residues in the catalytic cleft were investigated. No obvious binding residue was found in subsite +1 and the substrate-binding site was exposed to the molecular surface, which may account for the enzymatic properties of mannanases that can digest complex substrates such as glucomannan and branched mannan. (Link to Publication Site) ● Publication site (DOI): 10.1107/S1744309112037074 (Link to Search Site for Scientific Articles) ● CiNii @ National Institute of Informatics (CRID): 1050282810716626560 ● Search Scopus @ Elsevier (DOI): 10.1107/S1744309112037074 (DOI: 10.1107/S1744309112037074, CiNii: 1050282810716626560)
26.	Kinuka Isshiki, Shinya Matsuda, Akihiko Tsuji and Keizo Yuasa : cGMP-dependent protein kinase I promotes cell apoptosis through hyperactivation of death-associated protein kinase 2., Biochemical and Biophysical Research Communications, Vol.422, No.2, 280-284, 2012. (Summary) cGMP-dependent protein kinase-I (cGK-I) induces apoptosis in various cancer cell lines. However, the signaling mechanisms involved remain unknown. Using protein microarray technology, we identified a novel cGK substrate, death-associated protein kinase 2 (DAPK2), which is a Ca(2+)/calmodulin-regulated serine/threonine kinase. cGK-I phosphorylated DAPK2 at Ser(299), Ser(367) and Ser(368). Interestingly, a phospho-mimic mutant, DAPK2 S299D, significantly enhanced its kinase activity in the absence of Ca(2+)/calmodulin, while a S367D/S368D mutant did not. Overexpression of DAPK2 S299D also resulted in a twofold increase in apoptosis of human breast cancer MCF-7 cells as compared with wild-type DAPK2. These results suggest that DAPK2 is one of the targets of cGK-I in apoptosis induction. (Keyword) Animals / apoptosis / Apoptosis Regulatory Proteins / COS Cells / Calcium-Calmodulin-Dependent Protein Kinases / Cell Line, Tumor / Cercopithecus aethiops / Cyclic GMP-Dependent Protein Kinases / Enzyme Activation / Humans / Mice / Mutation / phosphorylation / Protein Array Analysis / Serine / Substrate Specificity (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2012.04.148 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 22580283 ● Search Scopus @ Elsevier (PMID): 22580283 ● Search Scopus @ Elsevier (DOI): 10.1016/j.bbrc.2012.04.148 (DOI: 10.1016/j.bbrc.2012.04.148, PubMed: 22580283)
27.	Keizo Yuasa, Takeshi Nagame, Makoto Dohi, Yayoi Yanagita, Shin Yamagami, Masami Nagahama and Akihiko Tsuji : cGMP-dependent protein kinase I is involved in neurite outgrowth via a Rho effector, rhotekin, in Neuro2A neuroblastoma cells., Biochemical and Biophysical Research Communications, Vol.421, No.2, 239-244, 2012. (Summary) Although the cGMP/cGMP-dependent protein kinase (cGK) signaling is involved in the regulation of neurite outgrowth, its mechanism remains to be clarified. In this study, we identified a Rho effector, rhotekin, as a cGK-I-interacting protein. Rhotekin was also a substrate for cGK-Iα. In neurite-extended Neuro2A neuroblastoma cells, cGK-Iα and rhotekin were colocalized in the plasma membrane and extended neurites, while treatment with cGMP resulted in translocation of rhotekin to the cytoplasm. In addition, we found that cGK-Iα and rhotekin synergistically suppressed Rho-induced neurite retraction. Our findings suggest that cGK-Iα interacts with and phosphorylates rhotekin, thereby contributing to neurite outgrowth regulation. (Keyword) Animals / Cell Line, Tumor / Cyclic GMP-Dependent Protein Kinases / Intracellular Signaling Peptides and Proteins / Mice / Neurites / phosphorylation / Serine (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2012.03.143 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 22503686 ● Search Scopus @ Elsevier (PMID): 22503686 ● Search Scopus @ Elsevier (DOI): 10.1016/j.bbrc.2012.03.143 (DOI: 10.1016/j.bbrc.2012.03.143, PubMed: 22503686)
28.	Keizo Yuasa, Kaori Tada, Genki Harita, Tomomi Fujimoto, Masao Tsukayama and Akihiko Tsuji : Sudachitin, a polymethoxyflavone from Citrus sudachi, suppresses lipopolysaccharide-induced inflammatory responses in mouse macrophage-like RAW264 cells., Bioscience, Biotechnology, and Biochemistry, Vol.76, No.3, 598-600, 2012. (Summary) Although some polymethoxyflavones possess several important biological properties, including neuroprotective, anticancer, and anti-inflammatory ones, sudachitin, a polymethoxyflavone from <I>Citrus sudachi</I>, has been little studied. In this study, we found that sudachitin inhibited nitric oxide production by suppressing the expression of inducible nitric oxide synthase in lipopolysaccharide-stimulated macrophages, indicating that sudachitin has an anti-inflammatory effect. (Keyword) sudachitin / polymethoxyflavone / anti-inflammation / nitric oxide (NO) (Link to Publication Site) ● Publication site (DOI): 10.1271/bbb.110800 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 22451408 ● CiNii @ National Institute of Informatics (CRID): 1390001206478414848 ● Search Scopus @ Elsevier (PMID): 22451408 ● Search Scopus @ Elsevier (DOI): 10.1271/bbb.110800 (DOI: 10.1271/bbb.110800, PubMed: 22451408, CiNii: 1390001206478414848)
29.	Akihiko Tsuji, Yoshinori Fujisawa, Takeru Mino and Keizo Yuasa : Identification of a plant aminopeptidase with preference for aromatic amino acid residues as a novel member of the prolyl oligopeptidase family of serine proteases., The Journal of Biochemistry, Vol.150, No.5, 525-534, 2011. (Summary) Genome analysis has indicated that plants, like animals, possess a variety of protease genes. However, bulk of putative proteases has not been characterized at the enzyme level. In this article, a novel enzyme that hydrolyses phenylalanyl-4-methylcoumaryl 7-amide (phenylalanyl-MCA) was purified from cotyledons of daikon radish by ammonium sulphate fractionation and successive chromatography with DEAE-cellulose, phenyl-Sepharose, Sephacryl S-200 and Mini-Q. The molecular mass of the enzyme was estimated to be 78 kDa by SDS-PAGE under reducing conditions and 74 kDa by gel filtration, indicating that the enzyme is a monomer. The deduced amino acid sequence from the cDNA nucleotide sequence indicated that the enzyme is an orthologue of Arabidopsis unidentified protein, acylpeptide hydrolase-like protein (AHLP; UniProt ID: Q9FG66) belonging to the prolyl oligopeptidase (POP) family of a serine-type peptidase predicted from genetic information. Good substrates identified for the enzyme include phenylalanyl-MCA, tyrosyl-MCA and enkephalin. Neither acylamino acid-releasing activity nor endopeptidase activity was detected. The enzyme cleaved enkephalin (YGGFM, YGGFL), whereas, BAM-12 P (YGGFMRRVGRPE) and dynorphin A (YGGFLRRIRPKLK) were not digested. These results suggested that the enzyme possesses strict size selectivity of substrate. We propose the name 'tyrosyl aminopeptidase' for the uncharacterized protein AHLP. (Keyword) Amino Acids, Aromatic / Aminopeptidases / Cotyledon / Electrophoresis, Polyacrylamide Gel / Plant Proteins / Raphanus / Serine Endopeptidases / Serine Proteases / Substrate Specificity (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvr092 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 21788307 ● Search Scopus @ Elsevier (PMID): 21788307 ● Search Scopus @ Elsevier (DOI): 10.1093/jb/mvr092 (DOI: 10.1093/jb/mvr092, PubMed: 21788307)
30.	Keizo Yuasa, Taito Matsuda and Akihiko Tsuji : Functional regulation of transient receptor potential canonical 7 by cGMP-dependent protein kinase Iα., Cellular Signalling, Vol.23, No.7, 1179-1187, 2011. (Summary) The cGMP/cGMP-dependent protein kinase (cGK) signaling pathway is implicated in the functional regulation of intracellular calcium levels. In the present study, we investigated the regulation of transient receptor potential canonical 7 (TRPC7) by the cGMP/cGK-I pathway. TRPC7 contains three putative cGK phosphorylation sites (Arg-Arg/Lys-Xaa-Ser/Thr). However, the role of cGK-I in the regulation of TRPC7 activity remains unclear. In vitro and in vivo kinase assays have revealed that cGK-Iα phosphorylates mouse TRPC7 but not mouse TRPC3. Site-directed mutagenesis analysis revealed that TRPC7 was phosphorylated by cGK-Iα at threonine 15. Phosphorylation of TRPC7 significantly suppressed carbachol-induced calcium influx and CREB phosphorylation. Furthermore, co-immunoprecipitation assay demonstrated that cGK-Iα interacted with the ankyrin repeat domain in the N terminus of TRPC7. cGK-Iβ also bound to TRPC7, while the type II regulatory subunit of cAMP-dependent protein kinase did not bind. These data indicate that cGK-Iα interacts with and phosphorylates TRPC7, contributing to the quick and accurate regulation of calcium influx and CREB phosphorylation. (Keyword) Amino Acid Sequence / Animals / COS Cells / Calcium Signaling / Carbachol / Cercopithecus aethiops / Cyclic AMP Response Element-Binding Protein / Cyclic GMP-Dependent Protein Kinases / Enzyme Assays / HEK293 Cells / Humans / Isoenzymes / Molecular Sequence Data / Mutagenesis, Site-Directed / Phosphorylation / Protein Binding / Recombinant Fusion Proteins / TRPC Cation Channels (Link to Publication Site) ● Publication site (DOI): 10.1016/j.cellsig.2011.03.005 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 21402151 ● Search Scopus @ Elsevier (PMID): 21402151 ● Search Scopus @ Elsevier (DOI): 10.1016/j.cellsig.2011.03.005 (DOI: 10.1016/j.cellsig.2011.03.005, PubMed: 21402151)
31.	Ismail Ismaliza Mohd Nor, Tsuyoshi Yuasa, Keizo Yuasa, Yuko Nambu, Makoto Nishimoto, Masaki GOTO, Hitoshi Matsuki, Masahiro Inoue, Masami Nagahama and Akihiko Tsuji : A critical role for highly conserved Glu⁶¹⁰ residue of oligopeptidase B Tryoanosoma brucei in thermal stability, The Journal of Biochemistry, Vol.147, No.2, 201-211, 2010. (Summary) Oligopeptidase B from Trypanosoma brucei (Tb OPB) is a virulence factor and therapeutic target in African sleeping sickness. Three glutamic acid residues at positions 607, 609 and 610 of the catalytic domain are highly conserved in the OPB subfamily. In this study, the roles of Glu(607), Glu(609) and Glu(610) in Tb OPB were investigated by site-directed mutagenesis. A striking effect on k(cat)/K(m) was obtained following mutation of Glu(607) to glutamine. In contrast, the heat stability of Tb OPB decreased markedly following the single mutation of Glu(610) to glutamine, although this mutation had significantly less effect on catalytic properties compared with the Glu(607) mutation. Although no differences were found in the tertiary and secondary structures between wild-type (WT) OPB and the E610Q mutant prior to heat treatment, the E610Q mutant is inactivated more rapidly than WT OPB following heat treatment in a manner correlating with its attendant structural changes. Trypsin digestion showed that the boundary regions between the beta-propeller and catalytic domain of the E610Q mutant are unfolded with heat treatment. It is concluded that Glu(607) is essential for the catalytic activity of Tb OPB and that Glu(610) plays a critical role in stabilization rather than catalytic activity despite their close proximity. (Keyword) Adrenocorticotropic Hormone / Animals / Calorimetry, Differential Scanning / Circular Dichroism / Enzyme Activation / Enzyme Stability / Glucagon / Glutamic Acid / Hot Temperature / Mutagenesis, Site-Directed / Mutation / Protozoan Proteins / Salts / Serine Endopeptidases / Trypanosoma brucei brucei / Trypsin / Urea (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvp156 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 19819899 ● Search Scopus @ Elsevier (PMID): 19819899 ● Search Scopus @ Elsevier (DOI): 10.1093/jb/mvp156 (DOI: 10.1093/jb/mvp156, PubMed: 19819899)
32.	Keizo Yuasa, Shotaro Uehara, Masami Nagahama and Akihiko Tsuji : Transcriptional regulation of cGMP-dependent protein kinase II (cGK-II) in chondrocytes., Bioscience, Biotechnology, and Biochemistry, Vol.74, No.1, 44-49, 2010. (Summary) The C-type natriuretic peptide/natriuretic peptide receptor-B/cGMP pathway plays an important role in the regulation of endochondral ossification. In chondrocytes, the physiological effect of cGMP is mediated primarily by the activation of cGMP-dependent protein kinase II (cGK-II). In this study, we investigated the transcriptional regulation of cGK-II in chondrocytes. The expression pattern of cGK-II transcripts was examined during chondrogenic differentiation of ATDC5 cells. cGK-II mRNA was not detectable in undifferentiated cells, but increased dramatically prior to differentiation to the hypertrophic stage. To analyze the transcriptional regulation of cGK-II, the 5'-flanking region of the mouse cGK-II gene was isolated and characterized. The promoter activity of the cGK-II gene decreased markedly following deletion and mutagenesis of the putative Nkx-binding site between nucleotide positions -292 and -286. These results suggest that the homeobox gene Nkx family is critical for the transcriptional regulation of cGK-II during chondrogenesis. (Keyword) Animals / Base Sequence / Cartilage / Cell Differentiation / Cell Line, Tumor / Chondrocytes / Chondrogenesis / Cyclic GMP-Dependent Protein Kinases / Gene Expression Regulation, Enzymologic / Mice / Molecular Sequence Data / Promoter Regions, Genetic / Rats / Transcription, Genetic (Link to Publication Site) ● Publication site (DOI): 10.1271/bbb.90529 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 20057151 ● CiNii @ National Institute of Informatics (CRID): 1390282681455121792 ● Search Scopus @ Elsevier (PMID): 20057151 ● Search Scopus @ Elsevier (DOI): 10.1271/bbb.90529 (DOI: 10.1271/bbb.90529, PubMed: 20057151, CiNii: 1390282681455121792)
33.	Tomoko Takahashi, Takanori Ida, Takahiro Sato, Yoshiki Nakashima, Yuki Nakamura, Akihiko Tsuji and Masayasu Kojima : Production of n-octanoyl-modified Ghrelin in Cultured Cells Requires Prohormone Proceeding Protease and Ghrelin O-acyltransferase, as well as n-octanoic Acid, The Journal of Biochemistry, Vol.146, No.5, 675-682, 2009. (Summary) Ghrelin was originally isolated from rat stomach as an endogenous ligand for the GH secretagogue receptor. The major active form of ghrelin is a 28-amino acid peptide modified by an n-octanoic acid on the serine 3 residue, and this lipid modification is essential for the biological activity of ghrelin. However, it is not clear whether prohormone convertase (PC) and ghrelin O-acyltransferase (GOAT) are the minimal requirements for synthesis of acyl-modified ghrelin in cultured cells. By using three cultured cell lines, TT, AtT20 and COS-7, in which the expression levels of processing proteases and GOAT vary, we examined the processing patterns of ghrelin precursor. We found that not only PC1/3 but also both PC2 and furin could process proghrelin to the 28-amino acid ghrelin. Moreover, the presence of PC and GOAT in the cells, as well as n-octanoic acid in the culture medium, was necessary to produce n-octanoyl ghrelin. (Keyword) Acyltransferases / Animals / Cells, Cultured / Chromatography, High Pressure Liquid / Fasting / Furin / Gene Expression Regulation / Genetic Vectors / Ghrelin / Humans / Male / Mice / Octanoic Acids / Proprotein Convertases / RNA, Messenger / Rats / Rats, Wistar / Stomach / Transfection (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvp112 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 19628676 ● Search Scopus @ Elsevier (PMID): 19628676 ● Search Scopus @ Elsevier (DOI): 10.1093/jb/mvp112 (DOI: 10.1093/jb/mvp112, PubMed: 19628676)
34.	Keizo Yuasa, Tetsuya Masuda, Chihiro Yoshikawa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Subtilisin-like Proprotein Convertase PACE4 is Required for Skeletal Muscle Differentiation, The Journal of Biochemistry, Vol.146, No.3, 407-415, 2009. (Summary) Most growth factors stimulate myoblast proliferation and prevent differentiation, whereas insulin-like growth factors (IGFs) promote myoblast differentiation through the phosphatidylinositol 3-kinase (PI3K) pathway. Subtilisin-like proprotein convertases (SPCs) are involved in cell growth and differentiation via activation of pro-growth factors. However, the role of SPCs in myogenesis remains poorly understood. Here we show that PACE4, a member of the SPC family, plays a critical role in myogenic differentiation of C2C12 cells. PACE4 mRNA levels increased markedly during myogenesis, whereas the expression of other member of SPC family, furin and PC6, remained unchanged. The expression pattern of pro-IGF-II, which is processed extracellularly by SPCs, was similar to that of PACE4. The expression of shRNA targeting PACE4, but not furin, suppressed the expression of the muscle-specific myosin light chain (MLC). Interestingly, reduced expression of MLC was restored following treatment with recombinant mature IGF-II. Finally, we demonstrated that the PI3K inhibitor LY294002 blocked the induction of PACE4 mRNA, a result not observed when another myogenic differentiation inhibitor, SB203580 (p38 MAP kinase inhibitor), was employed, indicating the presence of a positive feedback loop regulating PACE4 expression. These results suggest that PACE4 plays an important role in myogenic differentiation through its association with the IGF-II pathway. (Keyword) Animals / Cell Differentiation / Cell Line / Feedback, Physiological / Gene Knockdown Techniques / Insulin-Like Growth Factor II / Matrix Metalloproteinase 11 / Mice / Muscle Development / MyoD Protein / Myoblasts, Skeletal / Myogenin / Myosin Light Chains / Phosphatidylinositol 3-Kinases / Proprotein Convertases / Protein Precursors / Signal Transduction / Up-Regulation / p38 Mitogen-Activated Protein Kinases (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvp090 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 19520771 ● Search Scopus @ Elsevier (PMID): 19520771 ● Search Scopus @ Elsevier (DOI): 10.1093/jb/mvp090 (DOI: 10.1093/jb/mvp090, PubMed: 19520771)
35.	Masami Nagahama, Machi Ohnishi, Yumiko Kawate, Takayuki Matsui, Hitomi Miyake, Keizo Yuasa, Katsuko Tani, Mitsuo Tagaya and Akihiko Tsuji : UBXD1 is a VCP-interacting protein that is involved in ER-associated degradation, Biochemical and Biophysical Research Communications, Vol.382, No.2, 303-308, 2009. (Summary) AAA ATPase VCP and its yeast ortholog Cdc48, in a complex with the Ufd1-Npl4 heterodimer as an adaptor, play an essential role in endoplasmic reticulum-associated degradation (ERAD). Several UBX domain-containing proteins function to recruit ubiquitylated substrates to VCP/Cdc48 by binding both VCP/Cdc48 and other ERAD components such as ubiquitin ligases. Here we show that mammalian UBXD1 is an additional UBX domain-containing protein involved in the ERAD process. UBXD1 is a cytosolic protein that interacts with VCP and Derlin-1. Overexpression of UBXD1 in cells causes selective dissociation of Ufd1 from VCP, resulting in inhibition of mutant cystic fibrosis transmembrane conductance regulator (CFTR) degradation by ERAD. Additionally, depletion of endogenous UBXD1 protein by RNA interference also results in a defect in CFTR degradation. Collectively, these findings suggest that UBXD1 is a regulatory component of ERAD that may modulate the adaptor binding to VCP. (Keyword) Adenosine Triphosphatases / Carrier Proteins / Cell Cycle Proteins / Endoplasmic Reticulum / HeLa Cells / Humans / Membrane Proteins / Protein Structure, Tertiary (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2009.03.012 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 19275885 ● Search Scopus @ Elsevier (PMID): 19275885 ● Search Scopus @ Elsevier (DOI): 10.1016/j.bbrc.2009.03.012 (DOI: 10.1016/j.bbrc.2009.03.012, PubMed: 19275885)
36.	Takuya Yanagino, Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Transcriptional Regulation of Fibrillin-2 Gene by E2F Family Members in Chondrocyte Differentiation, Journal of Cellular Biochemistry, Vol.106, No.4, 580-588, 2009. (Summary) Mutation in fibrillin-2, a major structural component of extracellular microfibrils in connective tissue, results in the autosomal dominant disease congenital contractural arachnodactyly. This genetic disease is characterized by dolichostenomelia and arachnodactyly, in addition to contractures of the large joints and abnormal pinnae formation, thus indicating the significance of fibrillin-2 in chondrogenesis. In this study, we investigated the transcriptional regulation of fibrillin-2 in chondrogenic differentiation. Although mRNA expression of fibrillin-1, a highly homologous protein to fibrillin-2, remained almost unchanged during chondrogenesis of mouse ATDC5 cells, fibrillin-2 mRNA expression varied. Fibrillin-2 was highly expressed at the early stage and declined progressively during differentiation. The 5'-flanking region of the fibrillin-2 gene contains potential binding sites for E2F, Runx, AP-2, and Sox transcription factors. The promoter activity of fibrillin-2 decreased markedly following deletion and mutagenesis of the E2F binding site between -143 and -136 bp. Overexpression of E2F1 resulted in a marked increase in its promoter activity, whereas expression of other transcription factors including AP-2alpha and Runx2 had no effect. The increase in promoter activity by E2F1 was completely suppressed by the coexpression of E2F4. E2F2 and E2F3 had positive effects on the promoter activity. Although ATDC5 cells expressed transcripts for the E2F family genes at all stages of differentiation, the expression profiles differed. E2F1 expression remained almost unchanged, whereas E2F4 expression increased markedly at the late stage of differentiation. These results indicated that coordinated expression of the E2F family is critical for the transcriptional regulation of fibrillin-2 during chondrogenesis. (Keyword) Animals / Cell Differentiation / Cell Line / Chondrocytes / Chondrogenesis / E2F Transcription Factors / E2F1 Transcription Factor / E2F2 Transcription Factor / E2F3 Transcription Factor / E2F4 Transcription Factor / Gene Expression Regulation / Mice / Microfilament Proteins / RNA, Messenger / Transcription, Genetic (Link to Publication Site) ● Publication site (DOI): 10.1002/jcb.22029 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 19142862 ● Search Scopus @ Elsevier (PMID): 19142862 ● Search Scopus @ Elsevier (DOI): 10.1002/jcb.22029 (DOI: 10.1002/jcb.22029, PubMed: 19142862)
37.	Akihiko Tsuji, Yayoi Kikuchi, Kentaro Ogawa, Hiroko Saika, Keizo Yuasa and Masami Nagahama : Purification and characterization of cathepsin B-like cysteine protease from cotyledons of daikon radish, Raphanus sativus, The FEBS Journal, Vol.275, No.21, 5429-5443, 2008. (Summary) Plant cathepsin B-like cysteine protease (CBCP) plays a role in disease resistance and in protein remobilization during germination. The ability of animal cathepsin B to function as a dipeptidyl carboxypeptidase has been attributed to the presence of a dihistidine (His110-His111) motif in the occluding loop, which represents a unique structure of cathepsin B. However, a dihistidine motif is not present in the predicted sequence of the occluding loop of plant CBCP, as determined from cDNA sequence analysis, and the loop is shorter. In an effort to investigate the enzymatic properties of plant CBCP, which possesses the unusual occluding loop, we have purified CBCP from the cotyledons of daikon radish (Raphanus sativus) by chromatography through Sephacryl S-200, DEAE-cellulose, hydroxyapatite and organomercurial-Sepharose. The molecular mass of the enzyme was estimated to be 28 kDa by SDS/PAGE under reducing conditions. The best synthetic substrate for CBCP was t-butyloxycarbonyl Leu-Arg-Arg-4-methylcoumaryl 7-amide, as is the case with human cathepsin B. However, the endopeptidase activity of CBCP towards glucagon and adrenocorticotropic hormone showed broad cleavage specificity. Human cathepsin B preferentially cleaves model peptides via its dipeptidyl carboxypeptidase activity, whereas daikon CBCP displays both endopeptidase and exopeptidase activities. In addition, CBCP was found to display carboxymonopeptidase activity against the substrate o-aminobenzoyl-Phe-Arg-Phe(4-NO(2)). Daikon CBCP is less sensitive (1/7000) to CA-074 than human cathepsin B. Expression analysis of CBCP at the protein and RNA levels indicated that daikon CBCP activity in cotyledons is regulated by post-transcriptional events during germination. (Keyword) Cathepsin B / Chromatography / Cotyledon / Cysteine Endopeptidases / Gene Expression Regulation, Plant / Germination / Humans / Molecular Weight / Plant Proteins / RNA, Messenger / Raphanus / Substrate Specificity (Link to Publication Site) ● Publication site (DOI): 10.1111/j.1742-4658.2008.06674.x (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 18959767 ● Search Scopus @ Elsevier (PMID): 18959767 ● Search Scopus @ Elsevier (DOI): 10.1111/j.1742-4658.2008.06674.x (DOI: 10.1111/j.1742-4658.2008.06674.x, PubMed: 18959767)
38.	Yayoi Kikuchi, Hiroko Saika, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : Isolation and Biochemical Characterization of Two Forms of RD21 from Cotyledons of Daikon Radish (Raphanus sativus)., The Journal of Biochemistry, Vol.144, No.6, 789-798, 2008. (Summary) RD21 (Responsive to desiccation-21) is an Arabidopsis cysteine protease which possesses a granulin-like domain at the C-terminus. Although two forms of RD21 have been identified, consisting of an intermediate form (iRD21) containing a granulin domain and a mature form (mRD21) lacking this domain, the enzymatic properties of these enzymes remain poorly understood. In this study, mRD21 orthologue was purified to homogeneity from the cotyledons of daikon radish (Raphanus sativus). RD21 preferentially cleaved peptide bond that had an aromatic or hydrophobic amino acid at the P2 position. Furthermore, the presence of a polar amino acid at the P1 position enhanced the cleavage susceptibility of the peptide bond, although the importance of the type of amino acid residue at the P1 and P1' positions was not as significant as the residue located at the P2 position. The iRD21 was also identified as an oligomeric form by gel filtration and sedimentation analyses. The expression of RD21 mRNA was initiated by imbibition and continued at almost constant levels during germination. On the other hand, the enzyme activity increased markedly 5 days after imbibition. These results indicate that this elevation of RD21 activity is generated post-transcriptionally. (Keyword) Amino Acid Sequence / Angiotensin II / Base Sequence / Chromatography, Gel / Cotyledon / Cysteine Endopeptidases / Dynorphins / Hydrolysis / Molecular Sequence Data / Plant Proteins / Protein Isoforms / Raphanus / Substrate Specificity (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvn132 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 18838434 ● Search Scopus @ Elsevier (PMID): 18838434 ● Search Scopus @ Elsevier (DOI): 10.1093/jb/mvn132 (DOI: 10.1093/jb/mvn132, PubMed: 18838434)
