1997 Fiscal Year Annual Research Report

タンパク質の構造安定化機構への温度・圧力効果の研究

Research Project

Project/Area Number	08045025
Research Institution	Ritsumeikan University
Principal Investigator	澤村精治立命館大学, 理工学部, 助教授 (10167439)
Co-Investigator(Kenkyū-buntansha)	ペィティグレンフェルブリティッシュコロンビア大学, 理学部, 教授マクギリブレイロスブリティッシュコロンビア大学, 理学部, 教授マウクグラントブリティッシュコロンビア大学, 医学部, 教授加藤稔立命館大学, 理工学部, 講師 (00241258) 谷口吉弘立命館大学, 理工学部, 教授 (70066702)
Keywords	タンパク質 / 高圧力 / リゾチーム
Research Abstract	最初にタンパク質の存在する水中での溶質溶媒相互作用についても調べた。そのいくつかを示す。タンパク質の疎水部分のモデルとして四級アンモニウム塩を用い、この水溶液のNMR緩和の回転アニソトロピーから3種の水和水の存在を確認した。リゾチーム-アルコール-水系の熱測定からは、タンパク質表面の結合サイトのアルコールによる占有はそのアルコールにとって必ずしもタンパク質の性質に影響を及ぼさなくてもよいことを明らかにした。リゾチームの水への高圧溶解度を測定した。一方でタンパク質の構成部分であるアミノ酸の中でもっとも単純なグリシンについても高圧溶解度を測定した。その結果、リゾチームの溶解度は加圧とともに増加するのに対し、グリシンは減少した。両者とも加熱すると溶解度は増加することから、溶解度の圧力依存性を調べることの方が、タンパク質と水との相互作用についての情報が得られやすいことが示唆された。タンパク質については、クジラミオグロビン(Mb)のFT-IRスペクトルを数千気圧の高圧下で-10〜70℃で測定した。その結果、圧力によるMbの二次構造は二状態転移を示し、加熱によりMbの二次構造は分子間ベータシートの形成を伴いながらランダムコイルへと不可逆的に転移すること、さらに低温ではMbの二次構造はランダムコイルへと可逆的に変化する事を明らかにした。また、遺伝子工学的手法によるタンパク質試料の調製を試みた。その結果、大腸菌を利用する事により酵母チトクロムCを調整することが可能となり、ワイルドタイプのチトクロムCが比較的高収率で得られることとなった。このことによりさらに異なったタイプのチトクロムCも調製できる可能性を示し、耐熱・耐圧を有するタンパク質を調製する道が開けた。

Research Products

(33 results)

All Other

All Publications (33 results)

