渗透剂稳定蛋白质结构的分子动力学模拟研究
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摘要
只有正确折叠的蛋白质分子才能具有生物活性;因此在蛋白质工程和临床医学中,必须保证目标蛋白质处于合适的构象。渗透剂对蛋白质结构有着独特的稳定作用,因此被广泛应用于人工稳定和复性蛋白质技术中,其稳定机理也成为了生物技术领域中研究的热点。之前有研究发现渗透剂对蛋白质的稳定能力与它的分子表面的极性基团表面积比例呈反比关系,但忽略了分子体积所代表的空间位阻效应的影响,因此存在一定误差。我们推测分子体积也是影响渗透剂稳定作用的因素,对之前的理论进行了修正和发展,对于优化实验和生产条件、理性设计新型添加剂和配基,具有重要的指导意义。
本研究利用了计算机模拟的优势,以CI2为模型蛋白,通过分子动力学模拟考察8种典型渗透剂对CI2的热稳定性的影响;研究核心是根据模拟数据,利用统计学双参数拟合模型同时分析分子体积和极性基团表面积比例与渗透剂稳定能力的关系,加深了渗透剂稳定能力的影响因素的理性认识。之后将模型结果与具体的模拟数据分析手段结合,进一步的分析了多元醇类渗透剂与蛋白质作用的分子机理,阐述了造成多元醇类渗透剂独特性质的原因。
研究首先发现渗透剂对CI2有着不同程度的稳定能力;之后定义并计算了能够综合描述CI2结构特性的一维参数,从量化的角度定义了相应渗透剂对CI2的稳定能力。通过与实验结果的关联,认为模拟结果是比较可信的。
经过数学模型分析,发现分子体积的引入大大提高了模型精度,证明渗透剂的体积大小对其稳定能力有着重要的影响;由于分子体积与空间位阻效应直接相关,因此当极性基团表面积比例一定时,渗透剂的稳定能力随着分子体积的增大而增强;当分子体积一定时,渗透剂的稳定能力首先随着极性基团表面积比例的增加而下降,当这个比例超过某个阀值时,其稳定能力反而开始上升。
分析过程中发现多元醇类渗透剂与其他渗透剂的差异较大,因此单独对几种多元醇类渗透剂进行分析,发现其稳定能力与分子体积直接相关;而由于它们的极性基团表面积比例普遍较高,并且彼此差异不大,因此这个比例的影响很小,并且与相应的稳定能力存在正相关的现象。在分子机理层面,发现醇类渗透剂较大的分子体积造成的空间位阻效应,以及表面多个羟基导致的分子间彼此聚集现象,是造成它们独特性质的主要因素。
Only protein folded correctly could have bioactivity. Therefore, to protect the target protein in its native conformation is an important issue both in protein engineering and clinical medicine. Because osmolytes have particular stabilizing functions on protein structures in nature, they have been widely utilized into artificial protein stabilizing and refolding technologies, with their mechanisms also studied greatly in the biotechnology field. There was a research before which found that the osmolyte stabilizing effect have negative correlations with the ratio of polar group surface area of their molecules. However, it failed to consider the molecular volume which could represent the steric exclusion effect, therefore had some deviations. We speculate the molecular volume is also an influential factor of osmolyte effects. The research successfully revises and improves former mechanisms, shedding some lights on experiment and manufacture condition optimization, and new additive and ligand designs.
Our research employs CI2 as the model protein, using molecular dynamics simulation to study eight typical osmolytes’effect on CI2’s thermal stability. Basing on the simulation data, the core is to use statistical bivariate fit model to simultaneously analyze the correlations of both osmolyte molecular volume and ratio of polar group surface area between their stabilizing abilities. After that, by combining the model results with the simulation analysis methods, the molecular mechanism of polyol-protein interaction has been studied, then the reason inducing the particularity of polyols is subsequently illustrated.
The research first finds osmolytes have different stabilizing abilities on CI2; then we define and calculate the one-dimensional coefficient of CI2 structure, by which the stabilizing abilities of osmolytes are defined quantitatively. By its correlation with experimental results, the simulation results are believed reliable.
By model analysis, it is found that including volume effect increase the model precision greatly, which proves the osmolyte volume functions greatly. Since the molecular volume correlates with steric exclusion effect directly, osmolyte stabilizing ability increases as the volume increase; when volume is fixed, osmolyte stabilizing ability first decrease as the ratio of polar group surface area increase, when this ratio is larger a certain cut-off, osmolyte stabilizing ability reversely begin to increase.