39.	Keizo Yuasa, Shin Yamagami, Masami Nagahama and Akihiko Tsuji : Trafficking of cGMP-dependent protein kinase II via interaction with Rab11, Biochemical and Biophysical Research Communications, Vol.374, No.3, 522-526, 2008. (Summary) cGMP-dependent protein kinase II (cGK-II) is implicated in several physiological functions including intestinal secretion, bone growth, and learning and memory, but the detailed mechanisms are still unclear. To identify proteins that are involved in cGMP/cGK-II signaling, we performed yeast two-hybrid screening and identified Rab11b as a cGK-II-interacting protein that regulates the slow-recycling pathway. Interestingly, cGK-II interacted with the GDP-bound form of Rab11b (Rab11b S25N), but not the GTP-bound form, in mammalian cells. Immunofluorescence staining revealed that Rab11b S25N promoted the translocation of cGK-II from the plasma membrane to the cytoplasm and that the localization of cGK-II extensively overlapped with Rab11b. Furthermore, treatment with a membrane-permeable cGMP analog caused the rapid retranslocation of cGK-II and Rab11b S25N to the membrane. These data indicate that Rab11b is necessary for the trafficking of cGK-II and that the cGMP/cGK-II signaling pathway is closely related to Rab11b recycling pathway. (Keyword) Animals / Cell Line / Cell Membrane / Cyclic GMP-Dependent Protein Kinases / Guanine Nucleotides / Humans / Mice / Protein Transport / Signal Transduction / Two-Hybrid System Techniques / rab GTP-Binding Proteins (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2008.07.071 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 18656450 ● Search Scopus @ Elsevier (PMID): 18656450 ● Search Scopus @ Elsevier (DOI): 10.1016/j.bbrc.2008.07.071 (DOI: 10.1016/j.bbrc.2008.07.071, PubMed: 18656450)
40.	Youichi Tajima, Fumiko Matsuzawa, Sei-ichi Aikawa, Toshika Okumiya, Michiru Yoshimizu, Takahiro Tsukimura, Masahiko Ikekita, Seiichi Tsujino, Akihiko Tsuji, Tim Edmunds and Hitoshi Sakuraba : Structural and biochemical studies on Pompe disease and a "pseudodeficiency of acid α-glucosidase", Journal of Human Genetics, Vol.52, No.11, 898-906, 2007. (Summary) We constructed structural models of the catalytic domain and the surrounding region of human wild-type acid alpha-glucosidase and the enzyme with amino acid substitutions by means of homology modeling, and examined whether the amino acid replacements caused structural and biochemical changes in the enzyme proteins. Missense mutations including p.R600C, p.S619R and p.R437C are predicted to cause apparent structural changes. Nonsense mutation of p.C103X terminates the translation of acid alpha-glucosidase halfway through its biosynthesis and is deduced not to allow formation of the active site pocket. The mutant proteins resulting from these missense and nonsense mutations found in patients with Pompe disease are predictably unstable and degraded quickly in cells. The structural change caused by p.G576S is predicted to be small, and cells from a subject homozygous for this amino acid substitution exhibited 15 and 11% of the normal enzyme activity levels for an artificial substrate and glycogen, respectively, and corresponding amounts of the enzyme protein on Western blotting. No accumulation of glycogen was found in organs including skeletal muscle in the subject, and thus the residual enzyme activity could protect cells from glycogen storage. On the other hand, p.E689K, which is known as a neutral polymorphism, little affected the three-dimensional structure of acid alpha-glucosidase. Structural study on a mutant acid alpha-glucosidase in silico combined with biochemical investigation is useful for understanding the molecular pathology of Pompe disease. (Keyword) Amino Acid Sequence / Blotting, Western / Cells, Cultured / Glycogen Storage Disease Type II / Humans / Molecular Sequence Data / Sequence Homology, Amino Acid / alpha-Glucosidases (Link to Publication Site) ● Publication site (DOI): 10.1007/s10038-007-0191-9 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 17805474 ● CiNii @ National Institute of Informatics (CRID): 1521980703408428928 ● Search Scopus @ Elsevier (PMID): 17805474 ● Search Scopus @ Elsevier (DOI): 10.1007/s10038-007-0191-9 (DOI: 10.1007/s10038-007-0191-9, PubMed: 17805474, CiNii: 1521980703408428928)
41.	Keizo Yuasa, Kaori Suzue, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Transcriptional regulation of subtilisin-like proprotein convertase PACE4 by E2F: Possible role of E2F-mediated upregulation of PACE4 in tumor progression, Gene, Vol.402, No.1-2, 103-110, 2007. (Summary) PACE4, a member of the subtilisin-like proprotein convertase (SPC) family, is expressed at high levels in certain tumor cells and plays a role in metastatic progression through activation of matrix metalloproteinases. The mechanism leading to overexpression of PACE4 in tumor cells remains unclear. In this study, we show that the E2F1 transcription factor, which is implicated in carcinoma invasiveness, upregulates the expression of PACE4. HT1080 (highly tumorigenic and invasive) cells expressed much higher levels of PACE4 and E2F family (E2F1 and E2F2) transcripts than IMR90 (normal fibroblast) cells. Expression levels of other SPCs (furin and PC6) remained unchanged in these cells. Promoter analysis indicated that two E2F consensus binding sites (-117/-110 and -86/-79) in the 5'-flanking region of the human PACE4 gene function as positive regulatory elements. Mutation of these sites abolished PACE4 promoter response to E2F1 as well as binding of E2F1 in electrophoretic mobility-shift assays. Other E2F members, E2F2 and E2F3, also activated PACE4 expression, as in the case of E2F1. These results indicate a novel mechanism for E2F family-mediated promotion of carcinoma invasiveness through PACE4. (Keyword) Binding Sites / Cell Line, Tumor / Disease Progression / E2F Transcription Factors / E2F1 Transcription Factor / Electrophoretic Mobility Shift Assay / Gene Expression Regulation, Neoplastic / Humans / Neoplasms / Promoter Regions, Genetic / Proprotein Convertases / Serine Endopeptidases / Transcription, Genetic / Transfection / Up-Regulation (Link to Publication Site) ● Publication site (DOI): 10.1016/j.gene.2007.07.028 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 17825503 ● Search Scopus @ Elsevier (PMID): 17825503 ● Search Scopus @ Elsevier (DOI): 10.1016/j.gene.2007.07.028 (DOI: 10.1016/j.gene.2007.07.028, PubMed: 17825503)
42.	Akihiko Tsuji, Hiroki Kanie, Hirotaka Makise, Keizo Yuasa, Masami Nagahama and Yoshiko Matsuda : Engineering of α1-antitrypsin cariants selective for subtilisin-like proprotein convertases PACE4 and PC6: Importance of the P2' residue in stable complex formation of the serpin with proprotein convertase, Protein Engineering, Design & Selection, Vol.20, No.4, 163-170, 2007. (Summary) Furin and PACE4, members of the subtilisin-like proprotein convertase (SPC) family, have been implicated in the metastatic progression of certain tumors in addition to the activation of viral coat proteins and bacterial toxins, indicating that these enzymes are potential targets for therapeutic agents. Alpha1-Antitrypsin Portland is an engineered alpha1-antitrypsin designed as a furin-specific inhibitor and has been used as a tool in the functional analysis of furin. In this work, we engineered rat alpha1-antitrypsin to create a PACE4-specific inhibitor. Substituting Arg-Arg-Arg-Arg for Ala-Val-Pro-Met(352) at P4-P1 and Ala for Leu(354) at P2' created a potent PACE4- and PC6-specific inhibitor. This variant (RRRRSA) formed an SDS- and heat-stable serpin/proteinase complex with PACE4 or PC6 and inhibited both enzyme activities. The RRRRSA variant was efficiently cleaved by furin without formation of the stable complex. This is the first report of a highly selective protein-based inhibitor of PACE4 and PC6. This inhibitor will be useful in delineating the roles of PACE4 and PC6 localized in the extracellular matrix. (Keyword) Amino Acid Sequence / Amino Acid Substitution / Animals / Binding Sites / Proprotein Convertase 5 / Proprotein Convertases / Protein Engineering / Rats / Serpins / alpha 1-Antitrypsin (Link to Publication Site) ● Publication site (DOI): 10.1093/protein/gzm007 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 17351018 ● Search Scopus @ Elsevier (PMID): 17351018 ● Search Scopus @ Elsevier (DOI): 10.1093/protein/gzm007 (DOI: 10.1093/protein/gzm007, PubMed: 17351018)
43.	Masami Nagahama, Takeshi Yamazoe, Yoshimitsu Hara, Katsuko Tani, Akihiko Tsuji and Mitsuo Tagaya : The AAA-ATPase NVL2 is a component of pre-ribosomal particles that interacts with the DexD/H-box RNA helicase DOB1, Biochemical and Biophysical Research Communications, Vol.346, No.3, 1075-1082, 2006. (Summary) Nuclear VCP/p97-like protein 2 (NVL2) is a member of the chaperone-like AAA-ATPase family with two conserved ATP-binding modules. Our previous studies have shown that NVL2 is localized to the nucleolus by interacting with ribosomal protein L5 and may participate in ribosome synthesis, a process involving various non-ribosomal factors including chaperones and RNA helicases. Here, we show that NVL2 is associated with pre-ribosomal particles in the nucleus. Moreover, we used yeast two-hybrid and co-immunoprecipitation assays to identify an NVL2-interacting protein that could yield insights into NVL2 function in ribosome biogenesis. We found that NVL2 interacts with DOB1, a DExD/H-box RNA helicase, whose yeast homologue functions in a late stage of the 60S subunit synthesis. DOB1 can interact with a second ATP-binding module mutant of NVL2, which shows a dominant negative effect on ribosome synthesis. In contrast, it cannot interact with a first ATP-binding module mutant, which does not show the dominant negative effect. When the dominant negative mutant of NVL2 was overexpressed in cells, DOB1 appeared to remain associated with nuclear pre-ribosomal particles. Such accumulation was not observed upon overexpression of wild-type NVL2 or a nondominant-negative mutant. Taken together, our results suggest that NVL2 might regulate the association/dissociation reaction of DOB1 with pre-ribosomal particles by acting as a molecular chaperone. (Keyword) Adenosine Triphosphatases / Cell Line / Cell Nucleus / Humans / Mutation / Protein Binding / RNA Helicases / Ribosomes (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbrc.2006.06.017 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 16782053 ● Search Scopus @ Elsevier (PMID): 16782053 ● Search Scopus @ Elsevier (DOI): 10.1016/j.bbrc.2006.06.017 (DOI: 10.1016/j.bbrc.2006.06.017, PubMed: 16782053)
44.	Akihiko Tsuji, Yayoi Kikuchi, Yukimi Sato, Shizuyo Koide, Keizo Yuasa, Masami Nagahama and Yoshiko Matsuda : A proteomic approach reveals transient association of reticulocalbin-3, a novel member of the CREC family, with the precursor of subtilisin-like proprotein convertase, PACE4, The Biochemical Journal, Vol.396, No.1, 51-59, 2006. (Summary) SPCs (subtilisin-like proprotein convertases) are a family of seven structurally related serine endoproteases that are involved in the proteolytic activation of proproteins. In an effort to examine the substrate protein for PACE4 (paired basic amino-acid-cleaving enzyme-4), an SPC, a potent protein inhibitor of PACE4, an alpha1-antitrypsin RVRR (Arg-Val-Arg-Arg) variant, was expressed in GH4C1 cells. Ectopic expression of the RVRR variant caused accumulation of the 48 kDa protein in cells. Sequence analysis indicates that the 48 kDa protein is a putative Ca2+-binding protein, RCN-3 (reticulocalbin-3), which had previously been predicted by bioinformatic analysis of cDNA from the human hypothalamus. RCN-3 is a member of the CREC (Cab45/reticulocalbin/ERC45/calumenin) family of multiple EF-hand Ca2+-binding proteins localized to the secretory pathway. The most interesting feature of the RCN-3 sequence is the presence of five Arg-Xaa-Xaa-Arg motifs, which represents the target sequence of SPCs. Biosynthetic studies showed that RCN-3 is transiently associated with proPACE4, but not with mature PACE4. Inhibition of PACE4 maturation by a Ca2+ ionophore resulted in accumulation of the proPACE4-RCN-3 complex in cells. Furthermore, autoactivation and secretion of PACE4 was increased upon co-expression with RCN-3. Our findings suggest that selective and transient association of RCN-3 with the precursor of PACE4 plays an important role in the biosynthesis of PACE4. (Keyword) Adenoma / Amino Acid Sequence / Animals / COS Cells / Calcium-Binding Proteins / Cell Line / Cell Line, Tumor / Cercopithecus aethiops / Enzyme Activation / Furin / Humans / Kidney / Molecular Sequence Data / Neoplasm Proteins / Pituitary Neoplasms / Proprotein Convertase 5 / Proprotein Convertases / Protein Binding / Protein Precursors / Proteomics / Rats / Recombinant Fusion Proteins / Sequence Alignment / Sequence Homology, Amino Acid / Serine Endopeptidases / Transfection / alpha 1-Antitrypsin (Link to Publication Site) ● Publication site (DOI): 10.1042/BJ20051524 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 16433634 ● Search Scopus @ Elsevier (PMID): 16433634 ● Search Scopus @ Elsevier (DOI): 10.1042/BJ20051524 (DOI: 10.1042/BJ20051524, PubMed: 16433634)
45.	Akihiko Tsuji, Tadashi Yoshimoto, Keizo Yuasa and Yoshiko Matsuda : Pratamine: a unique and patent inhibitor of oligopeptidase B, Journal of Peptide Science, Vol.12, No.1, 65-71, 2006. (Summary) Oligopeptidase B is a serine endopeptidase found in prokaryotes, unicellular eukaryotes and higher plants. The enzyme has been shown recently to play a central role in the pathogenesis of several parasitic diseases such as African trypanosomiasis, and to be a potential therapeutic target. This study reports that protamine, a basic peptide rich in arginine, is a potent inhibitor at the nanomolar level of oligopeptidase B from E. coli and wheat. Protamines 1B, 2C, 3A and TP17 displayed similar inhibitory activities and were capable of binding strongly to oligopeptidase B without proteolytic cleavage. The concentration of protamine needed for 50% inhibition (IC50) of oligopeptidase B was 10(4)-fold lower than the IC50 of trypsin. Oligopeptidase B was highly sensitive to inhibition by protamines even in the presence of serum (IC50, 1 microM). These data indicate that protamines might provide information useful for the design of more specific synthetic oligopeptidase B inhibitors. (Keyword) Enzyme Inhibitors / Escherichia coli / Protamines / Serine Endopeptidases / Structure-Activity Relationship / Triticum (Link to Publication Site) ● Publication site (DOI): 10.1002/psc.683 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 15948139 ● Search Scopus @ Elsevier (PMID): 15948139 ● Search Scopus @ Elsevier (DOI): 10.1002/psc.683 (DOI: 10.1002/psc.683, PubMed: 15948139)
46.	Akihiko Tsuji, Keizo Yuasa and Yoshiko Matsuda : Identification of Oligopeptidase B in Higher Plants. Purification and Characterization of Oligopeptidase B from Quiescent Wheat Embryo, Triticum aestivum, The Journal of Biochemistry, Vol.136, No.5, 673-681, 2004. (Summary) 小麦発芽過程において，著明に活性が上昇するセリンプロテアーゼを見出し，小麦胚芽より各種クロマトグラフィーによって単一に精製した．精製酵素はオリゴぺプチダーゼbに類似した酵素学的性質を有するとともに，その部分アミノ酸配列も大腸菌やトリパノゾーマオリゴぺプチダーゼbの配列と極めて相同性が高かった．以上の結果より，今までバクテリアやトリパノゾーマのみ存在すると考えられていたオリゴペプチダーゼbは高等植物にも存在することが蛋白レベルで証明された． (Keyword) Adrenocorticotropic Hormone / Amino Acid Sequence / Electrophoresis, Polyacrylamide Gel / Escherichia coli / Hydrogen-Ion Concentration / Hydrolysis / Kinetics / Molecular Sequence Data / Molecular Weight / Seeds / Sequence Alignment / Sequence Homology, Amino Acid / Serine Endopeptidases / Sodium Chloride / Somatostatin / Somatostatin-28 / Species Specificity / Time Factors / Triticum (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvh163 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 15632308 ● Search Scopus @ Elsevier (PMID): 15632308 ● Search Scopus @ Elsevier (DOI): 10.1093/jb/mvh163 (DOI: 10.1093/jb/mvh163, PubMed: 15632308)
47.	Akihiko Tsuji, Manami Tsuji, Hirimiti Takami, Shingo Nakamura and Yoshiko Matsuda : Molecular cloning and expression analysis of novel wheat cysteine protease, Biochimica et Biophysica Acta (BBA) - General Subjects, Vol.1670, No.1, 84-89, 2004. (Summary) 小麦発芽過程で著明に発現量が増加する新規のシステインプロテアーゼのクローニングを行った．得られたcDNAから推定されるアミノ酸配列はこのプロテアーゼがパパインファミリーに属する新規のシステインプロテアーゼであることを示した．このプロテアーゼは小麦，大麦に存在し，米，シロイヌナズナには存在しなかった．抗体を作成し，その蛋白レベルにおける発現を調べると，発芽2日目以降に著明に増加することが判明し，小麦の発芽過程における細胞の増殖，分化に寄与している可能性が高い． (Keyword) Amino Acid Sequence / Base Sequence / Cloning, Molecular / Cysteine Endopeptidases / DNA, Complementary / Gene Expression Regulation, Enzymologic / Germination / Molecular Sequence Data / Sequence Alignment / Triticum (Link to Publication Site) ● Publication site (DOI): 10.1016/j.bbagen.2003.10.009 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 14729145 ● Search Scopus @ Elsevier (PMID): 14729145 ● Search Scopus @ Elsevier (DOI): 10.1016/j.bbagen.2003.10.009 (DOI: 10.1016/j.bbagen.2003.10.009, PubMed: 14729145)
48.	Shizuyo Koide, Ichiro Yoshida, Akihiko Tsuji and Yoshiko Matsuda : The expression of proprotein convertase PACE4 is highly regulated by Hash-2 in placenta: Possible role of placenta-specific basic helix-loop-helix transcription factor, human Achaete-Scute homologue-2, The Journal of Biochemistry, Vol.134, No.3, 433-440, 2003. (Summary) 培養細胞Bewoは絨毛癌に由来する培養細胞で，トロホブラスト細胞のモデル細胞として使われている．Bewo細胞にはPACE4が発現し，その発現はE-boxに結合するbHLH転写因子で制御されることを明らかにした．また，ヒト胎盤の免疫染色によってPACE4は胎盤トロホブラストの分化過程で発現量が変動することを見出した． (Keyword) Animals / Base Sequence / Cell Line, Tumor / E-Box Elements / Gene Expression Regulation / Helix-Loop-Helix Motifs / Humans / Placenta / Promoter Regions, Genetic / Proprotein Convertases / Rats / Serine Endopeptidases / Tissue Distribution / Transcription, Genetic / Transfection (Link to Publication Site) ● Publication site (DOI): 10.1093/jb/mvg161 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 14561729 ● Search Scopus @ Elsevier (PMID): 14561729 ● Search Scopus @ Elsevier (DOI): 10.1093/jb/mvg161 (DOI: 10.1093/jb/mvg161, PubMed: 14561729)
49.	Akihiko Tsuji, Kensuke Sakurai, Emi Kiyokage, Takahito Yamazaki, Shizuyo Koide, Kazunori Toida, Kazunori Ishimura and Yoshiko Matsuda : Secretory proprotein convertases PACE4 and PC6A are heparin-binding proteins which are localized in the extracellular matrix. Potential role of PACE4 in the activation of proproteins in the extracellular matrix, Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, Vol.1645, No.1, 95-104, 2003. (Summary) PACE4のヘパリンセファロースゲルに対する結合能を調べた．PACE4はヘパリンカラムに強く結合し，塩化ナトリウム1モル以上の濃度で溶出された．また，結合したPACE4はヘパリンで溶出されたが，コンドロイチン硫酸では全く溶出されなかった．PACE4のC末に塩基性アミノ酸に富んだヘパリン結合部位を同定した．さらに，培養細胞を用いてPACE4は細胞外マトリックスに存在するプロテオグリカン分子のヘパラン硫酸糖鎖に結合していることを証明した． (Keyword) Amino Acid Sequence / Binding Sites / Cells, Cultured / Enzyme Activation / Extracellular Matrix / Heparan Sulfate Proteoglycans / Heparin / Humans / Immunohistochemistry / Molecular Sequence Data / Placenta / Proprotein Convertase 5 / Proprotein Convertases / Serine Endopeptidases / Subtilisins / Trophoblasts (Link to Publication Site) ● Publication site (DOI): 10.1016/S1570-9639(02)00532-0 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 12535616 ● Search Scopus @ Elsevier (PMID): 12535616 ● Search Scopus @ Elsevier (DOI): 10.1016/S1570-9639(02)00532-0 (DOI: 10.1016/S1570-9639(02)00532-0, PubMed: 12535616)
50.	Miwa Bando, Atsushi Matsuoka, Akihiko Tsuji and Yoshiko Matsuda : The proprotein convertase PACE4 is upregulated by PDGF-BB in megakaryocytes: gene expression of PACE4 and furin is regulated differently in Dami cells, The Journal of Biochemistry, Vol.132, No.1, 127-134, 2002. (Summary) The differentiation of megakaryocytes into platelets is highly regulated by many cytokines and growth factors. PACE4 and furin are Ca(2+)-dependent serine endoproteases belonging to the subtilisin-like proprotein convertase (SPC) family. These enzymes are involved in the proteolytic activation of proteins that play essential roles in cell growth and differentiation. In this study, we examined the expression of PACE4 and furin during the differentiation of megakaryoblastic cell lines, Dami and HEL cells, induced by phorbol 12-myristate 13-acetate (PMA). PMA stimulates not only the expression of platelet-derived growth factor-B (PDGF-B) mRNA, but also PACE4 mRNA in these cell lines. The expression of PACE4 transcripts (both the PACE4A and PACE4C/CS isoforms) was upregulated more than 4-fold by PMA. Moreover, direct treatment with PDGF-BB also resulted in an increase in the level of PACE4 mRNA. Further, the effect of PDGF-BB on PACE4 expression was confirmed by promoter assay of the PACE4 gene. Although the furin mRNA level was increased by TGF-beta1 in Dami cells, it was not affected by PDGF-BB. These results indicate for the first time that PACE4 expression is specifically upregulated by PDGF-BB in differentiated megakaryoblasts, suggesting a unique role for PACE4 in platelet production. (Keyword) megakaryocytes / PACE4 / PDGF-BB / proprotein convertase / transcriprional regulation (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 12097169 ● CiNii @ National Institute of Informatics (CRID): 1390282679908516864 ● Summary page in Scopus @ Elsevier: 2-s2.0-0036330386 (PubMed: 12097169, CiNii: 1390282679908516864, Elsevier: Scopus)
51.	K. Utsumi, Akihiko Tsuji, R. Kase, A. Tanaka, T. Tanaka, E. Uyama, T. Ozawa, H. Sakuraba, Y. Komaba, M. Kawabe, Y. Iino and Y. Katayama : Western blotting analysis of the beta-hexosaminidase alpha and beta-subunits in cultured fibroblasts frim cases of various forms of GM2 gangliosidosis, Acta Neurologica Scandinavica, Vol.105, No.6, 427-430, 2002. (Summary) GM2ガングリオシドーシス患者皮膚培養繊維芽細胞のヘキソサミニダーゼ発現量や分子量をウエスタンブロット法で解析する方法を確立し，この方法は，簡便で正確なGM2ガングリオシドーシス診断法であることを証明した． (Keyword) Adult / Antibodies / Blotting, Western / Cells, Cultured / Female / Fibroblasts / Gangliosidoses, GM2 / Hexosaminidase A / Hexosaminidase B / Humans / Infant / Luminescent Measurements / Male / Sandhoff Disease / Skin / Tay-Sachs Disease / beta-N-Acetylhexosaminidases (Link to Publication Site) ● Publication site (DOI): 10.1034/j.1600-0404.2002.01097.x (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 12027830 ● Search Scopus @ Elsevier (PMID): 12027830 ● Search Scopus @ Elsevier (DOI): 10.1034/j.1600-0404.2002.01097.x (DOI: 10.1034/j.1600-0404.2002.01097.x, PubMed: 12027830)
52.	Akihiko Tsuji, Takayuki Ikoma, Emi Hashimoto and Yoshiko Matsuda : Development of selectivity of α1-antitrypsin variant by mutagenesis in its reactive site loop against proprotein convertase. A crutial role of the P4 arginine in PACE4 inhibition, Protein Engineering, Vol.15, No.2, 123-130, 2002. (Summary) ズブチリシンプロプロテインコンベルターゼ(SPC)は，哺乳類では7種類が同定されているが，これらの酵素は類似した基質特異性を持ち，さらに複数の酵素がひとつの細胞で発現しているため，その生理機能解析は困難である．本研究では，これらSPCの機能解析のツールとして使用できる選択的阻害特性を有する阻害タンパクを作成することを目的として，アンチトリプシンのタンパク工学的改変を行い，その阻害特異性について検討した．SPCの中でPACE4とFurinを識別可能な改変体を作成することができた． (Keyword) Animals / Arginine / Cadherins / DNA Primers / Furin / Humans / Mutagenesis, Site-Directed / Polymerase Chain Reaction / Precipitin Tests / Proprotein Convertase 5 / Proprotein Convertases / Rats / Recombinant Proteins / Serine Endopeptidases / Serine Proteinase Inhibitors / Subtilisins / Transfection / Tumor Cells, Cultured / alpha 1-Antitrypsin (Link to Publication Site) ● Publication site (DOI): 10.1093/protein/15.2.123 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 11917148 ● Search Scopus @ Elsevier (PMID): 11917148 ● Search Scopus @ Elsevier (DOI): 10.1093/protein/15.2.123 (DOI: 10.1093/protein/15.2.123, PubMed: 11917148)
53.	Takazumi Taniguchi, Rieko Kuroda, Kensuke Sakurai, Masami Nagahama, Ikuo Wada, Akihiko Tsuji and Yoshiko Matsuda : A Critical Role for the Carboxy Terminal Region of the Proprotein Convertase, PACE4A, in the Regulation of Its Autocatalytic Activation Coupled with Secretion, Biochemical and Biophysical Research Communications, Vol.290, No.2, 878-884, 2002. (Summary) ズブチリン様プロプロテインコンベルターゼファミリーであるPACE4は，自己活性化されたのち分泌されるが，培養細胞で過剰発現しても，その自己活性化速度が非常に遅いため，分泌される活性型酵素の量が少ない．本研究では，PACE4のC末端側に自己活性化を制御する領域があることを証明した． (Keyword) Amino Acid Sequence / Cell Line / Enzyme Activation / Humans / Kidney / Molecular Sequence Data / Mutagenesis, Site-Directed / Peptide Fragments / Proprotein Convertases / Protein Precursors / Protein Processing, Post-Translational / Protein Structure, Quaternary / Sequence Deletion / Serine Endopeptidases / structureactivity relationship / Subtilisins / Ultracentrifugation (Link to Publication Site) ● Publication site (DOI): 10.1006/bbrc.2001.6282 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 11785985 ● Search Scopus @ Elsevier (PMID): 11785985 ● Search Scopus @ Elsevier (DOI): 10.1006/bbrc.2001.6282 (DOI: 10.1006/bbrc.2001.6282, PubMed: 11785985)
54.	Ichiro Yoshida, Shizuyo Koide, Shin-ichi Hasegawa, Akira Nakagawara, Akihiko Tsuji and Yoshiko Matsuda : Proprotein convertase PACE4 is down-regulated by the basic helix-loop-helix transcription factor hASH-1 and MASH-1, The Biochemical Journal, Vol.360, No.3, 683-689, 2001. (Summary) Mash-1は，神経分化に必須のbHLH型転写因子であるが，その標的遺伝子は不明である．本研究では，種々のMash-1発現量の異なるヒト神経芽細胞腫を用いて，Mash-1のズブチリシン様プロプロテインコンベルターゼであるPACE4発現に及ぼす影響について検討し，Mash-1はPACE4遺伝子上流のE-boxクラスタに結合し，そのプロモーター活性をネガティブに制御する転写因子であることを証明した． (Link to Publication Site) ● Publication site (DOI): 10.1042/0264-6021:3600683 (Link to Search Site for Scientific Articles) ● CiNii @ National Institute of Informatics (CRID): 1573950399804603392 ● Summary page in Scopus @ Elsevier: 2-s2.0-0035893574 (DOI: 10.1042/0264-6021:3600683, CiNii: 1573950399804603392, Elsevier: Scopus)
55.	Kenji Mori, Akiyoshi Imamaki, Kaya Nagata, Yasuo Yonetomi, Ryoko Kiyokage-Yoshimoto, John T Martin, Matthew T Gillespie, Masami Nagahama, Akihiko Tsuji and Yoshiko Matsuda : Subtilisin-Like Proprotein Convertase, PACE4 and PC8, as Well as Furin, are Endogenous Proalbumin Convertases in HepG₂ Cells, The Journal of Biochemistry, Vol.125, No.3, 627-633, 1999. (Summary) 血清アルブミンはプロ体として肝臓で合成されたのち，細胞内でプロペプチドがプロセスされ血液中に分泌される．しかしこのプロセシング機構についてはまだ不明な点が多い．本研究では，アンチセンスRNA法を用いて，ズブチリシン様プロプロテインコンベルターゼであるFurin, PACE4, PC8のアルブミンプロセシングにおける役割を解析し，Furinのみならず，PACE4, PC8もアルブミンプロセシングに関わっていることを証明した．