[Publications] T.Nakai: "Effects of Pressure and Temperatur on the viscosity 13 coefficiant for Potassium Chloride in Water" Matter.Sci.Res.Int.,. 3. 204-209 (1997)
[Publications] N.Takeda: "Pressure-induced Structural rearrangements of bovine pancreatic trypsin inhibitor studied by FTIR Spectroscopy" Biospectroscopy. 印刷中. (1998)
[Publications] T.Matsumoto: "Effect of Pressure on the mechanism of hydrolysis of malcternose,maltopcrvtaose,and maltohexose catalyzed by porcine paticieatic α-amylase" Biochim.Biophys.Acta.1343. 243-250 (1997)
[Publications] K.Fumino: "NMR Studies on dynamic structure of hydrated water molecules in tetraalkylammonium bromide-D_2O Solutions at5-25℃" J.Mol.Liquids,. 72、(印刷中).
[Publications] K.Fumino: "Concontration dependence of nuclear magnetic relaxation vntes of alkalimetal ionic nucliein dilute solutions" Denki Kagaku. 65. 198-203 (1997)
[Publications] Y.Koga: ""Intermolecular Interactions"in Aqueous Solutions of Three Components including Lysozyme" Fluid Phase Egnil.(印刷中). (1998)
[Publications] I.J.F.Day: "Ion Solration in Binavy Mixtares" J.Chem.Phys.106. 2782-2793 (1997)
[Publications] G.Ayton: "Liquid Crystal Phases of Dipolar Discotic Particles" Phys.Rer.E. 55. 447-454 (1997)
[Publications] M.Twitchett: "Intramolecular Electron Transfer Kinetics of a Synthetin Flavocytochrome C" J.Am.Chem.Soc.119. 435-436 (1997)
[Publications] J.Bujons: "Charge Reversal of a Critical Active Site Residue of Cytochrome C Peroxidase:Cfaracterixation of the Afs 48Glu Variant" Eur.J.Biochem. 243. 72-84 (1997)
[Publications] C.L.Hunter: "Role of Heme Propionates in the Interaction of Heme with Apomyoglobin and Apocytochrome b5." Biochemistry. 36. 1010-1017 (1997)
[Publications] C.L.Hunter: "Dissociation of Heme from Myoglobin and Cytochrome b5:Comparison of Behavior in Solution and the Gas Phase" Biochemistry. 36. 1018-1025 (1997)
[Publications] J.M.Nocek: "Cyclic Electron Transfer with in the [Zn-Deuteremyoglobin,Cytochromb5]Complex:Redox-Linked Changes at theInterface" J.Am.Chem.Lec.119. 2146-2155 (1997)
[Publications] A.G.Mank: "Control of Metalloprotein Redox Potentials:What Does Site-Directed Mutagenesis of Hemoproteins Tell Us ?" J.Biol.Inors.Chem.2. 119-125 (1997)
[Publications] K.Kishi: "Site-Directed Mutations of Phonylalanire-190 of Manganese Peroxidase:Effects on Stability,Function and Coordination" Biochemistry. 36. 4268-4277 (1997)
[Publications] L.Konermann: "Acid-Induced Donaturation of Myoglobin Studied by Time-Resolved Electrospray Ionization Mass Spectrometry" Biochemistry. 36. 6448-6454 (1997)
[Publications] R.Maurus: "A Myoglobin Variant witha Polar Substitution in Conserved Hydrophobic Cluster in the Heme Binding Pocket" Biochim.Biophys.Acta.1341. 1-13 (1997)
[Publications] Cote.H.C.F: "A Novel approach for the characterization and epitope determination of alupus anticoagnlant-associatedneutralizing anti-prothrombin antibody" Am.J.Clin.Phthol.107. 197-205 (1997)
[Publications] P.Westh: "Intermolecular Interactions of Lysozyme and Small Alcohols:A Calorimetric Investigation" J.Phys.Chem. B101. 5755-5758 (1997)
[Publications] Y.Xu: "Light Scattering Study of Sol-Gel Trsnsformation of Sio_<> Colloid at Extremely High Concentrations" Clays and Clay Minerals. (印刷中). (1998)
[Publications] D.R.Berard: "Structive of the Metal-Aqueous Electrolyte Solution Interface" J.Chem.Phys.107. 4719-4728 (1997)
[Publications] J.C.Shelley: "Modeling and Structure of Mercury-Water Interface" J.Chem.Phys.107. 2122-2141 (1997)
[Publications] G.Ayton: "Ferroelectic and Dipolar Glass Phases of Non-Crystalline Systems" Phys.Rev.E. 56. 562-570 (1997)
[Publications] N.M.Cann: "An Investigation of the Infulence of Solute Size and Insertion on Solvation Thermodynamics" J.Chem.Phys.106. 8165-8195 (1997)
[Publications] Cote.H.C.F.: "Functional Characterization of recombinant human meizo-thrombin and meizothrombin(DesF1)" J.Biol.Chem.272. 6194-6200 (1997)
[Publications] M.A.Brown: "Vectors for expressing proteinsat the amino-terminus of an activationdomain for use in the yeast two-hybridsystem" Anal.Biochem.247. 451-452 (1997)
[Publications] Q.Y.He: "Effects of mutations of aspartic acid63 on the metal-binding properties of the vecanbinant N-lobe of humanserum transferrin" Biochemistry. 36. 5522-5528 (1997)
[Publications] L.A.Luck: "19F NMR Studies of the recombinant human transferrin N-lobe and three singlepoint mutants" Mag.Reson.Chem.35. 477-481 (1997)
[Publications] A.B.Mason: "Receptor recognition Sites reside in both lobes of Humann Serum transferrin" Biochem.J.326. 77-85 (1997)
[Publications] O.Zak: "A kinetically active site in the C-lobe of human transferrin" Biochemistry. 36. 11036-11043 (1997)
[Publications] M.M.Guarna: "On-line monitoring and control of methanol concentration in Shake-flask cultures of Pichia pastoris" Biotechnol.Bioerg.56. 279-286 (1997)
[Publications] Q-Y.He: "Inequivalence of the two tyrosine ligandsin the N-lobe of human serumtransferrin" Biochemistry. 36. 14853-14860 (1997)
[Publications] 谷口吉弘: "生物・環境産学のための非加熱プロセス事典" サイエンスフォーラム, 95-99 (1997)