During the analyzing process, the polyols are found with great discrepancies from other osmolyte types. Therefore, we separately analyze their data, and find their stabilizing effects almost only correlate with their molecular volumes. Because their ratios of polar group surface area are generally high and similar with each other, this ratio does not have significant impact any more, and owns a positive correlation with relative stabilizing abilities. On the level of molecular mechanism, it is found the main factors of their particularity are their steric exclusion effect induced by their huge volumes, as well as their self-association phenomena induced by multiple hydroxyl groups on their surfaces.
本研究利用了计算机模拟的优势,以CI2为模型蛋白,通过分子动力学模拟考察8种典型渗透剂对CI2的热稳定性的影响;研究核心是根据模拟数据,利用统计学双参数拟合模型同时分析分子体积和极性基团表面积比例与渗透剂稳定能力的关系,加深了渗透剂稳定能力的影响因素的理性认识。之后将模型结果与具体的模拟数据分析手段结合,进一步的分析了多元醇类渗透剂与蛋白质作用的分子机理,阐述了造成多元醇类渗透剂独特性质的原因。
研究首先发现渗透剂对CI2有着不同程度的稳定能力;之后定义并计算了能够综合描述CI2结构特性的一维参数,从量化的角度定义了相应渗透剂对CI2的稳定能力。通过与实验结果的关联,认为模拟结果是比较可信的。
经过数学模型分析,发现分子体积的引入大大提高了模型精度,证明渗透剂的体积大小对其稳定能力有着重要的影响;由于分子体积与空间位阻效应直接相关,因此当极性基团表面积比例一定时,渗透剂的稳定能力随着分子体积的增大而增强;当分子体积一定时,渗透剂的稳定能力首先随着极性基团表面积比例的增加而下降,当这个比例超过某个阀值时,其稳定能力反而开始上升。
分析过程中发现多元醇类渗透剂与其他渗透剂的差异较大,因此单独对几种多元醇类渗透剂进行分析,发现其稳定能力与分子体积直接相关;而由于它们的极性基团表面积比例普遍较高,并且彼此差异不大,因此这个比例的影响很小,并且与相应的稳定能力存在正相关的现象。在分子机理层面,发现醇类渗透剂较大的分子体积造成的空间位阻效应,以及表面多个羟基导致的分子间彼此聚集现象,是造成它们独特性质的主要因素。
Only protein folded correctly could have bioactivity. Therefore, to protect the target protein in its native conformation is an important issue both in protein engineering and clinical medicine. Because osmolytes have particular stabilizing functions on protein structures in nature, they have been widely utilized into artificial protein stabilizing and refolding technologies, with their mechanisms also studied greatly in the biotechnology field. There was a research before which found that the osmolyte stabilizing effect have negative correlations with the ratio of polar group surface area of their molecules. However, it failed to consider the molecular volume which could represent the steric exclusion effect, therefore had some deviations. We speculate the molecular volume is also an influential factor of osmolyte effects. The research successfully revises and improves former mechanisms, shedding some lights on experiment and manufacture condition optimization, and new additive and ligand designs.
Our research employs CI2 as the model protein, using molecular dynamics simulation to study eight typical osmolytes’effect on CI2’s thermal stability. Basing on the simulation data, the core is to use statistical bivariate fit model to simultaneously analyze the correlations of both osmolyte molecular volume and ratio of polar group surface area between their stabilizing abilities. After that, by combining the model results with the simulation analysis methods, the molecular mechanism of polyol-protein interaction has been studied, then the reason inducing the particularity of polyols is subsequently illustrated.
The research first finds osmolytes have different stabilizing abilities on CI2; then we define and calculate the one-dimensional coefficient of CI2 structure, by which the stabilizing abilities of osmolytes are defined quantitatively. By its correlation with experimental results, the simulation results are believed reliable.
By model analysis, it is found that including volume effect increase the model precision greatly, which proves the osmolyte volume functions greatly. Since the molecular volume correlates with steric exclusion effect directly, osmolyte stabilizing ability increases as the volume increase; when volume is fixed, osmolyte stabilizing ability first decrease as the ratio of polar group surface area increase, when this ratio is larger a certain cut-off, osmolyte stabilizing ability reversely begin to increase.
During the analyzing process, the polyols are found with great discrepancies from other osmolyte types. Therefore, we separately analyze their data, and find their stabilizing effects almost only correlate with their molecular volumes. Because their ratios of polar group surface area are generally high and similar with each other, this ratio does not have significant impact any more, and owns a positive correlation with relative stabilizing abilities. On the level of molecular mechanism, it is found the main factors of their particularity are their steric exclusion effect induced by their huge volumes, as well as their self-association phenomena induced by multiple hydroxyl groups on their surfaces.