56.	Akihiko Tsuji, Emi Hashimoto, Takayuki Ikoma, Takazumi Taniguchi, Kenji Mori, Masami Nagahama and Yoshiko Matsuda : Inactivation of Proprotein Convertase, PACE4 by α₁-Antitrypsin Portland (α₁-PDX) which is a Blocker of Proteolytic Activation of Bone Morphogenetic Protein during Embryogenesis., The Journal of Biochemistry, Vol.126, No.3, 591-603, 1999. (Summary) PACE4はSPCファミリーに属するプロテアーゼで，特に細胞分化過程でその発現量が高度に制御されているが，その機能に関しては不明な点が多い．本研究では，BMPの活性化を阻害し，細胞分化を抑制するアンチトリプシン改変体が，PACE4に対して強い阻害作用を持つことを証明し，PACE4がBMPプロセシングプロテアーゼとして機能していることを示唆した．またPACE4の酵素学的性質についても検討した．
57.	Akihiko Tsuji, Shigeru Yoshida, Shinichi Hasegawa, Miwa Bando, Ichiro Yoshida, Shizuyo Koide, Kenji Mori and Yoshiko Matsuda : Human Subtilisin-Like Proprotein Convertase, PACE4 (SPC4) Gene Expression Is Highly Regulated through E-Box Elements in HepG₂ and GH₄C₁ Cells, The Journal of Biochemistry, Vol.126, No.3, 494-502, 1999. (Summary) ヒトPACE4の発現制御機構を明らかにするため，その遺伝子5'上流域の構造，機能解析を行った．ヒトPACE4遺伝子上流域には多くの転写因子結合サイトが認められたが，特に上流1kbにE-box配列が12個存在した．リポータージーンアッセイにより，E4-E9のE-boxクラスタが強力な負のエンハンサーとして機能していることを明らかにした．E-boxに結合するbHLH型転写因子は細胞分化を制御するものが多く，PACE4と細胞分化の関わりを示唆した．
58.	Masami Nagahama, Takazumi Taniguchi, Emi Hashimoto, Akiyoshi Imamaki, Kenji Mori, Akihiko Tsuji and Yoshiko Matsuda : Biosynthetic processing and quoternary interactions of proprotein convertase SPC4 (PACE4)., FEBS Letters, Vol.434, No.1, 155-159, 1998. (Summary) ズブチリン様プロプロテインコンベルターゼファミリーに属するPACE4の活性化機構について解析した．プロPACE4のプロペプチドの切断はPACE4の活性基を変異した不活性体では起こらず，また不活性体と活性体を共発現しても不活性体のプロセシングがなかったことから，このプロセシングは分子内自己活性化であることを証明した．また自己活性化はPACE4の分泌に必須であり，PACE4の活性化と細胞内移行は共役していることを明らかにした．
59.	Masami Nagahama, Takazumi Taniguchi, Emi Hashimoto, Akiyoshi Imamaki, Kenji Mori, Akihiko Tsuji and Yoshiko Matsuda : Biosynthetic processing and quaternary interactions of proprotein convertase SPC4 (PACE4)., FEBS Letter, Vol.434, 155-159, 1998.
60.	Hideaki Nagamune, Hiroki Kourai, Ayako Katsuura, Yasuki Taoka, Kenzo Fushitani, Robert A. Whiley, Kikuji Yamashita, Akihiko Tsuji, Yoshiko Matsuda, Hiroki Kourai and Seiichiro Kitamura : Intermedilysin A cytolytic toxin specific for human cells of a Streptococcus intermedius isolated from human liver abscess in "Streptococci and The Host", Advances in Experimental Medicine and Biology, Vol.418, 773-775, 1997. (Summary) S. intermedius肝膿瘍分離株から単離されたヒト細胞特異的な細胞溶解毒素インターメディリシン(ILY)の性質を検討した．これまで知られていたチオール基修飾試薬への非感受性やコレステロールへの低感受性に加え，電子顕微鏡観察からILYは直径30-50nmの膜孔を形成し細胞を破壊することが示された．またILYの部分遺伝子構造解析の結果からILYはチオール活性化毒素群と同じ分子祖先から進化したものであることが示唆された．
61.	Akihiko Tsuji, Chiemi Hine, Yasuhiro Tamai, Kayoko Yonemoto, Kazushige Mori, Shigeru Yoshida, Miwa Bando, Eri Sakai, Kenji Mori, Tetsuya Akamatsu and Yoshiko Matsuda : Genomic organization and alternative splicing of human PACE4 (SPC4), Kexin-like processing endoprotease., The Journal of Biochemistry, Vol.122, No.2, 438-452, 1997. (Summary) 本研究ではプロセシング酵素Kexin familyの1つPACE4のヒトゲノムを単離し, その構造を解析し, ファミリーの中でも最も大きく複雑な構造であること, 多種類存在するアイソフォームが全て同一遺伝子から選択的スプライシングにより生じること, およびプロモーター領域がユニークな構造であること等を初めて明らかにした. (Keyword) Processing protease / PACE4 / Gene structure / Alternative splicing
62.	Tetsuya Akamatsu, Shigeo Daikoku, Hideaki Nagamune, Shigeru Yoshida, Kenji Mori, Akihiko Tsuji and Yoshiko Matsuda : Developmental expression of a novel Kexin family protease, PACE4E, in the rat olfactory system., Histochemistry and Cell Biology, Vol.108, No.2, 95-103, 1997. (Summary) 本研究では新規Kexin family proteaseであるPACE4Eがラット脳発生過程において時期·空間特異的に発現することを明らかにし, 特に嗅球においては僧帽細胞に極めて特異的に発現し, 僧帽細胞の分化成熟と密接に関わることを見い出した. (Keyword) Processing protease / Olfactory system / Mitral cell
63.	Kenji Mori, Sachiko Kii, Akihiko Tsuji, Masami Nagahama, Akiyoshi Imamaki, Keiko Hayashi, Tetsuya Akamatsu, Hideaki Nagamune and Yoshiko Matsuda : A Novel Human PACE4 Isoform, PACE4E Is an Active Processing Protease Containing a Hydrophobic Cluster at the Carboxy Terminus, The Journal of Biochemistry, Vol.121, No.5, 941-948, 1997. (Summary) ズブチリシン様プロプロテインコンベルターゼファミリーに属するPACE4の新規アイソフォームPACE4E-I, PACE4E-IIをヒト脳cDNAライブラリーよりクローニングした．PACE4Eは，PACE4Aと比べ，C末に75残基からなる，疎水性アミノ酸のクラスタを含むユニークな配列を持っていた．この部分を欠失したPACE4Eは分泌速度が上昇した．共発現実験によって，PACE4E-1, E-IIは，PACE4Aと同様にプロセシング活性を持っていることが証明された．
64.	Hideaki Nagamune, Kazuaki Muramatsu, Masakatsu Higashine, Tetsuya Akamatsu, Yasuhiro Tamai, Yasuo Yonetomi, Akihiko Tsuji, Keisuke Izumi, Takemasa Sakaguchi, Tetsuya Yoshida and Yoshiko Matsuda : Cyclic AMP-inducible procalcitonin processing in thyroidal parafollicular cells is regulated by the kexin family protease, PC1, Biomedical and Health Research, Vol.15, 22-33, 1997. (Summary) 甲状腺におけるカルシトニンのプロセシングに関与するズブチリシン様プロプロテインコンベルターゼを同定するため，特異抗体による免疫染色とモデル細胞TT細胞におけるcAMPの影響について調べた．カルシトニン産生細胞で発現していたのは，主にPC1とPC2であったが，カルシトニンの産生を刺激するcAMPによって発現が増加したのは，PC1であった．これらの結果は，PC1がプロカルシトニンの活性化酵素であることを示している．
65.	Hideaki Nagamune, Chikako Ohnishi, Ayako Katsuura, Kenzo Fushitani, Robert A. Whiley, Akihiko Tsuji and Yoshiko Matsuda : Intermedilysin, a novel cytotoxin specific for human cells, secreted by Streptococcus intermedius UNS46 isolated from a human liver abscess, Infection and Immunity, Vol.64, No.8, 3093-3100, 1996. (Summary) S. intermediusのヒト肝膿瘍分離株がヒト細胞特異的な細胞溶解毒素インターメディリシン(ILY)を分泌することを発見した． ILYは約54KDaの蛋白で，その部分一次構造はニューモリシンと高い相同性を示した．しかしSH修飾試薬により失活せず，コレステロール対しても感受性が低かった．またトリプシン処理したヒト赤血球はILYに対して抵抗性を示すようになることから，ILY作用には宿主細胞膜の蛋白質が関与している事が示された．
66.	Akihiko Tsuji, Keisuke Edazawa, Koji Sakiyama, Kaya Nagata, Yuka Sasaki, Hideaki Nagamune and Yoshiko Matsuda : Purification and characterization of a novel serine proteinase from the microsomal fraction of bovine pancreas., The Journal of Biochemistry, Vol.119, No.1, 100-105, 1996. (Summary) 牛膵臓膜画分より，CHAPSによる可溶化，ゲルろ過，トリプシンインヒビターセファロース，アルギニンーセファロースによって，新規のトリプシン様セリンプロテアーゼを単一に精製した．この酵素は分子量29500で，グルカゴンを用いて切断特異性を調べると，塩基性アミノ酸とロイシンのC末側を切断した．N末のアミノ酸配列は，エラスターゼIV, II, IIIに類似していた．
67.	Akihiko Tsuji, Hideaki Nagamune, Tetsuya Akamatsu, Chiemi Hine, Kazuaki Muramatsu, Kenji Mori, Yasuhiro Tamai, Kayoko Yonemoto and Yoshiko Matsuda : Novel members of mammalian kexin family proteases, PACE4C, PACE4D, PC7A and PC7B., Intracellular Protein Catabolism, Vol.389, 63-71, 1996.
68.	新本美智枝, Akihiko Tsuji, 楊瑞成, 鈴木義之, 野村房子 and 瀬川昌也 : 制限酵素断片長多型を利用した連鎖解析によるX染色体劣性遺伝病の遺伝子診断, 日本小児科学会雑誌, Vol.91, 3440-3447, 1996. (Summary) X染色体劣性遺伝病である筋ジストロフィーとオルニチントランスカルバミラーゼ欠損症について，その患者と家系のRFLP解析を行った．白人のRFLP解析で有用なプローブは，日本人においても有効であり，人種による差は認められなかった．この方法は，キャリアー診断においても有用であることが判明した．
69.	Hideaki Nagamune, Kazuaki Muramatsu, Tetsuya Akamatsu, Yasuhiro Tamai, Keisuke Izumi, Akihiko Tsuji and Yoshiko Matsuda : Distribution of the kexin family proteases in pancreatic islets., --- PACE4C is specifically expressed in B cells of pancreatic islets ---, Endocrinology, Vol.136, No.1, 357-360, 1995. (Summary) Kexin Family Proteasesはペプチドホルモン等のプロセシング酵素として同定され, 血糖調節ホルモンのインスリン等のプロセシングにも関与することが示唆されていたが, 本研究では我々が新規にクローニングしたPACE4CをはじめとするKexin Family Proteasesとインスリン等との膵島細胞での局在, 共存関係を初めて明らかにし, その成果は雑誌表紙にも掲載された. (Keyword) Precessing protease / Pancreatic islet / Insulin / Glucagon
70.	Akihiko Tsuji, Chiemi Hine, Kenji Mori, Yasuhiro Tamai, Kazuya Higashine, Hideaki Nagamune and Yoshiko Matsuda : A Novel Member, PC7, of The Mammalian Kexin-Like Protease Family, --- Homology to PACE4A, Its Brainspecific Expression and Identification of Isoforms ---, Biochemical and Biophysical Research Communications, Vol.202, No.3, 1452-1459, 1994. (Summary) 哺乳類には7種類のKexin様プロプロテインコンベルターゼが存在する．哺乳類Kexin様プロテアーゼ間で保存された触媒領域のプライマーを用い，ラット脳下垂体からcDNA断片をPCR増幅し，これをプローブにしてラット脳下垂体cDNAライブラリーからPC7をクローニングした．PC7(PC7AとPC7B)はRXKRの活性化サイトを持ち，触媒，HomoB及びシステインリッチ領域を持っており，PACE4Aと非常に高いホモロジーを示した．PC7は脳で強く発現し，肝，腎，心，脾では検出されなかった．
71.	Akihiko Tsuji, Kenji Mori, Chiemi Hine, Yasuhiro Tamai, Hideaki Nagamune and Yoshiko Matsuda : The tissue distribution of mRNAs for the PACE4 isoforms, kexin-like processing protease: PACE4C and PACE4D mRNAs are major transcripts among PACE4 isoforms, Biochemical and Biophysical Research Communications, Vol.202, No.3, 1215-1221, 1994. (Summary) 新たに見いだされたPACE4アイソフォーム(PACE4CとPACE4D)の組織分布をRT-PCR法を用いて解析した．両アイソフォームのmRNAともに，PACE4Aと同様に多くの組織で見いだされたが，それぞれの発現パターンは異なっていた．また，PACE4C及びPACE4DのmRNAはPACE4Aのそれより遙かに多く，PACE4 mRNAの主要な部分はPACE4CとPACE4DのmRNAである事が示唆された．
72.	Akihiko Tsuji, Ryoichi Oda, Kohji Sakiyama, Hideaki Nagamune, Kouji Itou, Ryoichi Kase, Hitoshi Sakuraba, Yoshiyuki Suzuki and Yoshiko Matsuda : Lysosomal enzyme replacement using α2-macroglobulin as a transport vehicle, The Journal of Biochemistry, Vol.115, No.5, 937-944, 1994. (Summary) α2-マクログロブリン(α2M)を運搬担体に用いたリソソーム酵素欠損性遺伝病の酵素補充療法システムを開発した． α2Mはレセプターを介してリソソームに取り込まれることから，豚肝臓から精製した α-グルコシダーゼ或いはα-ガラクトシダーゼにα2Mを運搬担体として二価架橋試薬で連結し，リソソーム指向性の酵素補充療法剤を構築した．これを用いてin vitro実験を行い，ポンペ病及びファブリー病細胞の細胞病態が改善されることを証明した．
73.	Akihiko Tsuji, Kazuya Higashine, Chiemi Hine, Kenji Mori, Yasuhiro Tamai, Hideaki Nagamune and Yoshiko Matsuda : Identification of Novel cDNAs Encording Human Kexin-like Protease, PACE4 Isoforms, Biochemical and Biophysical Research Communications, Vol.200, No.2, 943-950, 1994. (Summary) ヒト胎盤cDNAライブラリーより，新規のPACE4アイソフォーム，PACE4CとPACE4Dをクローニングし，塩基配列を決定した．PACE4Cは，PACE4AのC末に存在するシステインリッチ領域を持たず，ホモBドメインのC末にC-特異的配列を持っていた．PACE4Dはシグナルペプチド，プロペプチドおよびシステインリッチ領域を欠失していた．
74.	Akihiko Tsuji, Tetsuya Akamatsu, Hideaki Nagamune and Yoshiko Matsuda : Identification of targeting proteinase for rat α1-macroglobulin in vivo., --- Mast-cell tryptase is a major component of the α1-macroglobulin-proteinase complex endocytosed into rat liver lysosomes. ---, The Biochemical Journal, Vol.298, No.1, 79-85, 1994. (Summary) 血清蛋白質であるα-マクログロブリン(αM)は種々の酵素に対するインヒビターとして機能すると考えられていたが, 生体内で標的となる酵素は不明であった. 本研究ではαMが酵素と複合体形成後, リソゾームに運ばれ分解されることに着目し, リソゾーム内の分解を抑制したラット肝臓リソゾームよりαM-酵素複合体を単離し, 肥満細胞トリプターゼが生体内でαMの標的酵素の1つであることを初めて明らかにした. (Keyword) α-macroglobulin / Mast-cell tryptase / Lysosome
75.	Torres-Rosado Adrian, O'Shea K.Sue, Akihiko Tsuji, Chou Shan-Ho and Kurachi Kotoku : Hepsin, a putative cell-surface serine protease, is required for mammalian cell growth, Proceedings of the National Academy of Sciences of the United States of America, Vol.90, No.15, 7181-7185, 1993. (Summary) 細胞膜結合セリンプロテアセーゼ，ヘプシンの生理機能解析のため，抗体による阻害，またはアンチセンスRNAによる発現阻害の細胞に及ぼす影響を調べた．ヘプシンの活性を現象させると，著名な細胞増殖の抑制が見られ，ヘプシンは細胞増殖を制御するプロテアーゼであることが明らかになった．
76.	Akihiko Tsuji, Adrian Torres-Rosado, Toshiro Arai, Michelle M. LeBeau, Richard S. Lemons, Shan-Ho Chou and Kotoku Kurachi : Hepsin, a cell membrane-associated protease, --- characterization, tissue distribution and gene localization ---, The Journal of Biological Chemistry, Vol.266, No.25, 16948-16953, 1991. (Summary) ヘプシンはトリプシン様セリンプロテアーゼの活性セリン周辺の保存された配列から設計されたオリゴヌクレオチドプローブによって，肝臓からクローニングされたが，その機能に関しては全く不明である．本研究では，染色体局在，ノーザンブロットによる組織分布，ペプチド抗体によるヘプシンタンパクの同定を行った．ヘプシン遺伝子は第19染色体に局在し，特に肝臓で強く発現していた．免疫染色によって，ヘプシンは膜結合酵素として細胞膜に発現していることが判明した．
77.	Akihiko Tsuji, Tishiro Arai, P. S. Furcinitti, J. P. Langmore and Kotoku Kurachi : The major component of a large, intracellular proteinase accumulated by inhibitors is a complex of alpha-2 macroglobulin and thrombin, Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, Vol.1078, No.1, 85-93, 1991. (Summary) A large, intracellular proteinase accumulated by inhibitors (PABI) was found in cultured mammalian cells as a large, multicatalytic proteinase with a greatly elevated concentration in the presence of small peptide proteinase inhibitors (Tsuji and Kurachi (1989) J. Biol. Chem. 264, 16093). Electron microscopic analysis showed that the tertiary structure of PABI highly resembled that of alpha 2-macroglobulin complexed with a proteinase(s). Isolation of the anti-PABI cross-reacting material from calf serum added to the culture media of baby hamster kidney cells further supported that the primary component of PABI was alpha 2-macroglobulin. Immunoblot analyses and the substrate specificity of PABI indicated that the major proteinase component contained in PABI was thrombin. When alpha 2-macroglobulin was added to the PABI-depleted serum, a significant accumulation or a degradation of the intracellular alpha 2-macroglobulin was observed in the presence or absence of leupeptin, respectively. Similarly, when thrombin was added to the PABI-depleted fetal calf serum supplemented with fresh alpha 2-macroglobulin, a significant amount of intracellular thrombin was found only in the presence of leupeptin. These results indicate that the major component of the intracellular PABI molecules is a complex of alpha 2-macroglobulin with thrombin which is internalized from the culture media. Intracellular accumulation of PABI, therefore, is a phenomenon primarily relevant to the culture cells. Whether or not PABI is also generated in certain physiological or pathological conditions requires further study. (Keyword) Amino Acid Sequence / Animals / Cattle / Cells, Cultured / Cross Reactions / Endopeptidases / Image Processing, Computer-Assisted / Immunoblotting / Microscopy, Electron / Molecular Sequence Data / Protein Binding / Thrombin / alpha-Macroglobulins (Link to Publication Site) ● Publication site (DOI): 10.1016/0167-4838(91)90096-I (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 1710931 ● Search Scopus @ Elsevier (PMID): 1710931 ● Search Scopus @ Elsevier (DOI): 10.1016/0167-4838(91)90096-I (DOI: 10.1016/0167-4838(91)90096-I, PubMed: 1710931)
78.	P. Hu, A J J Reuser, H. C. Janse, W. J. Kleijer, D. Schindler, H. Sakuraba, Akihiko Tsuji, Y. Suzuki and O. van P. Diggelen : Biosynthesis of human alpha-N-acetylgalactosaminidase: Defective phosphorylation and maturation in infantile alpha-NAGA deficiency, Biochemical and Biophysical Research Communications, Vol.175, No.No 3, 1097-1103, 1991. (Summary) 正常者およびα-N-アセチルガラクトサミニダーゼ欠損症患者繊維芽細胞におけるα-N-アセチルガラクトサミニダーゼの生合成を解析した．正常者では分子量52,000の前駆体として合成された後，リン酸化された49,000の成熟体に変換したが，患者細胞では前駆体が合成されたが，リン酸化されず，細胞内で分解された．
79.	Yang Rei-Cheng, Akihiko Tsuji and Suzuki Yoshiyuki : Abnormal protein spots revealed by two-dimensional electrophoresis in mycoplasma-infected human fibroblasts, Electrophoresis, Vol.11, 344-346, 1990. (Summary) 培養細胞は病因蛋白質の解析に使われるが，マイコプラズマに感染した細胞では，その解析が困難になる．本研究では，マイコプラズマ感染によって特異的に発現が誘導される蛋白6種類を同定した．
80.	Akihiko Tsuji and Kotoku Kurachi : Isolation and characterization of a novel large protease accumulated mammalian cells in the presence of inhibitors, The Journal of Biological Chemistry, Vol.264, No.27, 16093-16099, 1989.
81.	Yuji Iwasaki, Akihiko Tsuji, Kiyoshi Omura and Yoshiyuki Suzuki : Purification and characterization of beta-mannosidase from human placenta, The Journal of Biochemistry, Vol.106, No.2, 331-335, 1989. (Summary) ヒト胎盤から約10,000倍にβマンノシダーゼを精製した．分子量は110,000で，等電点は4.7であった．主に高マンノース型糖鎖うぃ持っていた．至適pHは，pH 4.5であった．
82.	Rei-Cheng Yang, Yoshiyuki Suzuki and Akihiko Tsuji : Two-dimentional electrophoresis aided personal computer analusis for screening of mutant proteins in inherited diseases, Electrophoresis, Vol.10, No.11, 785-792, 1989. (Summary) 2次元電気泳動を用い，正常者と先天代謝異常症患者皮膚繊維芽細胞の発現蛋白質の違いを検討した．泳動パターンをコンピューターに取込み，移動度が異なる蛋白，発現量に差がある蛋白を敏速に検出できる方法を開発した．
83.	Akihiko Tsuji and Kotoku Kurachi : Isolation and characterization of a novel large protease accumulated in mammalian cells in the presence of inhibitors, The Journal of Biological Chemistry, Vol.264, No.No 27, 16093-16099, 1989. (Summary) トリプシン様セリンプロテアーゼおよびシステインプロテアーゼ阻害剤であるロイペプチン存在下で培養したBHK細胞に分子量の大きなプロテアーゼが蓄積することを見いだし，単一に精製し，その酵素学的性質について検討した．分子量約700,000で，トリプシン様活性を持ち，阻害剤を培養液から除くと敏速に細胞内で分解された．
84.	Ohshima Akihiro, Akihiko Tsuji, Nagao Yoshiro, Sakuraba Hitoshi and Suzuki Yoshiyuki : Cloning, sequencing, and expression of cDNA for human beta-galactosidase, Biochemical and Biophysical Research Communications, Vol.157, No.No 1, 238-244, 1988. (Summary) 完全長ヒトβガラクトシダーゼcDNAをヒト胎盤cDNAライブラリーより，抗体を用いたスクリーニング法でクローニングした．677アミノ酸からなる配列が推定された．7か所のN型糖鎖結合サイトが存在した．また培養細胞で，このcDNAを発現させ，βガラクトシダーゼ蛋白の発現と活性の上昇を確認した．このcDNAによって，GM1-ガングリオシドーシス症の遺伝学的研究が可能となった．
85.	Akihiko Tsuji and Yoshiyuki Suzuki : Solubilization of Membrane-Bound Acid α-Glucosidase by Proteolysis, Biochemical and Biophysical Research Communications, Vol.151, No.3, 1358-1363, 1988.
86.	Akihiko Tsuji, Kiyoshi Omura and Yoshiyuki Suzuki : Intracelluler Transport of Acid α-Glucosidase in Human Fibroblasts: Evidence for Involvement Phosphomannosyl Receptor-Independent System, The Journal of Biochemistry, Vol.104, 276-278, 1988. (Summary) 本論文は，正常Ⅰ-cell病皮膚繊維芽細胞におけるα-グルコシダーゼ及びβ-ヘキソサミニダーゼの活性，細胞内局在性を比較したものである．Ⅰ-cell病患者は，多くのリソゾーム酵素の活性が低下する．しかし酸性α-グルコシダーゼの活性，細胞内局在性はⅠ-cell病細胞においても異常が認められず，本酵素のリソゾーム転送機構は，従来報告されているホスホマンノシルレセプター系とは異なる可能性が示唆された．
87.	Yoshiyuki Suzuki, Akihiko Tsuji, Kiyoshi Omura, Gen Nakamura, Shoichi Awa, Mrian Kroos and Arnold J.J. Reuser : Km mutant of acid α-glucosidase in a case of cardiomyopathy without signs of skeletal muscle involvement, Clinical Genetics, Vol.33, 376-385, 1988. (Summary) 本論文は，16歳時に不整脈と診断され，17歳時に急性心不全で死亡した患者の酸性α-グルコシダーゼを生検試料，皮膚線維芽細胞を用いて研究したものである．その結果，患者の酸性α-グルコシダーゼはグリコーゲンに対するKm値が，正常酵素に比べ異常に高いことが判明し，この症例は今まで報告されていない新しいタイプのKm変異酸性α-グルコシダーゼによる糖原病Ⅱ型であることがわかった．
88.	Eiji Nanba, Akihiko Tsuji, Kiyoshi Omura and Yoshiyuki Suzuki : Galactosialidosis: molecular heterogeneity in biosynthesis and processing of peotective protein for β-galactosidase, Human Genetics, Vol.80, 329-332, 1988. (Summary) 本論文は，ガラクトシアリドーシス患者皮膚線維芽細胞におけるβ-ガラクトシダーゼの保護タンパク(プロテクティブプロテイン)の生合成，プロセッシングについて詳細に検討したものである．
89.	Shinmoto Michie, Akihiko Tsuji and Suzuki Yoshiyuki : Restriction fragment length polymorphism on the short arm of X-chromosome among the Japanese population, Japan Journal of Human Genetics, Vol.33, 333-338, 1988. (Summary) 日本人におけるX染色体短腕DNAプローブ(13種類)の制限酵素断片長多型性について，白人の多型性と比較した．プローブ782，754は，得られる断片は白人の結果より長く，またプローブC7, pERT87-8, XJ5.1は断片の比率が白人の場合と全く異なっていた．これらの結果は，RFLP解析で使用されるプローブは人種の違いを考慮する必要があることを示した．
90.	Akihiko Tsuji, Omura Kiyoshi and Suzuki Yosiyuki : I-cell disease: evidence for a mannose 6-phospahte independent pathway for translocation of lysosomal enzymes in lymphoblastoid cells, Clinica Chimica Acta, Vol.176, 115-122, 1988. (Summary) I-cell病患者と正常者のリンパ芽球細胞を培養し，リソソーム酵素の動態について解析，検討した．I-cell病患者皮膚繊維芽細胞ではほとんどのリソソーム酵素活性は著明に低下するが，リンパ芽球細胞ではガラクトシダーゼ以外の酵素活性はほぼ正常であり，細胞内局在性やプロセシングも正常細胞と差は見られず，マンノース6リン酸経路以外のリソソーム酵素局在化機構の存在が示唆された．
91.	Michie Shinmoto, Akihiko Tsuji, Rei-Cheng Yang, Yoriko Nomura, Masaya Segawa and Yosiyuki Suzuki : DNA deletion and recombination in the gene locus of X-linked muscular dystrophies, Journal of Inherited Metabolic Disease, Vol.11, 324-328, 1988. (Summary) 日本人のドゥシェンヌ型，ベッカー型筋ジストロフィー患者のRFLP解析結果をまとめた．日本人も白人と同様の比率で，遺伝子欠損または交差がみられた．また一つの家系では，ジストロフィーのプローブでは異常はないが，その近傍で遺伝子欠損が認められた．
92.	Akihiko Tsuji and Yoshiyuki Suzuki : Biosynthesis of two components of human acid α-glucosidase, Archives of Biochemistry and Biophysics, Vol.259, No.2, 234-240, 1987. (Summary) 本論文は，酸性α-グルコシダーゼの生合成過程を詳細に検討するために分子量の異なる2種類の本酵素(分子量8万と7万1千)を分離精製し，両タンパクの物理化学的，タンパク化学的，免疫学的性質を比較したものである．更に，ヒト線維芽細胞の標識実験により，80K，71K分子の合成過程も検討した．その結果，両酵素は同一前駆体からプロセスされるものの，アミノ末端部分に差がみられることがわかった．
93.	Akihiko Tsuji and Yoshiyuki Suzuki : The precursor of acid α-glucosidase is synthesized as a membrane bound enzyme, Biochemistry International, Vol.15, No.5, 945-952, 1987. (Summary) A pulse-chase study in human skin fibroblasts showed that a 110 kDa precursor of acid alpha-glucosidase was synthesized as a membrane-bound protein, which was solubilized in vitro not by mannose 6-phosphate or 1M KCl but by Triton X-100 or trypsin. This 110 kDa precursor form bound to the membrane was detected in control fibroblasts treated with tunicamycin and in I-cell disease fibroblasts as well. The precursors in human placenta were found also in the membrane fraction. It was concluded that the newly synthesized acid alpha-glucosidase precursor is located on the surface of the membrane, and the phosphomannosyl receptor does not participate in the enzyme-membrane binding. (Keyword) Cell Membrane / Cells, Cultured / Chromatography, Gel / Female / Fibroblasts / Humans / In Vitro Techniques / Leucine / Placenta / alpha-Glucosidases (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 3325063 ● Search Scopus @ Elsevier (PMID): 3325063 (PubMed: 3325063)
94.	Akihiko Tsuji and Yoshiyuki Suzuki : Purification of human placental acid α-mannosidase by an immunological method, Biochemistry International, Vol.15, No.3, 483-489, 1987. (Summary) An immunoaffinity column was used for the purification of alpha-mannosidase from human placenta. The enzyme was purified to homogeneity by extraction in the presence of various protease inhibitors, immunoaffinity chromatography, Ultrogel AcA-34 gel filtration and hydroxyapatite chromatography. Two subunits were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Their molecular weights were 65 kDa and 27 kDa. Heterogeneity of the molecular weight of the large subunit was not observed in our preparation. This method is relatively simple and rapid for obtaining the purified enzyme which is structurally not modified during purification procedures. (Keyword) Amino Acids / Centrifugation, Density Gradient / Chromatography, Affinity / Chromatography, Gel / Electrophoresis, Polyacrylamide Gel / Female / Humans / Immune Sera / Kinetics / Mannosidases / Placenta / alpha-Mannosidase (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 3426623 ● Search Scopus @ Elsevier (PMID): 3426623 (PubMed: 3426623)
95.	Eiji Nanba, Akihiko Tsuji, Kiyoshi Omura and Yoshiyuki Suzuki : Galactosialidosis: A direct evidence that 49-kilodalton protein restores deficient enzyme activities in fibroablast, Biochemical and Biophysical Research Communications, Vol.144, No.1, 138-142, 1987. (Summary) 本研究は，ガラクトシアリドーシス患者で，欠損する分子量49kDaのタンパク質を，正常繊維芽細胞培養液より分離後，患者細胞に加え，患者細胞中のβ-ガラクトシダーゼ，シアリダーゼ活性の変化を調べたものであり，49kDaタンパク質はβ-ガラクトシダーゼ，シアリダーゼの安定化に寄与していることが分かった．
96.	Akihiko Tsuji, Rei-Cheng Yang, Kiyoshi Omura, Tomoko Imabayashi and Yoshiyuki Suzuki : A simple differential immunoprecipitation assey of urinary asid and neutral α-glucosidase for glucogenosisII, Clinica Chimica Acta, Vol.167, 313-320, 1987. (Summary) 本論文は，糖源病Ⅱ型の診断法として，酸性α-グルコシダーゼ特異抗体を用いた尿中の酸性及び中性グルコシダーゼ活性測定法を開発したものである．本法によって，他の方法で糖源病Ⅱ型と診断されている患者尿と正常者の酸性α-グルコシダーゼ活性を測定すると，患者尿では活性がみられず，本法がこの遺伝病の診断に有効であることが判明した．
97.	Eiji Nanba, Akihiko Tsuji, Kiyoshi Omura and Yoshiyuki Suzuki : Galactosialidosis: Studies on residual enzymes in early and late onset clinical phenotypes, Journal of Clinical Biochemistry and Nutrition, Vol.3, No.2, 149-157, 1987. (Summary) 本論文は，β-ガラクトシダーゼ，シアリダーゼの欠損するガラクトシアリドーシスの乳児型，成人型患者皮膚繊維芽細胞の残存β-ガラクトシダーゼ，シアリダーゼの性質を比較したものである．乳児型，成人型とも著明に両酵素活性が低下しているものの，有意に乳児型に比べ成人型で両活性が高かった． (Link to Publication Site) ● Publication site (DOI): 10.3164/jcbn.3.149 (Link to Search Site for Scientific Articles) ● Search Scopus @ Elsevier (DOI): 10.3164/jcbn.3.149 (DOI: 10.3164/jcbn.3.149)
98.	Matsuko Moriyasu, Kichiko Koike, Yoshishige Urata, Akihiko Tsuji and Masahiko Koike : Rapid and simple isolation procedure for three component enzyme of pig heart 2-oxoglutarate dehydrogenase complex, Journal of Nutritional Science and Vitaminology, Vol.32, No.No 1, 33-40, 1986. (Summary) 本研究は，高速液体クロマトグラフィーを用いて敏速かつ簡単に，ブタ心筋より2-オキソグルタル酸脱水素酵素複合体の3つの成分酵素を精製する方法を開発したもので，この方法により，従来の方法より高収量で，3つの各成分酵素を分離，精製することが可能となった．
99.	Katsuko Yamashita, Yoko Tachibana, Akira Hitoi, Yoshiko Matsuda, Akihiko Tsuji, Nobuhiko Katunuma and Akira Kobata : Differences in the sugar chains of two subunits and isozymic forms of rat kidney γ-glutamyltranspeptidase, Archives of Biochemistry and Biophysics, Vol.227, No.1, 225-232, 1983. (Summary) 本研究は，ラット腎γ-グルタミルトランスぺプチダーゼを構成する2種のサブユニットの糖鎖構造を各々明らかにしたものである．
100.	Yoshiko Matsuda, Akihiko Tsuji, Nobuhiko Katunuma and N. Katunuma : Biosynthesis and degradation of gamma-glutamyltranspeptidase of rat kidney, The Journal of Biochemistry, Vol.94, No.3, 755-765, 1983. (Summary) ラット腎臓γ-glutamyltranspepidaseの生合成，分解について調べた．ラットに放射性アミノ酸を投与し，蛋白質を標識後，抗体によって腎臓のγ-glutamyltranspepidaseを分離し，生合成について解析した．その結果，本酵素は一本鎖で合成された後，重鎖と軽鎖にプロセスされ，細胞膜に局在化することが明らかになった．
101.	Katsuko Yamashita, Akira Hitoi, Yoshiko Matsuda, Akihiko Tsuji, Nobuhiko Katunuma and Akira Kobata : Structural studies of the carbohydrate moieties of rat kidney γ-glutamyltranspeptidase, The Journal of Biological Chemistry, Vol.258, No.2, 1098-1107, 1983. (Summary) 本研究は，ラット腎γ-グルタミルトランスペプチダーゼの糖鎖構造を調べたもので，極めて多様性に富む糖鎖構造の概要が明らかにされた．
102.	Yoshiko Matsuda, Akihiko Tsuji and Nobuhiko Katunuma : Studies on the structure of γ-glutamyltranspeptidase. III. Evidence that the amino terminus of the heavy subunit is membrane binding segment, The Journal of Biochemistry, Vol.93, No.5, 1427-1433, 1983. (Summary) γ-グルタミルトランスペプチダーゼの膜結合様式について詳細に検討し，ヘビーサブユニットのアミノ末端付近の疎水性アミノ酸に富む部分が膜との結合に関与していることを明らかにした．
103.	Tsutomu Miura, Yoshiko Matsuda, Akihiko Tsuji and Nobuhiko Katunuma : Immunological cross reactivity of γ-glutamyltranspeptidase from human and rat kidney, liver and bile, The Journal of Biochemistry, Vol.89, No.1, 217-222, 1981. (Summary) 本研究は，ヒト腎及びラット腎よりγ-グルタミルトランスペプチダーゼを精製し，各々特異抗体を調製したのち，ラット腎，肝及びヒト腎，胆汁中のγ-グルタミルトランスペプチダーゼの免疫学的性質を比較検討した．
104.	Yoshiko Matsuda, Akihiko Tsuji and Nobuhiko Katunuma : Studies on the structure of γ-glutamyltranspeptidase. I. Correlation between sialylation and isozymic, The Journal of Biochemistry, Vol.87, No.No 4, 1243-1248, 1980. (Summary) 本研究は，γ-グルタミルトランスペプチダーゼの等電点多様性とシアル酸含量の関係を調べたもので，等電点の多様性は，タンパク部分ではなく糖鎖中のシアル酸含量の違いによることを明らかにした．
105.	Akihiko Tsuji, Yoshiko Matsuda and Nobuhiko Katunuma : Studies on the structure of g-glutamyltranspeptidase. II. Location of the segment anchoring γ-glutamyltranspeptidase to the membrane, The Journal of Biochemistry, Vol.87, No.No 6, 1567-1571, 1980. (Summary) 本研究は，γ-グルタミルトランスペプチダーゼの膜結合様式について調べたもので，ヘビーサブユニットのアミノ末端付近に膜結合部位があることを明らかにした．
106.	Akihiko Tsuji, Yoshiko Matsuda and Nobuhiko Katunuma : Characterization of human serum γ-glutamyltranspeptidase, Clinica Chimica Acta, Vol.104, No.3, 361-366, 1980. (Summary) 本研究は，ヒト血清，胆汁中のγ-グルタミルトランスペプチダーゼの構造比較を行い，本酵素の血液·胆汁中への遊出機序について検討した．
107.	Akihiko Tsuji, Yoshiko Matsuda and Nobuhiko Katunuma : Purification and characterization of γ-glutamyltranspeptidase from human bile, Biomedical Research, Vol.1, 410-416, 1980. (Summary) 本研究は，ヒト胆汁よりγ-グルタミルトランスペプチダーゼを単一に精製し，その性質を調べた．
108.	Yoshiko Matsuda, Akihiko Tsuji, Nobuhiko Katunuma, Masanori Hayashi and Yoshiyata Takahashi : Studies on liver argininosuccinate synthetase in a patient with citrullinemia and in normal subjects, The Journal of Biochemistry, Vol.85, No.No 1, 192-195, 1979. (Summary) 本研究は，肝生検試料を用いてシトルリン血症患者のアルギノコハク酸合成酵素の性質を正常者酵素と比較したもので，患者酵素の動力学的性質に異常を認め，臨床症状と酵素異常の関係を論じた．
109.	上田敦子, 大西悦子, Shiro Saito, Yoshiko Matsuda, Akihiko Tsuji and 勝沼信彦 : 血清Ornithin carbamyltransferaseの脂肪肝に対する診断的意義, 臨床化学, Vol.6, No.2, 125-130, 1978. (Summary) 本研究は，実験的肝炎，脂肪肝のウサギ血清オルニチントランスカルバミルトランスフェラーゼ活性を調べ，血清オルニチントランスカルバミルトランスフェラーゼ活性値は，脂肪肝の指標になることを示した．