1997 Fiscal Year Annual Research Report

タンパク質の構造安定化機構への温度・圧力効果の研究

Principal Investigator

澤村 精治 立命館大学, 理工学部, 助教授 (10167439)

Research Products

[Publications] T.Nakai: "Effects of Pressure and Temperatur on the viscosity 13 coefficiant for Potassium Chloride in Water" Matter.Sci.Res.Int.,. 3. 204-209 (1997)

[Publications] N.Takeda: "Pressure-induced Structural rearrangements of bovine pancreatic trypsin inhibitor studied by FTIR Spectroscopy" Biospectroscopy. 印刷中. (1998)

[Publications] T.Matsumoto: "Effect of Pressure on the mechanism of hydrolysis of malcternose,maltopcrvtaose,and maltohexose catalyzed by porcine paticieatic α-amylase" Biochim.Biophys.Acta.1343. 243-250 (1997)

[Publications] K.Fumino: "NMR Studies on dynamic structure of hydrated water molecules in tetraalkylammonium bromide-D_2O Solutions at5-25℃" J.Mol.Liquids,. 72、(印刷中).

[Publications] K.Fumino: "Concontration dependence of nuclear magnetic relaxation vntes of alkalimetal ionic nucliein dilute solutions" Denki Kagaku. 65. 198-203 (1997)

[Publications] Y.Koga: ""Intermolecular Interactions"in Aqueous Solutions of Three Components including Lysozyme" Fluid Phase Egnil.(印刷中). (1998)

[Publications] I.J.F.Day: "Ion Solration in Binavy Mixtares" J.Chem.Phys.106. 2782-2793 (1997)

[Publications] G.Ayton: "Liquid Crystal Phases of Dipolar Discotic Particles" Phys.Rer.E. 55. 447-454 (1997)

[Publications] M.Twitchett: "Intramolecular Electron Transfer Kinetics of a Synthetin Flavocytochrome C" J.Am.Chem.Soc.119. 435-436 (1997)

[Publications] J.Bujons: "Charge Reversal of a Critical Active Site Residue of Cytochrome C Peroxidase:Cfaracterixation of the Afs 48Glu Variant" Eur.J.Biochem. 243. 72-84 (1997)

[Publications] C.L.Hunter: "Role of Heme Propionates in the Interaction of Heme with Apomyoglobin and Apocytochrome b5." Biochemistry. 36. 1010-1017 (1997)

[Publications] C.L.Hunter: "Dissociation of Heme from Myoglobin and Cytochrome b5:Comparison of Behavior in Solution and the Gas Phase" Biochemistry. 36. 1018-1025 (1997)

[Publications] J.M.Nocek: "Cyclic Electron Transfer with in the [Zn-Deuteremyoglobin,Cytochromb5]Complex:Redox-Linked Changes at theInterface" J.Am.Chem.Lec.119. 2146-2155 (1997)