引文
[1]阎隆飞,孙之荣,蛋白质分子结构,北京,清华大学出版社,1999,21l-221
[2] Levintal, C., How to fold graciously In Mossbauer Spectroscopy in Biological Systems. In: Monticello I L eds. Proceedings of a Meeting held at Allerton House, University of Illinois Press, 1969, 22-24
[3] Haber, E. and Anfinsen, C.B., Side-chain interactions governing the pairing of half-cystine residues in ribonuclease. J Biol Chem, 1962, 237: 1839-1844
[4] Anfinsen, C.B., Principles that govern the folding of protein chains. Science, 1973, 181 (96): 223-230
[5] Kim, P.S. and Baldwin, R.L., Intermediates in the folding reactions of small proteins. Annu Rev Biochem, 1990, 59: 631-660
[6] Fersht, A.R., Optimization of rates of protein folding: the nucleation-condensation mechanism and its implications. Proc Natl Acad Sci U S A, 1995, 92 (24): 10869-10873
[7] Schellman, J.A., The stability of hydrogen-bonded peptide structures in aqueous solution. C R Trav Lab Carlsberg [Chim], 1955, 29 (14-15): 230-259
[8] Vendruscolo, M. and Dobson, C.M., Towards complete descriptions of the free-energy landscapes of proteins. Philos Transact A Math Phys Eng Sci, 2005, 363 (1827): 433-50; discussion 450-452
[9] Yancey, P.H., Water Stress, Osmolytes, and Proteins. AMER. ZOOL, 2001, 41: 699-709
[10] Crowe, J.H., Hoekstra, F.A., and Crowe, L.M., Anhydrobiosis. Annu Rev Physiol, 1992, 54: 579-599
[11] Story, K.B., Organic Solutes in freezing tolerance. Comp. Biochem. Physiol, 1997, 117 (3): 319-326
[12] Siebenaller, J.F. and Somero, G.N., Biochemical adaptation to the deep sea. CRC Crit. Rev. Aq. Sci., 1989, 1: 1-25
[13] Hardy, J. and Selkoe, D.J., The amyloid hypothesis of Alzheimer's disease: progress and problems on the road to therapeutics. Science, 2002, 297 (5580): 353-356
[14] Dobson, C.M., Protein misfolding, evolution and disease. Trends Biochem Sci, 1999, 24 (9): 329-332.
[15] Goldberg, M.S. and Lansbury, P.T., Jr., Is there a cause-and-effect relationship between alpha-synuclein fibrillization and Parkinson's disease? Nat Cell Biol, 2000, 2 (7): E115-119
[16] Yancey, P.H. and Somero, G.N., Counteraction of urea destabilization of protein structure by methylamine osmoregulatory compounds of elasmobranch fishes. Biochem J, 1979, 183 (2): 317-323
[17] Yancey, P.H., et al., Living with water stress: evolution of osmolyte systems. Science, 1982, 217 (4566): 1214-1222
[18] Timasheff, S.N., A physicochemical basis for the selection of osmolytes by nature. In G. N. Somero, C. B. Osmond, and C. L. Bolis (eds.). Water and life: A comparative analysis of water relationships at the organismic, cellular, and molecular levels, Springer-Verlag, Berlin, 1992, 70-84
[19] Santoro, M.M., et al., Increased thermal stability of proteins in the presence of naturally occurring osmolytes. Biochemistry, 1992, 31 (23): 5278-5283
[20] Timasheff, S.N., The control of protein stability and association by weak interactions with water: how do solvents affect these processes? Annu Rev Biophys Biomol Struct, 1993, 22: 67-97
[21] De Virgilio, C., et al., Acquisition of thermotolerance in Saccharomyces cerevisiae without heat shock protein hsp 104 and in the absence of protein synthesis. FEBS Lett, 1991, 288 (1-2): 86-90
[22] Salvucci, M.E., Hendrix, D.L., and Wolfe, G.R., A thermoprotective role for sorbitol in the silverleaf whitefly, Bemisia argentifolii. J Insect Physiol, 1998, 44 (7-8): 597-603
[23] Yancey, P.H. and Walsh, L.P., Hyperthermic and hypertonic shock induce HSP70 accumulation and thermotolerance but not osmotic tolerance in cultured mammalian renal cells. The Physiologist, 1994, 37: A98
[24] Gillett, M.B., et al., Elevated levels of trimethylamine oxide in muscles of deep-sea gadiform teleosts: A high-pressure adaptation? J. Exper. Zool., 1997, 279: 386-391