Academic Paper (Unrefereed Paper):

1.	Yoshiko Matsuda, Hitoshi Hori, Akihiko Tsuji, Hideko Nagasawa, Yoshihiro Uto, Rieko Kuroda, 葛西宗江, Miwa Bando, 吉田一郎, 上田祐司 and サハルディン B. モハマッド : 新医薬品の開発をめざした脳神経特異的発現遺伝子産物のプロテオミクス解析, Bulletin of Faculty of Engineering, The University of Tokushima, Vol.46, 49-56, 2001. (Summary) 本論文では，新薬開発における新規標的分子の同定のための脳神経特異的遺伝子産物のプロテオミクス解析に関する成果を報告した．
2.	Masami Nagahama, Ichiro Yoshida, Keiichi Uyemura, Akihiko Tsuji and Yoshiko Matsuda : Proteolytic Cleavage of the Neural Cell Adhesion Molecule L1 by Subtilisin-Like Proprotein Convertases, Keio University Symposia for Life Science and Medicine, Vol.2, 261-266, 1999. (Summary) L1蛋白質は神経の分化，機能と関係していると考えられている接着蛋白である．L1蛋白質はプロセス部位にSubtilisin-like proprotein convertaseの切断配列である塩基性アミノ酸対を持っているが，そのプロセシング機構は不明である．本研究では，L1蛋白質のプロセシングに関与するプロテアーゼについて，特にSPCについて検討した．培養細胞でL1蛋白質前駆体とSPC (furin, PACE4, PC6, PC1, PC4, PC8)を共発現すると，furin, PACE4, PC6によってプロセシングされることが判明した．