[Publications] A.G.Mank: "Control of Metalloprotein Redox Potentials:What Does Site-Directed Mutagenesis of Hemoproteins Tell Us ?" J.Biol.Inors.Chem.2. 119-125 (1997)

[Publications] K.Kishi: "Site-Directed Mutations of Phonylalanire-190 of Manganese Peroxidase:Effects on Stability,Function and Coordination" Biochemistry. 36. 4268-4277 (1997)

[Publications] L.Konermann: "Acid-Induced Donaturation of Myoglobin Studied by Time-Resolved Electrospray Ionization Mass Spectrometry" Biochemistry. 36. 6448-6454 (1997)

[Publications] R.Maurus: "A Myoglobin Variant witha Polar Substitution in Conserved Hydrophobic Cluster in the Heme Binding Pocket" Biochim.Biophys.Acta.1341. 1-13 (1997)

[Publications] Cote.H.C.F: "A Novel approach for the characterization and epitope determination of alupus anticoagnlant-associatedneutralizing anti-prothrombin antibody" Am.J.Clin.Phthol.107. 197-205 (1997)

[Publications] P.Westh: "Intermolecular Interactions of Lysozyme and Small Alcohols:A Calorimetric Investigation" J.Phys.Chem. B101. 5755-5758 (1997)

[Publications] Y.Xu: "Light Scattering Study of Sol-Gel Trsnsformation of Sio_<> Colloid at Extremely High Concentrations" Clays and Clay Minerals. (印刷中). (1998)

[Publications] D.R.Berard: "Structive of the Metal-Aqueous Electrolyte Solution Interface" J.Chem.Phys.107. 4719-4728 (1997)

[Publications] J.C.Shelley: "Modeling and Structure of Mercury-Water Interface" J.Chem.Phys.107. 2122-2141 (1997)

[Publications] G.Ayton: "Ferroelectic and Dipolar Glass Phases of Non-Crystalline Systems" Phys.Rev.E. 56. 562-570 (1997)

[Publications] N.M.Cann: "An Investigation of the Infulence of Solute Size and Insertion on Solvation Thermodynamics" J.Chem.Phys.106. 8165-8195 (1997)

[Publications] Cote.H.C.F.: "Functional Characterization of recombinant human meizo-thrombin and meizothrombin(DesF1)" J.Biol.Chem.272. 6194-6200 (1997)

[Publications] M.A.Brown: "Vectors for expressing proteinsat the amino-terminus of an activationdomain for use in the yeast two-hybridsystem" Anal.Biochem.247. 451-452 (1997)

[Publications] Q.Y.He: "Effects of mutations of aspartic acid63 on the metal-binding properties of the vecanbinant N-lobe of humanserum transferrin" Biochemistry. 36. 5522-5528 (1997)

[Publications] L.A.Luck: "19F NMR Studies of the recombinant human transferrin N-lobe and three singlepoint mutants" Mag.Reson.Chem.35. 477-481 (1997)

[Publications] A.B.Mason: "Receptor recognition Sites reside in both lobes of Humann Serum transferrin" Biochem.J.326. 77-85 (1997)

[Publications] O.Zak: "A kinetically active site in the C-lobe of human transferrin" Biochemistry. 36. 11036-11043 (1997)

[Publications] M.M.Guarna: "On-line monitoring and control of methanol concentration in Shake-flask cultures of Pichia pastoris" Biotechnol.Bioerg.56. 279-286 (1997)

[Publications] Q-Y.He: "Inequivalence of the two tyrosine ligandsin the N-lobe of human serumtransferrin" Biochemistry. 36. 14853-14860 (1997)

[Publications] 谷口 吉弘: "生物・環境産学のための非加熱プロセス事典" サイエンスフォーラム, 95-99 (1997)

澤村精治立命館大学, 理工学部, 助教授 (10167439)

[Publications] 谷口吉弘: "生物・環境産学のための非加熱プロセス事典" サイエンスフォーラム, 95-99 (1997)