[25] Yancey, P.H., Compatible and counteracting solutes. In:Strange K eds. Cellular and molecular physilology of cell volume regulation. CRC Press, Boca Raton, 1994, 81-109
[26] Dong, X.Y., Huang, Y., and Sun, Y., Refolding kinetics of denatured-reduced lysozyme in the presence of folding aids. J Biotechnol, 2004, 114 (1-2): 135-142
[27] Caldas, T., et al., Thermoprotection by glycine betaine and choline. Microbiology, 1999, 145 ( Pt 9) 2543-2548
[28] Singer, M.A. and Lindquist, S., Multiple effects of trehalose on protein folding in vitro and in vivo. Mol Cell, 1998, 1 (5): 639-648
[29] Liu, R., et al., Trehalose differentially inhibits aggregation and neurotoxicity of beta-amyloid 40 and 42. Neurobiol Dis, 2005, 20 (1): 74-81
[30] Brown, A.D. and Simpson, J.R., Water relations of sugar-tolerant yeasts: the role of intracellular polyols. J Gen Microbiol, 1972, 72 (3): 589-591
[31] Pollard, A. and Wyn Jones, R.G., Enzyme activities in concentrated solutions of glycinebetaine and other solutes. Planta, 1979, 144 291-298
[32] Low, P.S., Molecular basis of the biological compatibility of nature's osmolytes. In R. Gilles and M. Gilles-Ballien (eds.). Transport processes, iono- and osmoregulation, Springer-Verlag, Berlin, 1985, 469-477
[33] Roccatano, D., Computer simulations study of biomolecules in non-aqueous or cosolvent/water mixture solutions. Curr Protein Pept Sci, 2008, 9 (4): 407-426
[34] Collins, K.D. and Washabaugh, M.W., The Hofmeister effect and the behaviour of water at interfaces. Q Rev Biophys, 1985, 18 (4): 323-422
[35] Washabaugh, M.W. and Collins, K.D., The systematic characterization by aqueous column chromatography of solutes which affect protein stability. J Biol Chem, 1986, 261 (27): 12477-12485
[36] Clark, M.E., The osmotic role of amino acids: Discovery and function. In R. Gilles and M. Gilles-Ballien (eds.). Transport processes, iono- and osmoregulation, Springer-Verlag, Berlin, 1985, 412-423
[37] Wu, J.W. and Wang, Z.X., New evidence for the denaturant binding model. Protein Sci, 1999, 8 (10): 2090-2097
[38] Cottone, G., Cordone, L., and Ciccotti, G., Molecular dynamics simulation of carboxy-myoglobin embedded in a trehalose-water matrix. Biophys J, 2001, 80 (2): 931-938
[39] Cottone, G., et al., Molecular dynamics simulation of sucrose- and trehalose-coated carboxy-myoglobin. Proteins, 2005, 59 (2): 291-302
[40] Arakawa, T. and Timasheff, S.N., The stabilization of proteins by osmolytes. Biophys J, 1985, 47 (3): 411-414
[41] Timasheff, S.N., Protein hydration, thermodynamic binding, and preferential hydration. Biochemistry, 2002, 41 (46): 13473-13482
[42] Timasheff, S.N., Protein-solvent preferential interactions, protein hydration, and the modulation of biochemical reactions by solvent components. Proc Natl Acad Sci U S A, 2002, 99 (15): 9721-9726
[43] Bolen, D.W. and Baskakov, I.V., The osmophobic effect: natural selection of a thermodynamic force in protein folding. J Mol Biol, 2001, 310 (5): 955-963
[44] Liu, Y. and Bolen, D.W., The peptide backbone plays a dominant role in protein stabilization by naturally occurring osmolytes. Biochemistry, 1995, 34 (39): 12884-91
[45] Street, T.O., Bolen, D.W., and Rose, G.D., A molecular mechanism for osmolyte-induced protein stability. Proc Natl Acad Sci U S A, 2006, 103 (38): 13997-14002
[46] Plaza del Pino, I.M. and Sanchez-Ruiz, J.M., An osmolyte effect on the heat capacity change for protein folding. Biochemistry, 1995, 34 (27): 8621-8630
[47] Berg, O.G., The influence of macromolecular crowding on thermodynamic activity: solubility and dimerization constants for spherical and dumbbell-shaped molecules in a hard-sphere mixture. Biopolymers, 1990, 30 (11-12): 1027-1037