Review, Commentary:

1.	Akihiko Tsuji : Extracellular processing protease PACE4 : Significance of diversity of SPC family proteases, Seikagaku, Vol.75, No.4, 305-310, Apr. 2003. (Summary) 哺乳類ズブチリシン様プロプロテインコンベルターゼが現在7種類同定されている．これらの酵素はシグナルペプチド，プロペプチド，触媒ドメイン，ホモBドメインからなる共通構造を有するが，そのC末構造は異なっている．Furin, PC8は膜結合領域を持ち，PACE4, PC6はシステインリッチドメインを持っている．PC1, PC2, PC4は比較的短いC末配列を有している．本論文では，これらSPCの異なる細胞内局在，発現制御，機能解析に関する最新の知見を紹介し，SPCの多様性の生理的意義について考察する． (Link to Search Site for Scientific Articles) ● CiNii @ National Institute of Informatics (CRID): 1520290884016266112 (CiNii: 1520290884016266112)
2.	森健二, Akihiko Tsuji and Yoshiko Matsuda : 先天性異常アルブミン症，無アルブミン血症，先天代謝異常症候群-遺伝子解析の進歩と成果-, Nihon Rinsho. Japanese Journal of Clinical Medicine, Vol.19, No.5, 657-670, May 1998. (Summary) 無アルブミン症，異常アルブミン症について，最近の解析結果を解説した．
3.	森健二, Akihiko Tsuji and Yoshiko Matsuda : 先天性異常アルブミン症，無アルブミン血症, 先天代謝異常症候群ー遺伝子解析の進歩と成果, Vol.19, 123-127, 1998. (Summary) アルブミンの異常症の分類，病因について，最新の知見を解説した．
4.	Yoshiko Matsuda and Akihiko Tsuji : 前駆体蛋白質の活性化とケキシンファミリー，プロセシングプロテアーゼ, 蛋白質核酸酵素, Vol.42, No.14, 2355-2361, Aug. 1997.
5.	Yoshiko Matsuda, Akihiko Tsuji, Hideaki Nagamune, Tetsuya Akamatsu, Chiemi Hine, Kazuaki Muramatsu, Kenji Mori, Yasuhiro Tamai and Kayoko Yonemoto : Novel members of mammalian Kexin family proteases, PACE4C, PACE4D, PC7A and PC7B., Advances in Experimental Medicine and Biology, Vol.389, 63-71, Sep. 1996. (Summary) ペプチドホルモンや増殖因子等は不活性な前駆体として合成され, 特定の塩基性アミノ酸部位での限定切断を経て初めて活性化される. Kexin family proteaseはこのプロセシングを担う重要な酵素であり, 本研究では我々が新規メンバーをクローニングし, その構造, 組織分布等を明らかにした. (Keyword) Processing protease
6.	Yoshiko Matsuda, Akihiko Tsuji, Hideaki Nagamune, Tetsuya Akamatsu, Chiemi Hine, Kazuaki Muramatsu, Kenji Mori, Yasuhiro Tamai and Kayoko Yonemoto : Novel members of mammalian Kexin family proteases, PACE4C, PACE4D, PC7A and PC7B, Intracellular protein catabolism, Vol.389, No.7, 63-71, Jan. 1996.
7.	Akihiko Tsuji, 窪野慎一, Yuka Sasaki, Yoshiko Matsuda, 中山久幸 and 寺山日出男 : ピロリドンカルボン酸ソーダによるビオプラーゼの安定化とレンズ洗浄剤としての応用, 日本コンタクトレンズ学会誌, Vol.38, 232-236, Jan. 1996. (Summary) ピロリドンカルボン酸は，ビオプラーゼ(ズブチリシン)の自己分解作用を抑制し，安定化した．ビオプラーゼはピロリドンカルボン酸溶液中では室温においても安定であり，冷却の必要がなく，コンタクトレンズの洗浄剤として有用であると考えられた．
8.	桜場均, 石井達, 加瀬良一, Kouji Itou, 奥宮敏可, Akihiko Tsuji and 佐藤能雅 : 遺伝性代謝病:病因解明と治療法開発への分子遺伝学的アプローチ, 小児科診療, Vol.59, No.5, 821-828, 1996. (Summary) Fabry病を例にとり，遺伝子上の変異の多様性と発現蛋白の性状について，さらに昆虫細胞におけるα-ガラクトシダーゼの大量発現，構造解析，酵素補充療法への応用について最新の知見について解説した．
9.	鈴木義之, 新本美智枝, Akihiko Tsuji and 楊瑞成 : Duchenne型筋ジストロフィーの診断と治療, 小児科, Vol.29, No.1, 27-40, Jan. 1988. (Summary) 本論文は，ドゥシェンヌ型筋ジストロフィーの遺伝子診断，チオールプロテアーゼ阻害剤による治療の試みについて述べたものである．
10.	Suzuki Yoshiyuki, Omura Kiyoshi, Nanba Eiji, Akihiko Tsuji, Yang Rei-Cheng, Kato Goro, Sakuraba Hitoshi and Igarashi Takashi : Lysosomopathies: Analysis of intracellular abnormalities of functional proteins and a survey of new therapeutic approach, Child Neurology and Developmental Disabilities, 21-27, 1988. (Summary) 遺伝子異常疾患における酵素異常の多様性とプロテアーゼ阻害剤投与による異常酵素の安定化と治療への応用について解説した．
11.	T. Izumi, Y. Fukuyama, Akihiko Tsuji, T. Yamanaka, Y. Hirabayashi and Y. Suzuki : GM2-Gangliosidosis: B1 variant with thermostable beta-hexosaminidase A and molecular analysis of the mutany enzyme, Lipid Storage Disorders, 237-245, 1988. (Summary) 新しいタイプのGM2-Gangliosidosis患者のヘキソサミニダーゼの生合成並びに酵素学的性質について，αおよびβサブユニットに対する特異抗体を用い解析した．
12.	Akihiko Tsuji : RFLPによる遺伝病の解明，診断, Chemistry & Chemical Industry, Vol.40, No.10, 882-883, 1987. (Summary) 制限酵素断片長多型性による遺伝病の病因遺伝子の検索法，および遺伝病の診断法について最近の進歩について解説する．
13.	鈴木義之, 新本美智枝, Akihiko Tsuji and 楊瑞成 : 神経，筋疾患のDNA診断, 組織培養, Vol.13, 346-351, 1987. (Summary) 本論文は，RFLP解析による神経，筋疾患の遺伝子診断法について解説したものである．
14.	Yoshiko Matsuda, Akihiko Tsuji and Nobuhiko Katunuma : Quantitative abnormality of argininosuccinate synthetase in a patient with citrullinemia, Advancesin Experimental Medicine and Biology, Urea Cycle Disease, Vol.153, 77-82, 1984.
15.	Yoshiko Matsuda, Akihiko Tsuji, 久野健夫 and 勝沼信彦 : γ-GTPの生体内運命，血清，胆汁への逸脱機構, 日本臨床化学会記録, Vol.23, 12-13, 1983. (Summary) 膜結合酵素であるγ-GTPは，特にアルコール性肝炎など肝障害時，胆汁のうっ滞時血中へ遊離しγ-GTP値が急増する．本研究では，γ-GTPの生合成と細胞内移行，胆汁，血中への遊離機構について，最新の知見を解説する．
16.	林真功, 高橋善弥太, Yoshiko Matsuda, Akihiko Tsuji and 勝沼信彦 : アルギニノコハク酸合成酵素の質的異常に起因するシトルリン血症, Metabolism and Disease, Vol.15, No.No 9, 1253-1257, 1978. (Summary) シトルリン血症の特殊な1例を経験し，本症の病因がASSの量には差がなく，質的異常に起因することを明らかにしたので，報告する．

Proceeding of International Conference:

