type: misc

label: url

en: Protein Crystal Growth Simply by Concentration

ja: 濃縮のみによるタンパク質結晶化

en: The three-dimensional (3D) structure of a protein molecule is usually solved by using crystals grown with copious amounts of precipitants. The obtained structure could be significantly different from those in vivo because the precipitant concentration in vivo is much less than those under normal crystallization conditions. For the first time, we have developed novel precipitant-free protein crystallization methods by centrifugal concentration and drying. For both methods, atomic-level 3D structures were successfully obtained. 3D molecular structure of hen egg-white lysozyme (HEWL) obtained without precipitants was significantly different from that obtained with precipitants, whereas the precipitant-free molecular structure of glucose isomerase (GI) was similar to those obtained with precipitants.

en: Journal of the Japanese Association for Crystal Growth

ja: 日本結晶成長学会誌


en: 圧力によって深まる溶解現象の本質の理解

ja: 圧力によって深まる溶解現象の本質の理解

en: 溶解現象は，様々な場面で重要な意味を持つことが多いが，本質的なところはまだ未解明な部分が多い．ここでは特に溶解に伴う体積変化と，そのメカニズムを解明するために有用な圧力の活用について，基本的なところから，創薬の分野等で必要となる高品質タンパク質結晶化に渡る部分まで紹介する．

en: Chemistry and Education

ja: 化学と教育


en: Effects of Gravity on the Growth Process of Colloidal Crystals with Depletion Force Mediated Attractive Interaction

ja: 枯渇凝集引力系コロイド結晶の成長プロセスに及ぼす重力効果

en: Although depletion-induced attractive colloidal crystallization processes are recently studied as a model system of crystal growth because of the size of crystal unit (more than several hundred micro meters), effects of gravity would be significant, since the size and density of particles are large, and their crystallization usually starts at the bottom of growth containers. In this study we successfully observed in situ adsorption, desorption, surface diffusion, and kink incorporation of particles on growth interfaces of the depletion-induced attractive colloidal crystals of polystyrene particles in aqueous sodium polyacrylate solutions under several gravity conditions during parabolic flights. Precise surface transportation and kink incorporation processes of the particles into the colloidal crystals with attractive interactions were observed in situ at the particle level. Surface diffusion coefficients of particles " were roughly estimated. " at 0 G became larger than those at 1 G. " at 0 G also showed scattered data compared to those at 1 G, since particles which moved largely had a propensity to increase under microgravity conditions. Obtained " values were 1 ~ 2 orders of magnitude smaller than those of bulk diffusion coefficients.

en: Space Utilization Research

ja: Space Utilization Research


en: Impurity Effects on the Step Velocity of a Spiral Hillock on a Crystal under Perfect Stagnant Conditions

ja: 完全対流停止条件における結晶のらせん成長ステップの前進速度に及ぼす不純物効果

en: Effects of gravity on step velocities step of hen egg-white lysozyme (HEWL) tetragonal crystals are studied. Parabolic flight experiments revealed that step in <110> directions of the crystals have a tendency to increase with a decrease in gravity. This increase is mainly due to the decrease in convection flow rates, since the decrease would result in the decrease in the transportation rates of covalently bound dimer of HEWL molecules (major impurity of this study) onto the growth interface of the crystals.


en: In-situ observation of colloidal crystallization

en: We report contents of colloidal crystallization experiments in ISSCG-16. The participants produce two kinds of colloidal crystals, i.e., close-packed opal type crystals, and non-close-packed charged colloidal crystals. They can observe the crystallization processes by using optical microscopy and reflection spectroscopy.