1.	Shogo Abe, Saki Hirose, Mami Nishitani, Akihiko Tsuji and Keizo Yuasa : Citrus peel polymethoxyflavones, sudachitin and nobiletin, induce distinct cellular responses in human keratinocyte HaCaT cells via the MAPK pathways, The 7th International Conference on Food Factors (ICoFF2019)/The 12th International Conference and Exhibition on Nutraceuticals and Functional Foods (ISNFF2019), Kobe, Japan, Dec. 2019.
2.	Matsuda Shinya, Akihiko Tsuji and Keizo Yuasa : PCTK3/CDK18 regulates cell migration by negatively modulating the FAK1 activity, 16th International Conference of Biochemistry and Molecular Biology: Signalling Pathways in Development, Disease and Aging, Vancouver, BC, Canada, Jun. 2016.
3.	Shinya Matsuda, Kyohei Kominato, Akihiko Tsuji and Keizo Yuasa : PCTAIRE kinase 3/cyclin dependent kinase 18 is activated through association with cyclin A and/or phosphorylation by protein kinase A, Experimental Biology 2015, Boston, Massachusetts, Mar. 2015. (Link to Publication Site) ● Publication site (DOI): 10.1074/jbc.M113.542936 (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 24831015 ● Search Scopus @ Elsevier (PMID): 24831015 ● Search Scopus @ Elsevier (DOI): 10.1074/jbc.M113.542936 (DOI: 10.1074/jbc.M113.542936, PubMed: 24831015)
4.	Akihiko Tsuji : Characterization of proteases expressed in cotyledons of daikon radish during germination., 6th General Meeting of the International Proteolysis Society, Gold Coast, Oct. 2009.
5.	Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : The role of subutilisin-like proproteinn connvertase PACE4 in myogenesis, 5th General Meeting of the International Proteolysis Society, Patras, Greece, Oct. 2007.
6.	Kentaro Ogawa, Tsuyoshi Yuasa, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : Characterization of proteases expressed in the embryo of germinating wheat seed, International Symposium on Medical and Biological Perspectives in Proteases and Their Inhibitors. Satellite meeting of the 20th IUBMB International congress and 11th FAOBMB Congress, Hyogo, Jun. 2006.
7.	Keizo Yuasa, Kaori Suzue, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Subtilisin-like proprotein convertase PACE4 is transcriptionally regulated by E2F/Rb, International Symposium on Medical and Biological Perspectives in Proteases and Their Inhibitors. Satellite meeting of the 20th IUBMB International congress and 11th FAOBMB Congress, Hyogo, Jun. 2006.
8.	Hiroki Kanie, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : Development of selectivity of 1-antitrypsin variant by mutagenesis in reactive site loop against proprotein convertase, International Symposium on Medical and Biological Perspectives in Proteases and Their Inhibitors. Satellite meeting of the 20th IUBMB International congress and 11th FAOBMB Congress, Hyogo, Jun. 2006.
9.	Tetsuya Masuda, Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Transcriptional Activation of processing protease PACE4 during myogenic differentiation., International Symposium on Medical and Biological Perspectives in Proteases and Their Inhibitors. Satellite meeting of the 20th IUBMB International congress and 11th FAOBMB Congress, Hyogo, Jun. 2006.
10.	Akihiko Tsuji : A novel protein which regulates autoactivation and secretion of subtilisin-like proprotein convertase. International Symposium on Medical and Biological Perspectives in Proteases and Their Inhibitors, Satellite meeting of the 20th IUBMB International congress and 11th FAOBMB Congress, Hyogo, Jun. 2006.
11.	Yayoi Kikuchi, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : RCN3 facilitates the secretion and activation of PACE4, 20th IUBMB International Congress of Biochemistry and Molecular Biology and 11th FAOBMB Congress, Kyoto, Jun. 2006.
12.	Keizo Yuasa, Shin Yamagami, Shotaro Uehara, Masami Nagahama and Akihiko Tsuji : cGMP-dependent protein kinase II is required for chondrogenic differentiation, 20th IUBMB International Congress of Biochemistry and Molecular Biology and 11th FAOBMB Congress, Kyoto, Jun. 2006.
13.	Takuya Yanagino, Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Transcriptional Regulation of Fibrillin-2 Gene during Chondrogenrsis, 20th IUBMB International Congress of Biochemistry and Molecular Biology and 11th FAOBMB Congress, Kyoto, Jun. 2006.
14.	Akihiko Tsuji, Keizo Yuasa, Masami Nagahama and Yoshiko Matsuda : Role of PACE4 and furin in cell differentiation, International Conference on Serine-Carboxyl Peptidases 2005, Kyoto, Nov. 2005.
15.	Akihiko Tsuji, Yayoi Kikuchi and Yoshiko Matsuda : Reticulocalbin-3: A novel protein which regulates biosynthesis of subtilisin-like proprotein convertase, The 1st Pacific-Rim International Conference on Protein Science, 129, Yokohama, Apr. 2004. (Summary) レチキュロカルビン3は，遺伝子構造から推定される新規のCRECファミリーに属するカルシウム結合蛋白質であるが，これまで蛋白レベルの研究はまったく行われていない．我々はSPC阻害蛋白を安定発現するGH4C1細胞に特異的にレチキュロカルビン3が蓄積することを発見し，この蛋白質のカルシウム結合能，細胞分布，細胞内局在性を調べた．また，この蛋白質はSPCの前駆体と小胞体で一時的に結合することを見出し，SPCの生合成にシャペロンとして関わることを示唆した．
16.	Go Futamatsu, Tomonori Shimada, Akihiko Tsuji and Yoshiko Matsuda : Functional analysis of the processing protease PACE4 during chondrogenic differentiation of ATDC5 cells, International Symposium on Medical and Biological Perspectives in Proteases and Their Inhibitors., 36, Hyogo, Nov. 2003.
17.	Akihiko Tsuji, Shizuyo Koide, Keizo Yuasa and Yoshiko Matsuda : Involvement of PACE4 in activation of TGF beta-related factors in the ECM, International Symposium on Medical and Biological Perspectives in Proteases and Their Inhibitors., 20, Hyogo, Nov. 2003.
18.	Akihiko Tsuji, Yayoi Kikuchi, Yukimi Sato, Shizuyo Koide and Yoshiko Matsuda : Reticulocalbin-like protein (RCN3): A novel protein which regulates biosynthesis of subtilisin-like proprotein convertase (SPC), 3rd General Meeting of the International Proteolysis Society, 187, Nagoya, Nov. 2003.
19.	Akihiko Tsuji and Yoshiko Matsuda : Enhancement of selectivity of α1-antitrypsin variant against subtilisin-like proprotein convertase by mutagenesis in RSL and application for functional analysis, Second General Meeting of International Proteolysis Society, 59, Germany, Oct. 2001. (Summary) 非常に基質特異性の類似した哺乳類ズブチリシン様プロプロテインコンベルターゼ，PACE4, furin, PC6に選択性にある阻害蛋白質を，α1アンチトリプシンのタンパク工学的改変によって作成することができた，
20.	Akihiko Tsuji : Biosynthesis of PACE4, Hormonal and neural peptide biosynthesis Gordon Research Conference, New Hampshire, Jul. 2000.
21.	Akihiko Tsuji and Yoshiko Matsuda : Unique Mechanism Governing Cell-Specific Expression and Biosynthetic Processing of PACE4, International Proteolysis Society First Gemeral Meeting, 38, Michigan, USA, Sep. 1999.
22.	Akihiko Tsuji and Yoshiko Matsuda : Regulation of PACE4 expression under dynamic bHLH network and its physiological importance, 6th CGGH Symposium "Roles of Proteolysis in Health and Disease", 3, Tokushima, May 1999.
23.	Kenji Mori, Akihiko Tsuji and Yoshiko Matsuda : Subtilisin-like Proprotein Convertase, PACE4, participates in the Processing of Plasma protein precursors in human hepatoma HepG2 cells, 6th CGGH Symposium "Roles of Proteolysis in Health and Disease", 80, Tokushima, May 1999.
24.	Shin-ichi Hasegawa, Ichiro Yoshida, Shizuyo Koide, Akihiko Tsuji and Yoshiko Matsuda : Transcriptional Regulation of the Human PACE4 Gene Expression in HepG2 and GH₄C₁ cells, 6th CGGH Symposium "Roles of Proteolysis in Health and Disease", 83, Tokushima, May 1999.
25.	Emi Hashimoto, Takayuki Ikoma, Akihiko Tsuji and Yoshiko Matsuda : Proprotein Convertase PACE4 is inhibited by α₁-Antitrypsin cariant Portland, 6th CGGH Symposium "Roles of Proteolysis in Health and Disease", 82, Tokushima, May 1999.
26.	Miwa Bando, Atsushi Matsuoka, Akihiko Tsuji and Yoshiko Matsuda : Physiological Function of PACE4 on megakaryoblastic differentiation, 6th CGGH Symposium "Roles of Proteolysis in Health and Disease", 84, Tokushima, May 1999.
27.	Ichiro Yoshida, Shizuyo Koide, Akira Nakagawara, Akihiko Tsuji and Yoshiko Matsuda : Regulation of PACE4 Expression by bHLH factor in neuroblastoma cells, 6th CGGH Symposium "Roles of Proteolysis in Health and Disease", 85, Tokushima, May 1999.
28.	Akihiko Tsuji : Enzymatic characterization of human SPC4, Hormonal and neural peptide biosynthesis Gordon Research Conference, New Hampshire, Aug. 1998.
29.	Hideaki Nagamune, Chikako Ohnishi, Ayako Katsuura, Yasunori Taoka, Kenzo Fushitani, Robert A. Whiley, Kikuji Yamashita, Akihiko Tsuji, Yoshiko Matsuda, Hiroki Kourai and Seiichiro Kitamura : Intermedilysin A cytolytic toxin specific for human cells of a Streptococcus intermedius isolated from human liver abscess, XIII Lancefield International Symposium on Streptococci and Streptococcal Diseases; In "Streptococci and The Host"(T. Horaud, A. Bouvet, R. Leclercq, H. de Montclos, M. Sicard, eds.): Advances in Experimental Medicine and Biology, Vol.418, 773-775, Paris, Sep. 1997. (Summary) S. intermedius肝膿瘍分離株から単離されたヒト細胞特異的な細胞溶解毒素インターメディリシン(ILY)の性質を検討した．これまで知られていたチオール基修飾試薬への非感受性やコレステロールへの低感受性に加え，電子顕微鏡観察からILYは直径30-50nmの膜孔を形成し細胞を破壊することが示された．またILYの部分遺伝子構造解析の結果からILYはチオール活性化毒素群と同じ分子祖先から進化したものであることが示唆された．
30.	Kenji Mori, 今牧明義, Tetsuya Akamatsu, Akihiko Tsuji, Masami Nagahama and Yoshiko Matsuda : A novel human PACE4 isoform, PACE4E, is an active protease containing a hydrophobic cluster at the carboxy terminus, 17th International Congress of Biochemistry and Molecular Biology, San Francisco, Aug. 1997.
31.	Akihiko Tsuji, Chiemi Hine, Shigeru Yoshida, Miwa Bando, Kenji Mori, Tetsuya Akamatsu and Yoshiko Matsuda : Gene organization and Alternative Splicing of Human PACE4, Subtilisin-Like Proprotein Convertase, 17th International Congress of Biochemistry and Molecular Biology, FASEB.J., Vol.11, No.9, 1224, San Francisco, Aug. 1997.
32.	Hideaki Nagamune, Kazuaki Muramatsu, Masakatsu Higashine, Tetsuya Akamatsu, Yasuhiro Tamai, Yasuo Yonetomi, Akihiko Tsuji, Keisuke Izumi, Takemasa Sakaguchi, Tetsuya Yoshida and Yoshiko Matsuda : Cyclic AMP-inducible procalcitonin processing in thyroidal parafollicular cells is regulated by the Kexin family protease, PC1., International Symposium on Medical Aspects of Protease Inhibitors; In "Medical Aspects of Proteases and Protease Inhibitors": Biomedical and Health Research, Vol.15, 22-33, Amsterdam, May 1997. (Summary) ラット甲状腺においてKexin様プロテアーゼの免疫組織化学的解析を行った．その結果，カルシトニンを作る傍濾胞細胞ではPC1とPC2が強く発現しており，FurinやPC6の発現はわずかで，PACE4は検出できなかった．傍濾胞細胞株TTのウエスタンブロットでもPC1，PC2， Furin，PC6は検出されたが，cAMP刺激による成熟カルシトニン量の上昇と連動して急増する酵素はPC1だけであり，cAMP刺激に応答してカルシトニン量を制御するKexin様プロテアーゼはPC1である事を示した． (Keyword) Calcitonin / Processing / PC1
33.	Tetsuya Akamatsu, 吉田茂, 大黒成夫, Hideaki Nagamune, 長濱正巳, Akihiko Tsuji and Yoshiko Matsuda : Expression of a novel PACE4 isoform, PACE4E, in developmental rat brain, The 12th International Symposium of Federation of Asian and Oceanian Biochemists and Molecular Biologists, Tokushima, Jun. 1996.
34.	Akihiko Tsuji, Adrian Torres-Rosado, Toshihiro Arai, Shan-Ho Chou and Kotoku Kurachi : Characterization of hepsin, a membrane bound protease, Biomedica Biochimica Acta, Vol.50, No.4-6, 791-793, 1991. (Summary) Hepsin is a membrane bound protease of 51 kDa present in mammalian cells. It contains a stretch of hydrophobic sequence of 27 amino acid residues in its N-terminal region. By employing fluorescent immunostaining of cells and western blot analysis of the various cell subfractions, the catalytic subunit (carboxyl terminal half) of hepsin is found at the cell surface. The hepsin gene is expressed in most tissues of a young adult baboon, particularly in the liver at high levels. (Keyword) Animals / Cell Line / Cell Membrane / Gene Expression / Humans / Liver / Papio / Serine Endopeptidases / Tissue Distribution (Link to Search Site for Scientific Articles) ● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 1801756 ● Search Scopus @ Elsevier (PMID): 1801756 (PubMed: 1801756)

Proceeding of Domestic Conference:

1.	Sun Xiaomei, Ye Yuxin, Sakurai Naofumi, Kato Koji, Yu Jian, Keizo Yuasa, Akihiko Tsuji and Yao Min : Characterization and ligand-binding manner of EHEP and BGL for producing biofuel from brown algae, 日本結晶学会令和2年(2020年)度年会, Nov. 2020.
2.	Mika Nishida, Maki Shimada, Kenji Miyamoto, Akihiko Tsuji and Keizo Yuasa : ナトリウム利尿ペプチド受容体NPR-Cのグアニンヌクレオチド交換因子GEF-H1を介した新たなシグナル伝達機構, 日本農芸化学会2020年度中四国支部大会(第57回講演会), Sep. 2020.
3.	Mika Nishida, Kenji Miyamoto, Yuki Shimizu, Akihiko Tsuji and Keizo Yuasa : ナトリウム利尿ペプチド受容体NPR-Cの新たなシグナル伝達機構, 日本農芸化学会中四国支部第54回講演会, Jun. 2019.
4.	shogo Abe, Mami Nishitani, Akihiko Tsuji and Keizo Yuasa : 柑橘類果皮ポリメトキシフラボンであるスダチチンは，MAPK経路を介してアポトーシスを誘導する, 日本農芸化学会中四国支部第54回講演会, Jun. 2019.
5.	Akihiko Tsuji and Keizo Yuasa : ユニークな基質特異性をもつアメフラシトリプシン, 第4回日本生物工学会西日本支部講演会, Dec. 2018.
6.	Daichi Takagi, Tomonori Asada, Akihiko Tsuji, 大森謙司 and Keizo Yuasa : Serratia marcescens Lipシステムを利用した大腸菌による低分子抗体分泌産生系の構築, 第4回日本生物工学会西日本支部講演会, Dec. 2018.
7.	Ikuta Shirai, Shota Fujino, Rie Mukai, Akihiko Tsuji and Kei Yamamoto : 骨格筋におけるリン脂質代謝系の網羅的脂質メタボローム解析．第41回日本分子生物学会年会, 第41回日本分子生物学会, Nov. 2018.
8.	Shota Fujino, Akihiko Tsuji and Kei Yamamoto : 栄養飢餓時に変容するリン脂質代謝の役割, 第41回日本分子生物学会, Nov. 2018.
9.	Akihiko Tsuji, Keizo Yuasa and 姚閔 : 海藻ポリフェノールと結合する蛋白質(EHEP)の機能・構造解析, 第91回日本生化学会大会, Sep. 2018.
10.	Mayuki Tamai, Haruna Kataoka, Akihiko Tsuji and Keizo Yuasa : Death-associated protein kinase 2によるオートファジー制御機構の解明, 第91回日本生化学会大会, Sep. 2018.
11.	Akihiko Tsuji and Keizo Yuasa : アメフラシ消化液に含まれるフロロタンニン結合タンパク質の性質, 日本農芸化学会中四国支部第52回講演会, Sep. 2018.
12.	shogo Abe, saki Hirose, 吉田一郎, Akihiko Tsuji and Keizo Yuasa : 柑橘類果皮ポリメトキシフラボンであるスダチチンとノビレチンは，異なる細胞応答を誘導する, 日本農芸化学会中四国支部第52回講演会, Sep. 2018.
13.	Akihiko Tsuji and Keizo Yuasa : GHF45に属する21kDaセルラーゼの作用機構, 第59回日本生化学会中国・四国支部例会, May 2018.
14.	Rumi Morikawa, 小出(吉田) 静代, Jumpei Yamashita, Daichi Takagi, Akihiko Tsuji, 大森謙司 and Keizo Yuasa : Serratia marcescensヘム捕捉タンパク質HasA分泌の分子機構の解明, 日本農芸化学会2018年度大会, Mar. 2018.
15.	Masahiro Okuda, Keizo Yuasa and Akihiko Tsuji : ダイコンの発芽過程における各種システインプロテアーゼの発現解析, 日本農芸化学会中四国支部第50回講演会, Jan. 2018.
16.	Sara Murakami, Yuna Oue, Akihiko Tsuji and Keizo Yuasa : 細胞死関連キナーゼDRAK1とp53の細胞内局在とその役割, 日本農芸化学会中四国支部第50回講演会, Jan. 2018.
17.	Kohei Kawamoto, Matsuda Shinya, Akihiko Tsuji and Keizo Yuasa : 細胞運動・接着におけるPCTK3によるFAKの制御, 2017年度生命科学系学会合同年次大会第40回日本分子生物学会年会,第90回日本生化学会大会, Dec. 2017.
18.	Akihiko Tsuji, Keizo Yuasa, Chikako Asada and Yoshitoshi Nakamura : アメフラシ21K セルラーゼのセルロース分解における機能解析, 第69回日本生物工学会, Sep. 2017.
19.	Daichi Takagi, Shuhei Hamagaki, Akihiko Tsuji, 大森謙司 and Keizo Yuasa : Serratia marcescens type 1 secretion systemを利用した大腸菌による低分子抗体分泌産生系の構築, 第69回日本生物工学会, Sep. 2017.
20.	Daichi Takagi, Shuhei Hamagaki, Akihiko Tsuji, 大森謙司 and Keizo Yuasa : 大腸菌を用いた低分子抗体の分泌生産系の構築, 日本農芸化学会中四国支部第48回講演会, Jun. 2017.
21.	Yuna Oue, Sara Murakami, Akihiko Tsuji and Keizo Yuasa : 細胞死関連プロテインキナーゼDRAK1の細胞内局在化機構, 日本農芸化学会中四国支部第48回講演会, Jun. 2017.
22.	吉田一郎, Chihiro Ito, Matsuda Shinya, Akihiko Tsuji, 矢中規之 and Keizo Yuasa : メラニン産生抑制効果を有する沢瀉成分Alisol Bの作用機序の解明, 日本農芸化学会中四国支部第48回講演会, Jun. 2017.
23.	Norio Kamemura, Sara Murakami, Hiroaki Komatsu, Masahiro Sawanoi, Kenji Miyamoto, 石堂一巳, Koji Kishimoto, Akihiko Tsuji and Keizo Yuasa : 軟骨形成におけるcGMP-dependent protein kinase Ⅱの作用機構の探索., 第58回日本生化学会中国・四国支部例会, May 2017.
24.	Sara Murakami, Yuna Oue, Akihiko Tsuji and Keizo Yuasa : DNA傷害によるアポトーシス誘導におけるDRAK1の関連性, 日本農芸化学会2017年度大会, Mar. 2017.
25.	Toshihiro Kawauchi, Masahiro Okuda, Keizo Yuasa and Akihiko Tsuji : ダイコン由来システインプロテアーゼResponsive to Dehydration 19の同定, 第3回日本生物工学会西日本支部講演会, Dec. 2016.
26.	Shuhei Hamagaki, Daichi Takagi, Takumi Ichihara, Akihiko Tsuji, 大森謙司 and Keizo Yuasa : 大腸菌による1本鎖抗体分泌系の構築, 第3回日本生物工学会西日本支部講演会, Dec. 2016.
27.	Jumpei Yamashita, 小出(吉田) 静代, Akihiko Tsuji, 大森謙司 and Keizo Yuasa : Serratia marcescens HasAの分泌に関わるHasD領域の同定, 第3回日本生物工学会西日本支部講演会, Dec. 2016.
28.	Matsuda Shinya, Kohei Kawamoto, Mana Sawamoto, Akihiko Tsuji and Keizo Yuasa : PCTK3/CDK18による細胞形態・運動の制御, 第39回日本分子生物学会年会, Dec. 2016.
29.	Akihiko Tsuji, 桑村修司 and Keizo Yuasa : アメフラシ消化液より単離したフロロタンニン結合蛋白質，EHEPの特性解析, 第68回日本生物工学会, Sep. 2016.
30.	Shuhei Hamagaki, 高木大地, 市原拓巳, Akihiko Tsuji, 大森謙司 and Keizo Yuasa : Serratia marcescens type 1 secretion systemを用いた大腸菌による1本鎖抗体分泌系の構築, 第68回日本生物工学会, Sep. 2016.
31.	Jumpei Yamashita, 小出(吉田) 静代, 中西智美, 市原拓巳, Akihiko Tsuji, 大森謙司 and Keizo Yuasa : Serratia marcescens type 1 secretion systemによるHasA分泌機構の解明, 第68回日本生物工学会, Sep. 2016.
32.	Sara Murakami, Norio Kamemura, Hiroaki Komatsu, Akihiko Tsuji and Keizo Yuasa : PKGIIによるFGF/FGFR/ERK経路阻害機構の解明, 日本農芸化学会中四国支部第46回講演会, Sep. 2016.
33.	Matsuda Shinya, Kohei Kawamoto, Akihiko Tsuji and Keizo Yuasa : サイクリン依存性キナーゼPCTK3/CDK18による細胞運動調節機構の解明, 日本農芸化学会中四国支部第45回講演会, Jun. 2016.
34.	桑村修司, Keizo Yuasa and Akihiko Tsuji : アメフラシ消化液に含まれるフロロタンニン結合タンパク質の機能解析, 第57回日本生化学会中国・四国支部例会, May 2016.
35.	Hiroaki Komatsu, Norio Kamemura, Akihiko Tsuji and Keizo Yuasa : PKGIIはRaf1のSer43のリン酸化を介してMAPK経路を負に制御する, 日本農芸化学会2016年度大会, Mar. 2016.
36.	Keizo Yuasa, Kinuka Isshiki, Reina Ota, Matsuda Shinya, 雅広 Inoue and Akihiko Tsuji : 14-3-3から解離したDAPK2はチューブリンとの結合を介してアポトーシスを誘導する, 日本農芸化学会2016年度大会, Mar. 2016.
37.	Matsuda Shinya, Kenji Miyamoto, Akihiko Tsuji and Keizo Yuasa : PCTK3/CDK18はFAK1を抑制して細胞形態を制御する, 日本農芸化学会2016年度大会, Mar. 2016.
38.	Yurie Kawabata, Shuji Kuwamura, 澤田茉菜, Keizo Yuasa and Akihiko Tsuji : 褐藻類に含まれる摂食阻害物質に対する巻貝の戦略, 日本農芸化学会中四国支部第44回講演会, Jan. 2016.
39.	Shuji Kuwamura, 白石将孝, 佐藤仁昭, Keizo Yuasa and Akihiko Tsuji : アメフラシ消化液由来アラメ糖化促進タンパク質の特性解析, 第67回日本生物工学会, Oct. 2015.
40.	Hiroaki Komatsu, Matsuda Shinya, Akihiko Tsuji and Keizo Yuasa : タンパク質結晶構造解析に基づくPKGII の活性化機構の解明, 日本農芸化学会2015年度中四国・西日本支部合同大会(中四国支部第43回・西日本支部第312回講演), Sep. 2015.
41.	Chihiro Ito, 吉田一郎, Akihiko Tsuji, 矢中規之 and Keizo Yuasa : メラニン産生抑制効果を有する新規生薬成分の同定及びその作用機序の解明, 日本農芸化学会2015年度中四国・西日本支部合同大会(中四国支部第43回・西日本支部第312回講演), Sep. 2015.
42.	Koji Kishimoto, Takashi Hraguchi, Kenji Shimizu, Aiko Yamaguchi, Toshitada Yoshihara, Mikiko Kishi, Munenori Ide, Tetsunari Oyama, Yoshito Tsushima, Seishi Yobita, Akihiko Tsuji and Takashi Izumi : A GPCR, G2A, regulates stemness of cancer stem cells, 第56回日本生化学会中国四国支部例会(口頭), 16, May 2015.
43.	宮本賢治, 清水友紀, 松田真弥, Akihiko Tsuji and Keizo Yuasa : ナトリウム利尿ペプチド受容体Cの新規結合タンパク質同定, 日本農芸化学会2015年度大会, Mar. 2015.
44.	一色衣香, 平瀬大志, Akihiko Tsuji and Keizo Yuasa : Death-associated protein kinase-2 (DAPK2)と tubulinの相互作用解析及びアポトーシスへの関連性, 日本農芸化学会2015年度大会, Mar. 2015.
45.	Teruaki Ito, Yasuhiko Kawamura, Akihiko Tsuji, Masaki Hashizume and Toshihiro Moriga : Academic collaboration towards manufacturing system globalization, 日本機械学会生産システム部門研究発表講演会2015・講演論文集, Vol.15, No.8, 45-46, Mar. 2015. (Summary) TokushimaU-UTeM Academic Center (TMAC) has been successfully established in 2014 to promote the joint collaboration between the two institutions in terms of both education and research. One of the critical factors of this success was a good relationship between the two institutions based on the academic collaboration with alumni network in the past one decade. Presenting the overview of TMAC with its history and some of the critical on-going activities, this paper shows the feasibility of manufacturing system globalization through academic collaboration, and emphasized the importance of global engineer education. (Keyword) 大学間国際交流 / 教育研究交流 / 研究ユニット / 生産システム / 国際展開 (Link to Publication Site) ● Publication site (DOI): 10.1299/jsmemsd.2015.45 (Link to Search Site for Scientific Articles) ● CiNii @ National Institute of Informatics (CRID): 1390282680877479168 ● Search Scopus @ Elsevier (DOI): 10.1299/jsmemsd.2015.45 (DOI: 10.1299/jsmemsd.2015.45, CiNii: 1390282680877479168)
46.	佐藤仁昭, 河野剛士, Keizo Yuasa and Akihiko Tsuji : Subtilisin-like protein convertase(SPC)によるbone morphogenetic protein(BMP)のプロセシング, 第87回日本生化学会大会, Oct. 2014.
47.	太田玲奈, 松田真弥, 井上雅広, Akihiko Tsuji and Keizo Yuasa : 14-3-3は細胞死関連プロテインキナーゼDAPK2を負に制御する, 第87回日本生化学会大会, Oct. 2014.
48.	Keita Kinoshita, Toshihiro Kawauchi, Masaki Kubo, Keizo Yuasa and Akihiko Tsuji : Oligopeptidase Bのフォールディングと活性化機構の解明, 第87回日本生化学会大会, Oct. 2014.
49.	Jumpei Yamashita, 小出(吉田) 静代, 田中亮, 赤塚浩之, Akihiko Tsuji, 大森謙司 and Keizo Yuasa : S. marcescens type 1 secretion systemによるシグナルペプチド非依存的蛋白質分泌機構の解明, 第87回日本生化学会大会, Oct. 2014.
50.	松田真弥, 宮本賢治, 小松弘明, Akihiko Tsuji and Keizo Yuasa : PCTK3はアクチン動態を制御する, 日本農芸化学会2014年度中四国支部大会, Sep. 2014.
51.	白石将孝, 桑村修司, 大島美紀, 馬庭沙織, Keizo Yuasa and Akihiko Tsuji : アメフラシβ-グルコシダーゼのクローニング, 第66回日本生物工学会大会, Sep. 2014.
52.	桑村修司, 白石将孝, 大島美紀, 馬庭沙織, Keizo Yuasa and Akihiko Tsuji : アメフラシ消化液由来β-グルコシダーゼのラミナランの完全分解, 第66回日本生物工学会大会, Sep. 2014.
53.	白石将孝, 桑村修司, 馬庭沙織, 大島美紀, Keizo Yuasa and Akihiko Tsuji : アメフラシβ-グルコシダーゼの構造解析, 第55回日本生化学会中国・四国支部例会, Jun. 2014.
54.	桑村修司, 白石将孝, 大島美紀, 馬庭沙織, Keizo Yuasa and Akihiko Tsuji : アメフラシのラミナリン分解システム, 第55回日本生化学会中国・四国支部例会, Jun. 2014.
55.	松田真弥, 小湊恭平, 小出(吉田) 静代, 宮本賢治, Akihiko Tsuji and Keizo Yuasa : PCTK3はcyclin A及びPKAによって活性調節を受け，アクチン動態を制御する, 日本農芸化学会2014年度大会, Mar. 2014.
56.	伊藤千尋, 小出(吉田) 静代, Reger Albert, Akihiko Tsuji, Kim Choel and Keizo Yuasa : cGMP-dependent protein kinase II と小胞輸送制御因子 Rab11B との相互作用の解析, 日本農芸化学会2014年度大会, Mar. 2014.
57.	大島美紀, 西山奈見, 馬庭沙織, 桑村修司, 白石将孝, Keizo Yuasa and Akihiko Tsuji : アメフラシの海藻消化システムに関する研究, 第130回徳島生物学会, Dec. 2013.
58.	金惠珍, Atsushi Tabata, Toshifumi Tomoyasu, 上野友美, 内山成人, Keizo Yuasa, Akihiko Tsuji and Hideaki Nagamune : エストロゲン作用物質が示す骨芽細胞分化促進作用の解析, 徳島生物学会, Dec. 2013. (Keyword) エストロゲン / 骨芽細胞分化
59.	松田真弥, 小湊恭平, 小出(吉田) 静代, 宮本賢治, Akihiko Tsuji and Keizo Yuasa : サイクリン依存性キナーゼ18/PCTAIREキナーゼ3はサイクリンA2及びPKAによって活性化される, 第36回日本分子生物学会年会, Dec. 2013.
60.	Miki Oshima, Nami Nishiyama, Saori Maniwa, Keizo Yuasa and Akihiko Tsuji : アメフラシのα-グルカン分解機構, 第65回日本生物工学会大会, Sep. 2013.
61.	Saori Maniwa, Miki Oshima, Keizo Yuasa and Akihiko Tsuji : アメフラシのβ-グルコシダーゼの固定化と応用, 第65回日本生物工学会大会, Sep. 2013.
62.	Reina Ota, Shinya Matsuda, 雅広 Inoue, Akihiko Tsuji and Keizo Yuasa : 14-3-3による細胞死関連プロテインキナーゼDAPK2の活性制御機構, 第86回日本生化学会大会, Sep. 2013.
63.	Kyohei Kominato, Shinya Matsuda, Akihiko Tsuji and Keizo Yuasa : cyclin AおよびPKAによるCDKファミリーPCTK3の活性化機構, 第86回日本生化学会大会, Sep. 2013.
64.	松田真弥, 小湊恭平, 宮本賢治, Akihiko Tsuji and Keizo Yuasa : CDKファミリーPCTK3はcyclin AおよびPKAによって活性化する, 日本農芸化学会関西・中四国・西日本支部および日本ビタミン学会近畿・中国四国・九州沖縄地区合同大会2013年度合同広島大会, Sep. 2013.
65.	Akihiko Tsuji, Koichiro Hada, Kinuka Isshiki, Shinya Matsuda and Keizo Yuasa : プロセシングプロテアーゼ，Furinを特異的に阻害するalpha1-antitrypsin改変体の作成, 第13回日本蛋白質科学会年会, Jun. 2013.
66.	Kyohei Kominato, Shinya Matsuda, Akihiko Tsuji and Keizo Yuasa : CDKファミリーPCTK3の活性化機構の解明, 第54回日本生化学会中国・四国支部例会, May 2013.
67.	Reina Ota, Shinya Matsuda, 雅広 Inoue, Akihiko Tsuji and Keizo Yuasa : 14-3-3タンパク質によるDAPK2活性制御機構の解析, 第54回日本生化学会中国・四国支部例会, May 2013.
68.	Saori Maniwa, Miki Oshima, Keizo Yuasa and Akihiko Tsuji : アメフラシβ-グルコシダーゼの応用研究, 第54回日本生化学会中国・四国支部例会, May 2013.
69.	Miki Oshima, Nami Nishiyama, Saori Maniwa, Keizo Yuasa and Akihiko Tsuji : アメフラシのデンプン分解システム, 第54回日本生化学会中国・四国支部例会, May 2013.
70.	松田真弥, 篠倉悠久, Akihiko Tsuji and Keizo Yuasa : CDKファミリーPCTK3の活性化因子の同定, 日本農芸化学会2013年大会, Mar. 2013.
71.	一色衣香, Keizo Yuasa and Akihiko Tsuji : cGMP依存性プロテインキナーゼ(cGK)によるDeath-associated protein kinase-2 (DAPK2)調節機構, 第85回日本生化学大会, Dec. 2012.
72.	河野剛士, Keizo Yuasa and Akihiko Tsuji : 内軟骨性骨化過程におけるPACE4によるproBMP6の活性化, 第85回日本生化学大会, Dec. 2012.
73.	Junki Fukumoto, Ismaliza Ismail Nor Mohd, 雅広 Inoue, Keizo Yuasa and Akihiko Tsuji : Oligopeptidase Bにおけるβ-プロペラドメインと触媒ドメインの相互作用に重要な塩橋の同定, 第85回日本生化学大会, Dec. 2012.
74.	Hyejin Kim, Atsushi Tabata, Toshifumi Tomoyasu, Tomomi Ueno, Naruhito Uchiyama, Keizo Yuasa, Akihiko Tsuji and Hideaki Nagamune : A study on the mechanism of osteoblast differentiation induced by estrogen-stimuli, 日本生化学会, Dec. 2012. (Keyword) osteoblast / エストロゲン / イソフラボン
75.	西山奈美, 富永景子, Keizo Yuasa and Akihiko Tsuji : アメフラシ由来α-アミラーゼの精製と特性解析, 第64回日本生物工学会大会, Oct. 2012.
76.	植野勇司, 西山奈美, 富永景子, Keizo Yuasa and Akihiko Tsuji : アメフラシ由来β-グルコシダーゼの特性解析, 第64回日本生物工学会大会, Oct. 2012.
77.	富永景子, 西山奈美, Keizo Yuasa, Chizuru Sasaki, Yoshitoshi Nakamura and Akihiko Tsuji : アメフラシのセルロース消化システムの解析, 第64回日本生物工学会大会, Oct. 2012.
78.	Akihiko Tsuji, 福元淳生 and Keizo Yuasa : オリゴペプチダーゼBにおけるベータープロペラドメインと触媒ドメインの相互作用解析-触媒ドメインの安定化に重要な塩橋の同定, 第17回日本病態プロテアーゼ学会, Aug. 2012.
79.	Akihiko Tsuji, 佐藤しおり, 富永景子, Keizo Yuasa, Chizuru Sasaki and Yoshitoshi Nakamura : アミエビセルラーゼの基礎と応用研究, 日本生物工学会西日本支部第2回講演会, Jul. 2012.
80.	金惠珍, Atsushi Tabata, Toshifumi Tomoyasu, 上野友美, 内山成人, Keizo Yuasa, Akihiko Tsuji and Hideaki Nagamune : エストロゲン作用物質が示す骨芽細胞の分化促進作用, 第53回日本生化学会中国・四国支部例会, May 2012.
81.	西山奈見, 造田莉沙, 富永景子, Keizo Yuasa and Akihiko Tsuji : アメフラシ由来α-アミラーゼの精製と特性解析, 第53回日本生化学会中国・四国支部例会, May 2012.
82.	河野剛士, Keizo Yuasa and Akihiko Tsuji : 軟骨分化におけるPACE4の基質の同定, 第53回日本生化学会中国・四国支部例会, May 2012.
83.	一色衣香, Keizo Yuasa and Akihiko Tsuji : cGMP依存性プロテインキナーゼ(cGK)の新規基質同定, 第53回日本生化学会中国・四国支部例会, May 2012.
84.	須藤遥, 吉勝雄希, Keizo Yuasa, Akihiko Tsuji, 石田洋一 and Masami Nagahama : 核小体シャペロンNVL2によるリボソーム生合成制御機構;rRNA プロセシング複合体の分子間相互作用解析, 日本薬学会第132年会, Mar. 2012.
85.	播田元輝, Keizo Yuasa, 多田かおり, 藤本智美, 津嘉山正夫 and Akihiko Tsuji : スダチ果皮ポリメトキシフラボンの抗炎症作用に関する研究, 日本農芸化学会2012年度大会, Mar. 2012.
86.	一色衣香, Keizo Yuasa and Akihiko Tsuji : cGMP依存性プロテインキナーゼ(cGK)の新規基質同定, 日本農芸化学会2012年度大会, Mar. 2012.
87.	美野健, Yoshinori Fujisawa, Keizo Yuasa and Akihiko Tsuji : セリンプロテアーゼS9ファミリーに属する新規アミノペプチターゼの同定, 第63回日本生物工学会大会, Sep. 2011.
88.	富永景子, Chika Ikeda, 近藤あゆみ, 佐藤しおり, Keizo Yuasa and Akihiko Tsuji : アメフラシのセルロース消化システム解析, 第63回日本生物工学会大会, Sep. 2011.
89.	福元淳生, NOR MOHD BINTI ISMALIZA ISMAIL, Keizo Yuasa and Akihiko Tsuji : トリパノソーマ症の病原因子であるoligopeptidase Bのドメイン間相互作用の安定化機構, 第84回日本生化学大会, Sep. 2011.
90.	吉勝雄希, 石田洋一, Keizo Yuasa, Akihiko Tsuji and Masami Nagahama : rRNAプロセシング複合体の分子間相互作用におけるMPP6の役割, 第84回日本生化学大会, Sep. 2011.
91.	土肥真, Keizo Yuasa, Takeshi Nagame and Akihiko Tsuji : 神経細胞におけるcGMP依存性プロテインキナーゼとRhoエフェクターrhotekinの相互作用の解析, 第84回日本生化学大会, Sep. 2011.
92.	近藤あゆみ, Shiori Sato, Keiko Tominaga, 池田千佳, Keizo Yuasa and Akihiko Tsuji : サザエセルラーゼの酵素学的研究, 第84回日本生化学大会, Sep. 2011.
93.	佐藤しおり, Ayumi Kondo, Keiko Tominaga, 池田千佳, Keizo Yuasa and Akihiko Tsuji : アミエビセルラーゼの酵素学的研究, 第84回日本生化学大会, Sep. 2011.
94.	Akihiko Tsuji, Hiroshi Ishikawa and Keizo Yuasa : BMP活性化におけるPACE4とFurinの機能分担, 第16回日本病態プロテアーゼ学会, Aug. 2011.
95.	Junki Fukumoto, NOR MOHD BINTI ISMALIZA ISMAIL, Keizo Yuasa and Akihiko Tsuji : トリパノソーマ症の病原因子であるOligopeptidase Bのドメイン間相互作用の安定化機構, 第11回日本蛋白質科学会年会, Jun. 2011.
96.	Genki Harita, Keizo Yuasa, 津嘉山正夫 and Akihiko Tsuji : スダチ由来ポリフェノールsudachitinによる神経保護作用, 日本農芸化学会中四国支部第30回講演会, May 2011.
97.	土肥真, Keizo Yuasa, 長目健 and Akihiko Tsuji : cGMP依存性プロテインキナーゼとRhoエフェクターrhotekinの相互作用部位および細胞内局在の解析, 第52回日本生化学会中国・四国支部例会, May 2011.
98.	Keiko Tominaga, Chika Ikeda, Ayumi Kondo, 佐藤しおり, Keizo Yuasa and Akihiko Tsuji : アメフラシのセルロース消化システムの解析, 第52回日本生化学会中国・四国支部例会, May 2011.
99.	Takeru Mino, Yoshinori Fujisawa, Keizo Yuasa and Akihiko Tsuji : カイワレダイコンフェニルアラニルアミノぺプチダーゼの酵素特性解析及びクローニング, 第52回日本生化学会中国・四国支部例会, May 2011.
100.	Genki Harita, Keizo Yuasa, 津嘉山正夫 and Akihiko Tsuji : Effect of the polymethoxyflavone in citrus sudachi on neuroprotection, 日本農芸化学会2011年度大会, Mar. 2011.
101.	Keizo Yuasa, 松田泰斗 and Akihiko Tsuji : Functional regulation of transient receptor potential channel TRPC7 by cGMP-dependent protein kinase, 日本農芸化学会2011年度大会, Mar. 2011.
102.	Keizo Yuasa, 長目健, Makoto Dohi, Yayoi Yanagita, Masami Nagahama and Akihiko Tsuji : cGMP-dependent protein kinase/rhotekinによる神経突起形成の制御, 第33回日本分子生物学会年会・第83回日本生化学会大会合同大会, Dec. 2010.
103.	松田泰斗, Keizo Yuasa and Akihiko Tsuji : cGMP依存性プロテインキナーゼによる非選択的カチオンチャネルTRPC7の活性制御機構の解析, 第33回日本分子生物学会年会・第83回日本生化学会大会合同大会, Dec. 2010.
104.	港紗央里, 本木陽, Keizo Yuasa, Akihiko Tsuji and Masami Nagahama : VCP/p97結合タンパク質UBXD1 ( UBX domain-containing 1 ) の小胞体関連分解における機能解析, 第33回日本分子生物学会年会・第83回日本生化学会大会合同大会, Dec. 2010.
105.	本木陽, 港紗央里, 大西満智, Keizo Yuasa, Akihiko Tsuji and Masami Nagahama : 小胞体関連分解にかかわるVCP/p97結合タンパク質UBXD1のドメイン機能解析, 第33回日本分子生物学会年会・第83回日本生化学会大会合同大会, Dec. 2010.
106.	藤澤慶典, Akihiko Tsuji and Keizo Yuasa : S9セリンプロテアーゼに属する新規メンバー，フェニルアラニルアミノペプチダーゼの同定, 第33回日本分子生物学会年会・第83回日本生化学会大会合同大会, Dec. 2010.
107.	Akihiko Tsuji, 羽田浩一郎 and Keizo Yuasa : プロテインエンジニアリングによるFurin得意阻害タンパク質の作成, 第15回日本病態プロテアーゼ学会, Aug. 2010.
108.	Akihiko Tsuji, NOR MOHD BINTI ISMALIZA ISMAIL, Takeshi Nakai, Keizo Yuasa and 井上雅広 : モデリングより推定されたTrypanosoma bruceiオリゴペプチダーゼBのS1, S2サブサイトに位置するグルタミン酸残基の機能解析, 第51回日本生化学会中国・四国支部例会, May 2010.
109.	Keizo Yuasa, 長目健, 柳田弥生, Masami Nagahama and Akihiko Tsuji : 神経突起形成におけるcGMP依存性プロテインキナーゼの機能解析, 日本農芸化学会2010年度大会, Mar. 2010.
110.	松田泰斗, Keizo Yuasa, 北清水百合花 and Akihiko Tsuji : cGKIαによるTRPC7の調節機構, 日本農芸化学会2010年度大会, Mar. 2010.
111.	松田泰斗, Keizo Yuasa and Akihiko Tsuji : cGMP依存性プロテインキナーゼ(cGK)の新規基質の同定およびその機能解析, 第82回日本生化学大会, Oct. 2009.
112.	Keizo Yuasa, 長目健, 山上真, Masami Nagahama and Akihiko Tsuji : cGMP依存性プロテインキナーゼとRhoエフェクターrhotekinとの相互作用の解析, 第82回日本生化学大会, Oct. 2009.
113.	吉勝雄希, 谷田渚, 藤澤大輔, Keizo Yuasa, Akihiko Tsuji and Masami Nagahama : 核小体シャペロンNVL2によるエキソソーム複合体のrRNAプロセシングにおける機能制御, 第82回日本生化学大会, Oct. 2009.
114.	羽田浩一郎, Akihiko Tsuji and Keizo Yuasa : α1-Antitrypsinを用いたFurin特異的阻害剤の開発, 第82回日本生化学大会, Oct. 2009.
115.	塚本佳那, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : Germination specific cysteine protease 1 (GCP1)の酵素学的研究, 第82回日本生化学大会, Oct. 2009.
116.	岩本桂子, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : カイワレ大根の発芽初期過程で発現するプロテアーゼの解析, 第82回日本生化学大会, Oct. 2009.
117.	Ismaliza Ismail Nor Mohd, 湯浅剛, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : トリパノソーマオリゴペプチダーゼの熱安定性に重要なアミノ酸残基の同定, 第82回日本生化学大会, Oct. 2009.
118.	室下大樹, Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : BMP6の活性化を介したサチライシン様プロプロテインコンベルターゼPACE4による軟骨分化誘導, 第82回日本生化学大会, Oct. 2009.
119.	Akihiko Tsuji, Keizo Yuasa, Ismaliza Ismail Nor Mohd, 湯浅剛 and Masami Nagahama : トリパノソーマオリゴペプチダーゼの基質結合部位の機能解析, 第82回日本生化学大会シンポジウム「S9 セリンプロテアーゼの構造，機能から応用へ」, Oct. 2009.
120.	Akihiko Tsuji, Keizo Yuasa, 泰地裕美 and Masami Nagahama : 内軟骨性骨分化におけるBMP6の活性化機構, 第50回日本生化学会中国・四国支部例会, May 2009.
121.	Keizo Yuasa, 長目健, 山上真, Masami Nagahama and Akihiko Tsuji : cGMP依存性プロテインキナーゼ結合タンパク質の同定および機能解析, 日本農芸化学会2009年度大会, Mar. 2009.
122.	Akihiko Tsuji, 菊池弥生, 雑賀裕子, Keizo Yuasa and Masami Nagahama : オクルーディングループ構造の違いによる植物カテプシンBの特性, 第31回日本分子生物学会年会・第81回日本生化学大会合同大会, Dec. 2008.
123.	菊池弥生, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : 植物システインプロテアーゼRD21の精製と生化学的性質の解析, 第31回日本分子生物学会年会・第81回日本生化学大会合同大会, Dec. 2008.
124.	Keizo Yuasa, 山上真, 長目健, Masami Nagahama and Akihiko Tsuji : II型cGMP依存性プロテインキナーゼと小胞輸送制御因子Rab11bとの相互作用の解析, 第31回日本分子生物学会年会・第81回日本生化学大会合同大会, Dec. 2008.
125.	長目健, Keizo Yuasa, 山上真, Masami Nagahama and Akihiko Tsuji : cGMP依存性プロテインキナーゼとRhoエフェクターRhotekinとの相互作用の解析, 第31回日本分子生物学会年会・第81回日本生化学大会合同大会, Dec. 2008.
126.	Keizo Yuasa, 泰地裕美, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : 軟骨分化過程におけるサチライシン様プロプロテインコンベルターゼPACE4の機能解析, 第13回病態と治療におけるプロテアーゼとインヒビター学会, Aug. 2008.
127.	Masami Nagahama, machi Ohnishi, Keizo Yuasa and Akihiko Tsuji : 小胞体異常タンパク質分解におけるVCP/p97結合タンパク質UBXD1の役割, 第13回病態と治療におけるプロテアーゼとインヒビター学会, Aug. 2008.
128.	Akihiko Tsuji, Hiroko Saika, Yayoi Kikuchi, Keizo Yuasa and Masami Nagahama : 植物カテプシンB様システインプロテアーゼと動物カテプシンBの比較生化学, 第49回日本生化学会中国・四国支部例会, May 2008.
129.	Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : サチライシン様プロプロテインコンベルターゼPACE4は軟骨分化に必須である, 日本農芸化学会2008年度大会, Mar. 2008.
130.	大西満智, 河手裕美子, 松井隆幸, 三宅仁美, Keizo Yuasa, 谷佳津子, 多賀谷光男, Akihiko Tsuji and Masami Nagahama : UBXD1はVCP/97と結合しERADを制御する, 第80回日本生化学会大会, Dec. 2007.
131.	Kazuaki Muramatsu, 関光太, 鈴木龍介, 前野貴則, Akihiko Tsuji and Yoshiko Matsuda : オステオカルシンの成熟化に関与するプロセシング酵素の解析, 第80回日本生化学会大会, Dec. 2007.
132.	湯浅剛, 小田英里奈, Keizo Yuasa, Masami Nagahama, 井上雅広 and Akihiko Tsuji : Trypanosoma brucei oligopeptidase Bの基質結合部位の解析, 第80回日本生化学会大会, Dec. 2007.
133.	小川健太郎, 雑賀裕子, 林絢子, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : 植物カテプシンB様システインプロテアーゼの酵素学的特性, 第80回日本生化学会大会, Dec. 2007.
134.	柳野卓也, Keizo Yuasa, 木村道江, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : 軟骨分化におけるフィブリリン-2の機能解析, 第80回日本生化学会大会, Dec. 2007.
135.	山上真, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : cGMP依存性プロテインキナーゼと相互作用する因子の検索, 第80回日本生化学会大会, Dec. 2007.
136.	Keizo Yuasa, 影山陽子, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : ズブチリシン様プロプロテイン変換酵素PACE4は軟骨分化に必須である, 第80回日本生化学会大会, Dec. 2007.
137.	Akihiko Tsuji : SPCを選択的に阻害できるアンチトリプシン改変体の作成と応用, 第12回病態と治療におけるプロテアーゼとインヒビター学会, Aug. 2007.
138.	Akihiko Tsuji, 菊池弥生, Masami Nagahama, Keizo Yuasa and 井上英史 : サチライシン様プロプロテインコンベルターゼプロペプチドの機能, 第7回日本蛋白質科学会年会ワークショップ4(「分子内シャペロン」としてのプロペプチド研究:分子機構から生理機能まで), May 2007.
139.	Keizo Yuasa, 増田哲也, 吉川千尋, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : ズブチリシン様プロプロテイン変換酵素PACE4は骨格筋分化に必須である, 第48回日本生化学会中国・四国支部例会, May 2007.
140.	糟谷拓, Ismaliza Ismail Nor Mohd, 上川健一, 蟹江裕樹, Keizo Yuasa, Masami Nagahama, 助野晃子, 桑島正道 and Akihiko Tsuji : ヒト内臓・皮下脂肪組織におけるアドレノメジュリンとその活性化に必要なプロセシング酵素群の発現, 第48回日本生化学会中国・四国支部例会, May 2007.
141.	谷田渚, Keizo Yuasa, Akihiko Tsuji and Masami Nagahama : 核小体分子シャペロンNVL2とrRNAプロセシング複合体の相互作用解析, 第48回日本生化学会中国・四国支部例会, May 2007.
142.	上原正太郎, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : 軟骨分化過程におけるcGMP-dependent protein kinase IIの遺伝子発現調節機構, 日本分子生物学会2006フォーラム, Dec. 2006.
143.	柳野卓也, Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Transcriptional Regulation of Fibrilin-2 Gene during Chondrogenesis, 日本分子生物学会2006フォーラム, Dec. 2006.
144.	Ismaliza Ismail Nor Mohd, 蟹江裕樹, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : 脂肪細胞の分化過程におけるSPCファミリーの機能解析, 日本分子生物学会2006フォーラム, Dec. 2006.
145.	Syunji Miki, Keizo Yuasa, Akihiko Tsuji and Masami Nagahama : リボソーム生合成およびmRNA前駆体スプライシングにおけるAAA型分子シャペロンNVL2の機能, 第118回徳島生物学会, Nov. 2006.
146.	増田哲也, Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Subtilisin-like Proprotein Convertase (SPC)は骨格筋分化制御の重要因子である, 第11回病態と治療におけるプロテアーゼとインヒビター研究会, Aug. 2006.
147.	Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : 軟骨分化におけるsubtilisin-like proprotein convertase (SPC)の機能解析, 第11回病態と治療におけるプロテアーゼとインヒビター研究会, Aug. 2006.
148.	上川健一, Ismaliza Nor Ismail, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : 脂肪組織におけるプロセシングプロテア-ゼ群の機能解析, 第47回日本生化学会中国・四国支部例会, May 2006.
149.	蟹江裕樹, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : α1-Antitrypsinを用いたSPC特異的阻害剤の開発, 第6回日本蛋白質科学会年会, Apr. 2006.
150.	池田真理, 池内健, 志賀篤志, 幡野紘樹, 高橋俊昭, 原賢寿, Akihiko Tsuji, 西沢正豊 and 小野寺理 : 分泌型セリンプロテアーゼHtrA1とserpinとの高分子複合体形成の解析, 第28回日本分子生物学会年会, Dec. 2005.
151.	Keizo Yuasa, Yoshiko Matsuda and Akihiko Tsuji : 軟骨分化におけるSubtilisin-like proprotein convertase(SPC)の機能解析, 第28回日本分子生物学会年会ワークショップ, Dec. 2005.
152.	Takuya Yanagino, Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Transcriptional regulation of fibrillin-2 during chondrogenesis, 第78回日本生化学会大会, Oct. 2005.
153.	Tetsuya Masuda, Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : Transcriptional activation of proprotein convertase PACE4 during skeletal muscle differentiation, 第78回日本生化学会大会, Oct. 2005.
154.	Akihiko Tsuji, Kentaro Ogawa, Tsuyoshi Yuasa, Keizo Yuasa, Masami Nagahama and Tadashi Yoshimoto : Protamine: a unique and potent inhibitor of oligopeptidase B, 第78回日本生化学会大会, Oct. 2005.
155.	Kentaro Ogawa, Keizo Yuasa, Masami Nagahama and Akihiko Tsuji : Expression of cysteine proteases and oligopeptidase B in early seedling growth of wheat, 第78回日本生化学会大会, Oct. 2005.
156.	Yayoi Kikuchi, Keizo Yuasa, Masami Nagahama, Yoshiko Matsuda and Akihiko Tsuji : The role of Reticulocalbin-3 in regulation of PACE4 activation and secretion, 第78回日本生化学会大会, Oct. 2005.
157.	Akihiko Tsuji, Keizo Yuasa and Yoshiko Matsuda : サチライシン様プロプロテインコンベルターゼのユニークな自己活性化制御機構, 第5回日本蛋白質科学会年会ワークショップ9, Jun. 2005.
158.	Keizo Yuasa, Akihiko Tsuji and Yoshiko Matsuda : 骨格筋分化におけるプロセシングプロテアーゼPACE4の遺伝子発現調節の解析, 日本農芸化学会2005年度大会, Mar. 2005.
159.	Akihiko Tsuji, Keizo Yuasa and Yoshiko Matsuda : 哺乳類ズブチリシン様セリンプロテアーゼの特性，多様性と生理的意義, 日本農芸化学会2005年度大会ワークショップ, Mar. 2005.
160.	鈴江加織, Keizo Yuasa, Akihiko Tsuji and Yoshiko Matsuda : E2F1によるPACE4遺伝子発現調節の解析, 第27回日本分子生物学会年会, Dec. 2004.
161.	Keizo Yuasa, Kaori Suzue, Akihiko Tsuji and Yoshiko Matsuda : Transcriptional activation of proprotein convertase PACE4 by E2F1, 第77回日本生化学会大会, Oct. 2004.
162.	Akihiko Tsuji, Keizo Yuasa and Yoshiko Matsuda : Purification and characterization of oligopeptidase B from wheat germ. First identification of oligopeptidase B at the protein level in higer plant, 第77回日本生化学会大会, Oct. 2004.
163.	Yoshiko Matsuda, Akihiko Tsuji and Keizo Yuasa : Subtilisin like proprotein convertase familyの多様性と役割分担:ECM局在PACE4の細胞分化における役割, 第51回マトリックス研究会大会, 20-21, Apr. 2004.
164.	牧瀬広孝, Akihiko Tsuji and Yoshiko Matsuda : α1-アンチトリプシン変異体を用いたSPC ファミリー基質特異性の解析, 第26回日本分子生物学会年会, Dec. 2003.
165.	Akihiko Tsuji, 菊池弥生 and Yoshiko Matsuda : ズブチリシン様プロプロテインコンベルターゼ生合成におけるEF-Hand Ca結合タンパク，Reticulocalbin-like Protein (RLP)の機能同定, 第26回日本分子生物学会, Dec. 2003.
166.	Yayoi Kikuchi, Yukimi Sato, Akihiko Tsuji and Yoshiko Matsuda : Reticulocabin: A novel protein which regulates biosynthesis of subtilisin-like proprotein convertase, 第76回日本生化学会大会, Oct. 2003.
167.	Yoshiko Matsuda and Akihiko Tsuji : A unique 26-kDa cysteine protease induced in wheat embryo during germination, 第76回日本生化学会大会, Oct. 2003.
168.	Akihiko Tsuji, Hirotaka Makise and Yoshiko Matsuda : Antitrypsin mutants: Tool for functional analysis of serine-endopeptidases, 第76回日本生化学会大会, Oct. 2003.
169.	Akihiko Tsuji : 蛋白質の分析技術, 第40回分析化学講習会, 49-54, Aug. 2003.
170.	Akihiko Tsuji, 佐藤幸美 and Yoshiko Matsuda : PACE4生合成におけるレティキュロカビンの機能解析—シャペロンとしての役割—, 第3回日本蛋白質科学会年会, Jun. 2003.
171.	二松豪, 嶋田知訓, Akihiko Tsuji and Yoshiko Matsuda : ATDC5細胞の内軟骨分化過程におけるプロセシングプロテアーゼPACE4の機能解析, 第25回日本分子生物学会年会, Dec. 2002.
172.	Akihiko Tsuji, 生駒貴之, 橋本英美, 山下大 and Yoshiko Matsuda : ズブチリシン様プロプロテインコンベルターゼ(SPC)に特異阻害活性を持つα1-アンチトリプシン改変体の作製と機能解析への応用, 平成14年度日本生物工学会大会, Oct. 2002.
173.	佐藤幸美, 田頭和幸, Akihiko Tsuji and Yoshiko Matsuda : プロテオーム解析によるSPCファミリーの基質タンパクの検索:Ca結合タンパクReticulocabinについて, 第75回日本生化学会大会, Oct. 2002.
174.	田中幸子, Akihiko Tsuji and Yoshiko Matsuda : ズブチリシン様プロプロテインコンヴェルターゼ(SPC)に選択的阻害活性を有するα1アンチトリプシン改変体の作成と応用, 第75回日本生化学会大会, Oct. 2002.
175.	高橋保充, 櫻井謙介, Akihiko Tsuji and Yoshiko Matsuda : 細胞外マトリックスにおけるズブチリシン様プロセシングプロテアーゼPACE4の役割, 第75回日本生化学会大会, Oct. 2002.
176.	小出静代, Kazunori Toida, Kazunori Ishimura, Akihiko Tsuji and Yoshiko Matsuda : 胎盤特異的bHLH型転写因子MASH-2によるPACE4の胎盤発生過程における制御, 第75回日本生化学会大会, Oct. 2002.
177.	Miwa Bando, 山下大, Hiroshi Kido, Akihiko Tsuji and Yoshiko Matsuda : 巨核球分化におけるプロセシングプロテアーゼPACE4とfurinの転写調節機構, Seikagaku, Vol.74, No.8, 1016, Oct. 2002.
178.	Akihiko Tsuji, 生駒貴之, 橋本英美, 山下大 and Yoshiko Matsuda : ズブチリシン様プロプロテインコンベルターゼに選択的阻害作用を有するアンチトリプシン改変体の開発, 第2回日本蛋白質科学会年会, Jun. 2002.
179.	吉田聡子, Kikuji Yamashita, Akihiko Tsuji, Seiichiro Kitamura and Yoshiko Matsuda : 造血及び間質幹細胞培養の開発, 徳島生物学第110回総会, 110, 2002.
180.	小出静代, 清蔭恵美, 藤原良次, 川並由季, Akihiko Tsuji and Yoshiko Matsuda : ヒト胎盤でのPACE4遺伝子発現と生理機能の解析, 第24回日本分子生物学会年会, Dec. 2001.
181.	吉田一郎, 小出静代, Kazunori Toida, Kazunori Ishimura, Akihiko Tsuji and Yoshiko Matsuda : 神経特異的bHLH型転写因子Mash-1による前駆体タンパク質活性化酵素PACE4の神経発生過程における制御, 第24回日本分子生物学会年会, Dec. 2001.
182.	嶋田知訓, Miwa Bando, 開裕司, 宿南知佐, Akihiko Tsuji and Yoshiko Matsuda : ATDC5細胞の軟骨分化過程におけるSPCファミリーの動態, 第74回日本生化学会大会, Oct. 2001.
183.	Akihiko Tsuji, 生駒貴之 and Yoshiko Matsuda : サブチリシン様プロプロテインコンベルターゼ，PACE4, Furin, PC6に対する選択的阻害作用を持つアンチトリプシン改変体の作成, 第74回日本生化学会大会, Oct. 2001.
184.	Miwa Bando, 嶋田知訓, Akihiko Tsuji and Yoshiko Matsuda : 巨核球分化におけるプロセシングプロテアーゼPACE4の生理機能の解析, 第74回日本生化学会大会, Oct. 2001.
185.	Akihiko Tsuji : サチライシン様プロセシングプロテアーゼの発現制御と機能, 第8回長崎大学バイオフォーラム, Aug. 2001.
186.	櫻井謙介, 谷口孝純, Akihiko Tsuji and Yoshiko Matsuda : プロセシングプロテアーゼPACE4の活性発現と細胞外での機能の解析, 第53回日本細胞生物学会, Oct. 2000.
187.	櫻井謙介, 谷口孝純, 清蔭恵美, Akihiko Tsuji and Yoshiko Matsuda : 細胞外マトリックスにおけるヒト前駆体タンパク質プロセシングプロテアーゼPACE4の機能解析, 第73回日本生化学会大会, Oct. 2000.
188.	松岡厚志, Miwa Bando, 平山幸子, Akihiko Tsuji and Yoshiko Matsuda : ズブチリシン様前駆体タンパク質プロセシングプロテアーゼPACE4アイソフォーム:PACE4Cの生理的役割, 第73回日本生化学会大会, Oct. 2000.
189.	吉田一郎, 小出静代, 中川原章, Akihiko Tsuji and Yoshiko Matsuda : 神経芽細胞腫におけるbHLH型転写因子hASH-1による前駆体タンパク質活性化酵素PACE4遺伝子の発現制御, 第73回日本生化学会大会, Oct. 2000.
190.	Tetsuya Akamatsu, 吉田茂, Hideaki Nagamune, Akihiko Tsuji and Yoshiko Matsuda : ラット脳発生過程でのプロセシング酵素PACE4の発現, 日本生化学会第70回大会, Sep. 1997.
191.	Tetsuya Akamatsu, 吉田茂, 大黒成夫, Hideaki Nagamune, 長濱正巳, Akihiko Tsuji and Yoshiko Matsuda : プロセシング酵素PACE4Eのラット脳発生過程での発現, 日本生化学会第69回大会・日本分子生物学会第19回年会合同年会, Aug. 1996.
192.	Tetsuya Akamatsu, 吉田茂, 水野勝弘, 長田香弥, 米本佳余子, 森健二, Hideaki Nagamune, Akihiko Tsuji, Eric Grouzmann and Yoshiko Matsuda : Kexin様プロテアーゼPC7の中枢神経系における細胞特異的発現, 日本生化学会第68回大会, Sep. 1995.
193.	Akihiko Tsuji, Hideaki Nagamune, Tetsuya Akamatsu, 日根智恵美, 村松和明, 森健二, 玉井靖啓, 米本佳余子 and Yoshiko Matsuda : 新Kexin様プロテアーゼの構造と組織分布, 日本先天代謝異常学会第36回大会, Nov. 1994.
194.	Tetsuya Akamatsu, 今村恵美, Hideaki Nagamune, Akihiko Tsuji and Yoshiko Matsuda : 新しいkexinファミリープロテアーゼPC7の細胞特異的発現-in situ hybridizationによる解析, 日本細胞生物学会第47回大会, Sep. 1994.
195.	玉井靖啓, 村松和明, Tetsuya Akamatsu, 日根智恵美, 森健二, Hideaki Nagamune, Akihiko Tsuji, Keisuke Izumi and Yoshiko Matsuda : 新ペプチドホルモンプロセッシングプロテアーゼPACE4Cの細胞特異的発現, 日本細胞生物学会第47回大会, Sep. 1994.
196.	Akihiko Tsuji : DNAと蛋白質の微量定量法, 第40回分析化学講習会, 49-54, May 1994.
197.	Akihiko Tsuji : α₂ マクログロブリン-酵素複合体:リソゾーム病の酵素補充療法, 第9回タンパク質ハイブリッド研究会, May 1994.
198.	Hideaki Nagamune, Akihiko Tsuji, 村松和明, Tetsuya Akamatsu, 東根一也, 日根智恵美, Keisuke Izumi, 坂口剛正, 吉田哲也, Sumihare Noji and Yoshiko Matsuda : 甲状腺傍濾胞細胞におけるサチラーゼファミリープロテアーゼの発現, 日本生化学会第66回大会, Oct. 1993.
199.	Tetsuya Akamatsu, Akihiko Tsuji, Hideaki Nagamune and Yoshiko Matsuda : ラット肝α-マクログロブリン-プロテアーゼ複合体の性質, 日本生化学会第65回大会, Oct. 1992.
200.	Akihiko Tsuji, Tetsuya Akamatsu, 尾田領一, Hideaki Nagamune and Yoshiko Matsuda : ラット肝α-マクログロブリン-プロテアーゼ複合体の性質, 日本生化学会中四国支部第55回例会, Apr. 1992.
201.	Akihiko Tsuji : 細胞膜結合プロテアーゼ - ヘプシン, 平成2年度筑波大学遺伝子実験センター「バイオテクノロジーレクチャーコース」, Aug. 1990.