en: Progress in Crystal Growth and Characterization of Materials


en: Hydration / Dehydration Processes from the Viewpoints of Effects of Pressure on Solubilities and Growth Rates of Crystals

ja: 溶解度・結晶成長速度の圧力依存性から見る水和・脱水和プロセス

en: Precise understandings of hydration and dehydration processes in crystallization processes are prerequisites for deeper insights into fundamentals of crystal growth in aqueous solutions. In this review, we present the great potentialities of thermodynamic and kinetic analyses of the dependence of solubilities and crystal-growth-rates on pressure on experimentally accessible information of hydration and hydration processes in crystallization processes. The dependence of solubility of crystals on pressure provides molar volume change of dissolution. Positive and negative slopes of solubility curves with pressure result in the decrease and increase in the molar volume of solutes during dissolution processes, respectively. Activation volumes during solute incorporation processes at kink sites are calculated using the dependence of step kinetic constant on pressure. In practice, solubilities of alkali halide, amino acid, and protein crystals, and step velocity of glucose isomerase crystals are discussed.


en: Step Velocities of Protein Crystals in Ground-based Convection Free Experiments

ja: 地上対流抑制実験におけるタンパク質結晶のステップ前進速度

en: Convection flows induced by formation of solute depletion zone around growing protein crystals are visualized to clarify the reason why step velocities of a hen egg-white lysozyme (HEWL) tetragonal crystal grown at a ceiling position (top wall of a growth container) are not different from those at a sitting position (bottom wall of the container). Parabolic flight experiments revealed that flow rates of polystyrene particles (as marker particles) around a ceiling crystal do not indicate zero at hyper gravity conditions, and the particles completely stop at zero gravity, although the flow rates show slightly less values than those around a sitting crystal. Ceiling experiments cannot reproduce the effective suppression of convection flows around growing protein crystals in microgravity at this stage. Thus microgravity experiments are still indispensable to attain complete convection free conditions.


en: Change in interface growth kinetics and interface morphology of a protein crystal

ja: 溶液相の状態変化によるタンパク質結晶の界面成長kineticsおよび界面morphologyの変化

en: We focus on the growth mechanism of protein crystals and their perfection. The growth mechanism will be studied by surface observation and by measuring growth rates. The crystal defect will be studied by X-ray topography. We successfully carried out Nanostep project as an international space station (ISS) experiment in 2012 using lysozyme crystals as model protein crystals. From the results of the Nanostep project, we concluded that larger-size impurities are effectively filtered during crystallization owing to smaller diffusion constants of the larger impurities, and this filtration mechanism results in the increase in growth rate of lysozyme crystals in microgravity. However, from this assumption, we can easily anticipate that same-size impurities will not lead the faster growth rates, and smaller impurities will lead more suppression of growth rates in microgravity. To confirm these anticipations, a new ISS experiment is planned in near future using glucose isomerase (GI) crystals. We need to obtain ground based data for the comparison with the Nanostep data. Until now, we successfully confirm regrowth of GI crystals on chemically fixed seed crystals, conduct X-ray topography of regrown crystals on the seed crystals, and observe elementary steps on ceiling lysozyme crystals.


en: Exact Measument of Crystal Growth Rate of Lysozyme on The Ground and in ``Kobo''

ja: 地上とKiboで精密測定されたリゾチーム結晶成長速度

en: Growth rate of lysozyme crystals vs supersaturation has exactly been measured for the first time by means of laser interferometry at Kibo in odder to analyze the optimum condition and/or the growth mechanism for good quality protein crystals. The growth rate and shape of spiral growth hillocks were compared as obtained in gravity using the same growth cells. We here found the larger growth rate under microgravity than the rate in gravity in the wide range of supersaturation, which has commonly been used for growing crystals in gravity. The higher growth rate in space is attributed to the suppression of the adsorption of dimer-molecules that are present as impurity molecules, which block step movement along specific crystallographic directions depending on supersaturation. The lack of 2D heterogeneous nucleation growth regime in space is also due to the reduction of this impurity concentration at the growth surface. These effects would eventually widen the growth regime that is suitable for getting high quality protein crystals.


en: Effects of Microgravity on Crystallizaion of Glucose Isomerase --- Collaborative Studies toward the Proposal for Nanostep 2 Project

ja: グルコースイソメラーゼの結晶化に及ぼす微小重力効果 --- Nanostep 2 プロジェクト提案に向けた共同研究

en: Pressure is an attractive thermodynamic parameter. In this review, I present the great potentialities of high pressure for the promotion of studies on the enhancement of protein crystallization processes and correlation between functions and 3D structures of protein molecules. As a tool for enhancing the crystal growth, three of eight proteins show the decrease in its solubility under high pressure. Acceleration of growth and nucleation kinetics of glucose isomerase crystals occurred under high pressure. Step velocities under high pressure provided us direct information on activation volumes during incorporation processes of protein molecules at a kink site of crystals. Such activation volumes were negative in the case of glucose isomerase crystals. Precise discussion on the activation volume will be useful for understanding dehydration mechanisms during the incorporation processes. Usefulness of our standalone-type Be vessel for high-pressure protein crystallography was confirmed. With the vessel, precise high-pressure 3D structure analysis of protein crystals which are also grown under high pressure will be achieved.

en: International Journal of Microgravity Science and Application


en: Pressure effects on crystal growth of proteins

ja: 圧力を使ったタンパク質の結晶成長の研究

ja: Pressure is an attractive thermodynamic parameter. In this review, I present the great potentialities of high pressure for the promotion of studies on the enhancement of protein crystallization processes and correlation between functions and 3D structures of protein molecules. As a tool for enhancing the crystal growth, three of eight proteins show the decrease in its solubility under high pressure. Acceleration of growth and nucleation kinetics of glucose isomerase crystals occurred under high pressure. Step velocities under high pressure provided us direct information on activation volumes during incorporation processes of protein molecules at a kink site of crystals. Such activation volumes were negative in the case of glucose isomerase crystals. Precise discussion on the activation volume will be useful for understanding dehydration mechanisms during the incorporation processes. Usefulness of our standalone-type Be vessel for high-pressure protein crystallography was confirmed. With the vessel, precise high-pressure 3D structure analysis of protein crystals which are also grown under high pressure will be achieved.

en: Journal of the Japanese Association for Crystal Growth


en: X-ray structure analysis of glucose isomerase molecules in crystals prepared under high pressure

ja: 高圧力下で成長したグルコースイソメラーゼ結晶の常圧下でのX線結晶構造解析

en: 高圧バイオサイエンスとバイオテクノロジー 第15回生物関連高圧研究会20周年記念シンポジウム抄録集


en: In Situ Observation of Molecular Steps on the Surface of Protein Crystal under High Pressure

ja: 高圧力下における蛋白質結晶表面の分子ステップのその場観察

en: Velocities of elementary steps and two-dimensional nucleation rates on the {101} face of glucose isomerase (GI) crystals were measured at pressures of 0.1, 25 and 50 MPa. We conducted in situ observation of the steps by using a laser confocal microscope combined with differential interference contrast microscope (LCM-DIM) and high-pressure vessel with a sapphire window (t = 1 mm). Both step velocities and 2D nucleation rates increased with increasing pressure at T = 26.4°C and C = 5.6 mg ml-1. Solubilities Ce at high pressures were also measured by in situ observation of the moving steps around equilibrium temperatures. The solubility decreased with increasing pressure.

en: The Review of High Pressure Science and Technology

ja: 高圧力の科学と技術


en: 圧力印加による酵素の結晶成長の促進

ja: 圧力印加による酵素の結晶成長の促進

en: 食品と容器

ja: 食品と容器


en: タンパク質の結晶化に及ぼす圧力効果

ja: タンパク質の結晶化に及ぼす圧力効果

en: 静水圧はタンパク質の結晶化を制御するのに有用なパラメタである．タンパク質結晶の溶解度は，圧力を変化させることで急速に変化させることが可能である．タンパク質分子の特異的な表面に対する水和に関する情報は，溶解度の圧力依存性から得られる．タンパク質分子が疎水的であるか，その溶液中の分子体積が結晶中よりも十分大きい時，圧力印加は，結晶化を促進する効果的な手法となる．圧力はまた，タンパク質結晶の成長kineticsにも影響を及ぼす．典型的なタンパク質について，成長速度の結晶化駆動力に対する依存性を測定し，二次元核成長の多核成長様式を用いて解析した．成長kineticsに及ぼす高圧力効果は，結晶の表面エネルギー，分子が結晶に取り込まれる過程における活性化エネルギー，そして結晶表面のステップ上の平均キンク間距離の変化で説明出来る．しかし，成長kineticsに及ぼす圧力効果に対して，一般的·普遍的な解釈はまだ得られてはいない．