Et cetera, Workshop:

1.	Hirokazu Miyoshi, Hiroshi Maezawa, Akio Adachi, Takuya Saze, Namiko Irikura, Yoshinori Kuwahara, 立花さやか, Tatsuhiro Ishida, Akihiko Tsuji, Takao Hanabusa, Toshihiro Moriga, 鬼島明洋, Eiichi Honda, Seiji Iwamoto, 佐藤一雄, Shunsuke Furutani and Yasuhiko Morita : 徳島大学アイソトープ総合センターニュース, 徳島大学アイソトープ総合センターニュース, Vol.6, 1-42, Mar. 2007.

Patent:

1.	Yoshiko Matsuda and Akihiko Tsuji : ズブチリシン様プロプロテインコンベルターゼ·PACE4に対するモノクローナル抗体及びその利用, 2002-292869 (Oct. 2002), .

Grants-in-Aid for Scientific Research (KAKEN Grants Database @ NII.ac.jp)

海藻アラメからエタノールと食品機能成分を生産する新技術開発と海藻産業への応用研究 (Project/Area Number: 17K07936 )
Development of enzymatic saccharification method for complete utilization of seaweed biomass (Project/Area Number: 26450284 )
Production of useful products from cellulosic biomass usingultra-high temperature and pressure steam explosion (Project/Area Number: 22310048 )
プロセシングプロテアーゼ,PACE4(SPC4)の発現調節と細胞機能制御 (Project/Area Number: 11144226 )
Structure and Regulation of Processing Protease SPC4, and Identification of Its Physiological Substrates (Project/Area Number: 10680615 )
プロセシングプロテアーゼ,SPC4(PACE4)の発現調節と細胞機能制御 (Project/Area Number: 10163226 )
Physiological function of proprotein convertase PACE4 (Project/Area Number: 09670129 )
ケキシンファミリー,PACE4アイソフォームの生成機構と細胞機能制御 (Project/Area Number: 09267223 )
生理活性ペプチドプロセシングブロテアーゼ・PACE4アイソフォームの発現調節 (Project/Area Number: 08680690 )
ケキシンファミリー,PACE4アイソフォームの生成機構と細胞機構制御 (Project/Area Number: 08278221 )
生理活性ペプチドプロセシングプロテアーゼ・PACE4の機能 (Project/Area Number: 07680689 )
新Kex2様プロテアーゼの同定と構造及びその生理機能の研究 (Project/Area Number: 06680617 )
前駆体タンパク質の小胞体ゴルジ系におけるプロセシングの細胞特異性の研究 (Project/Area Number: 06248102 )
炎症時のαマクログログリン-肥満細胞トリプターゼ複合体の機能 (Project/Area Number: 05807011 )
膜融合タンパク質の活性化とウイルス感染のトポロジー (Project/Area Number: 05670143 )
ウイルス膜融合タンパク質の宿主細胞内輸送とトランスゴルジ領域での活性化機構 (Project/Area Number: 05252102 )
宿主細胞によるウイルスタンパク質活性化機構 (Project/Area Number: 04670163 )
ゴルジ領域に於けるウイルス膜融合タンパク質のブロシングとその生理的意義 (Project/Area Number: 04259105 )
ゴルジ膜結合プロセシングプロテア-ゼの生理的意義 (Project/Area Number: 03670143 )
肝ゴルジ膜結合プロテア-ゼのアルブミン及びウィルスタンパク生合成における役割 (Project/Area Number: 02670123 )
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Professor Emeritus : Tsuji, Akihiko

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Grants-in-Aid for Scientific Research (KAKEN Grants Database @ NII.ac.jp)