en: 日本結晶学会誌

ja: 日本結晶学会誌


en: A Novel Approach to Protein Crystallization : Use of a Magnetic Field and High-Pressure

ja: タンパク質結晶化への新たなアプローチ : 磁場および高圧力の利用

en: A magnetic field and high-pressure are effective parameters to control protein crystallization. A magnetic field significantly affects the number, orientation and growth rate (morphology) of protein crystals and damps the buoyancy convection in protein solutions. Quality of protein crystals can be also improved by a magnetic field. The improvement in the crystal quality is due to a magnetic orientation effect. High-pressure can also give large effects on the solubility and growth kinetics of protein crystals. Changes in the solubility with pressure results from a difference in the volume between bulk water and the water hydrating the contact regions of protein molecules inside the crystal.

ja: 磁場と高圧力は，タンパク質の結晶化を制御する有効なパラメタである．磁場は，タンパク質結晶の個数·方位·成長速度(モルフォロジー)に影響を及ぼし，タンパク質溶液中の浮力に伴う対流を抑える効果がある．タンパク質結晶の品質も磁場によって改善されうる．結晶品質の向上は，磁気配向効果によるものである．高圧力も溶解度や成長kineticsに大きな影響を及ぼす．溶解度の加圧に伴う変化は，タンパク質分子が結晶中で結合している部位に水和している水とバルク水の間の体積変化による．

en: Seibutsu Butsuri

ja: 生物物理


en: Protein Crystallization under High Hydrostatic Pressure

ja: 高圧力下におけるタンパク質の結晶化

en: In this article, recent advances in the fundamental research in the protein crystallization under high pressure were reviewed. Pressure is expected to be an important parameter to control protein crystallization, since hydrostatic pressure affects the whole system uniformly and can be changed very rapidly. So far, a lot of studies of protein crystallization under high pressure have been done, some less systematic than others. We have particularly focused on fundamental aspects (the solubility and the crystal growth kinetics) of protein crystallization under high pressure. The studies reviewed contribute significantly to high-pressure crystallography and high-pressure protein science.

ja: 高圧力下におけるタンパク質結晶化に関する基礎的研究の近年の発展について述べた．静水圧は系の中に均一に伝播し，かつ非常に迅速に変化出来るため，タンパク質結晶化において重要なパラメタである．これまで，高圧力下でタンパク質結晶化についての研究が多く行われてきたが，いずれも他のタンパク質結晶化の研究に比べ系統的ではなかった．本解説では，特に溶解度や結晶成長速度論といった基礎的な側面に注目した．溶解度の圧力依存性から溶解エンタルピーや溶解エントロピーの圧力依存性や，溶解に伴うモル体積変化といった情報が明らかになった．また，結晶成長速度の速度論的解析によって，分子表面自由エネルギーの圧力依存性を明らかにすることができた．ここで紹介する研究結果は高圧力下結晶構造解析や高圧力下タンパク質科学に重要な貢献をできるであろう．


en: Protein crystallization under high pressure

en: Pressure is expected to be an important parameter to control protein crystallization, since hydrostatic pressure affects the whole system uniformly and can be changed very rapidly. So far, a lot of studies on protein crystallization have been done. Solubility of protein depends on pressure. For instance, the solubility of tetragonal lysozyme crystal increased with increasing pressure, while that of orthorhombic crystal decreased. The solubility of subtilisin increased with increasing pressure. Crystal growth rates of protein also depend on pressure. The growth rate of glucose isomerase was significantly enhanced with increasing pressure. The growth rate of tetragonal lysozyme crystal and subtilisin decreased with increasing pressure. To study the effects of pressure on the crystallization more precisely and systematically, hen egg white lysozyme is the most suitable protein at this stage, since a lot of data can be used. We focused on growth kinetics under high pressure, since extensive studies on growth kinetics have already been done at atmospheric pressure, and almost all of them have explained the growth mechanisms well. The growth rates of tetragonal lysozyme decreased with pressure under the same supersaturation. This means that the surface growth kinetics significantly depends on pressure. By analyzing the dependence of supersaturation on growth rate, it was found that the increase in average ledge surface energy of the two-dimensional nuclei with pressure explained the decrease in growth rate. At this stage, it is not clear whether the increase in surface energy with increasing pressure is the main reason or not. Fundamental studies on protein crystallization under high pressure will be useful for high pressure crystallography and high pressure protein science.

ja: [Invited review] 圧力は系全体に均一にかつ非常に速く伝播するため，タンパク質の結晶化を制御するための重要なパラメタである．これまで多くの研究がなされてきた．まず結晶の溶解度は圧力に依存する．例えば正方晶系リゾチーム結晶の溶解度は加圧に伴い増加し，斜方晶系リゾチーム結晶の溶解度は逆に減少した．サブチリシン結晶の溶解度は加圧とともに増加した．また，結晶成長速度も圧力に依存する．グルコースイソメラーゼ結晶の成長速度は，加圧に伴い非常に促進されるが，正方晶系リゾチーム結晶とサブチリシン結晶の成長速度は逆に減少する．我々は高圧力下における成長kineticsに注目した．なぜなら，既に常圧下で多くの研究がなされており，そのほとんど全てが結晶成長機構をうまく説明できているからである．正方晶系リゾチーム結晶の成長速度の過飽和度依存性を解析した結果，二次元核のステップレッジ表面自由エネルギーが加圧に伴い増加するという結果が得られた．高圧力下におけるタンパク質結晶化の基礎的な研究は，高圧力下における結晶構造解析や高圧力下におけるタンパク質科学に有用であろう．

en: Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology


en: Pressure Effects of Crystallization of Hen Egg White Lysozyme

ja: リゾチームの結晶化に及ぼす圧力の効果

en: タンパク質の結晶化に及ぼす圧力効果についてニワトリ卵白リゾチームをモデルタンパクとして研究した．ダイヤモンドアンビルセルを用いた直接観察により，加圧に伴う正方晶系結晶の溶解度の増加と，核生成頻度·結晶成長速度の減少が明らかになった．定量的な研究を行うために，高圧力下で二光束干渉計を用いた新規な溶解度測定法を開発した．その結果，正方晶系結晶では加圧に伴い溶解度が増加したのに対し，斜方晶系結晶では減少した．この傾向については，結晶中の分子結合サイトに対する水和を考えることで議論した．結晶の成長速度を同じ結晶化駆動力で比較しても高圧力下の方が結晶成長速度が遅くなり，圧力は速度論的にも結晶化に影響を及ぼすことが明らかになった．これは正方晶系，斜方晶系とも同様であった．二次元核成長モデルで成長速度の解析を行った結果，正方晶系では，加圧に伴う結晶の平均ステップレッジ表面自由エネルギーの増加が成長速度の減少の原因であることが分かった．

en: Journal of the Japanese Association for Crystal Growth


en: 化学応用工学科のFD活動

ja: 化学応用工学科のFD活動

en: 平成24年度FD研究報告書

ja: 平成24年度FD研究報告書

misc_type: others


en: 化学応用工学科のFD活動について

ja: 化学応用工学科のFD活動について

en: 平成23年度FD研究報告書

ja: 平成23年度FD研究報告書


en: Development of techniques for creating high-quality colloidal c

ja: 高品質コロイド結晶成長技術の確立

en: Bulletin of Faculty of Engineering, The University of Tokushima

ja: 徳島大学工学部研究報告