[48] Parsegian, V.A., Rand, R.P., and Rau, D.C., Osmotic stress, crowding, preferential hydration, and binding: A comparison of perspectives. Proc Natl Acad Sci U S A, 2000, 97 (8): 3987-3992
[49] Davis-Searles, P.R., et al., Interpreting the effects of small uncharged solutes on protein-folding equilibria. Annu Rev Biophys Biomol Struct, 2001, 30: 271-306
[50] RCSB Protein Data Bank, An Information Portal to Biological Macromolecular Structures [EB/OL]. http://www.rcsb.org/pdb/home/home.do, 2006, 02: 1411
[51] Duan, Y. and Kollman, P.A., Pathways to a protein folding intermediate observed in a 1-microsecond simulation in aqueous solution. Science, 1998, 282 (5389): 740-744
[52] Cheatham, T.E., 3rd, Simulation and modeling of nucleic acid structure, dynamics and interactions. Curr Opin Struct Biol, 2004, 14 (3): 360-367
[53] Callen, H.B., Thermodynamics and an Introduction to Thermostatistics. Wiley:New York, 1985
[54] Alder, B.J. and Wainwright, T.E., Journal of Chemistry Physics, 1957, 27: 1208-1209
[55] McCammon, J.A., Molecular Dynamics study of the bovine pancreatic trypsin inhibitor. In Models for Protein Dynamics. CECAM: Orsay, France, 1976, 137
[56] McPhalen, C.A., et al., Crystal and molecular structure of chymotrypsin inhibitor 2 from barley seeds in complex with subtilisin Novo. Proc Natl Acad Sci U S A, 1985, 82 (21): 7242-7246
[57] Otzen, D.E., et al., Structure of the transition state for the folding/unfolding of the barley chymotrypsin inhibitor 2 and its implications for mechanisms of protein folding. Proc Natl Acad Sci U S A, 1994, 91 (22): 10422-10425
[58] Li, L. and Shakhnovich, E.I., Constructing, verifying, and dissecting the folding transition state of chymotrypsin inhibitor 2 with all-atom simulations. Proc Natl Acad Sci U S A, 2001,98 (23): 13014-13018
[59] Day, R., et al., Increasing temperature accelerates protein unfolding without changing the pathway of unfolding. J Mol Biol, 2002, 322 (1): 189-203
[60] Day, R. and Daggett, V., Sensitivity of the folding/unfolding transition state ensemble of chymotrypsin inhibitor 2 to changes in temperature and solvent. Protein Sci, 2005, 14 (5): 1242-1252
[61] Bennion, B.J. and Daggett, V., The molecular basis for the chemical denaturation of proteins by urea. Proc Natl Acad Sci U S A, 2003, 100 (9): 5142-5147
[62] Bennion, B.J. and Daggett, V., Counteraction of urea-induced protein denaturation by trimethylamine N-oxide: a chemical chaperone at atomic resolution. Proc Natl Acad Sci U S A, 2004, 101 (17): 6433-6438
[63] Van Der Spoel, D., et al., GROMACS: fast, flexible, and free. J Comput Chem, 2005, 26 (16): 1701-1718
[64] Van Gunsteren, W.F., et al., Biomolecular simulation: The GROMOS96 manual and user guide. 1996
[65] Berendsen, H.J.C., et al., Intermolecular Forces. 1981
[66] Verlet, L., Computer experiments on classical fluids. I. Thermodynamical properties of Lennard-Jones molecules. Phys. Rev., 1967, 159: 98-103
[67] van Gunsteren, W.F. and Berendsen, H.J.C., Algorithms for macromolecular dynamics and constraint dynamics. Mol. Phys., 1977, 34: 1311-1327
[68] Humphrey, W., Dalke, A., and Schulten, K., VMD: visual molecular dynamics. J Mol Graph, 1996, 14 (1): 33-38, 27-28
[69] Kabsch, W. and Sander, C., Dictionary of protein secondary structure: pattern recognition of hydrogen-bonded and geometrical features. Biopolymers, 1983, 22 (12): 2577-2637
[70] Cohen, F.E. and Sternberg, M.J., On the prediction of protein structure: The significance of the root-mean-square deviation. J Mol Biol, 1980, 138 (2): 321-333
[71] Beck, D.A. and Daggett, V., A one-dimensional reaction coordinate for identification of transition states from explicit solvent P(fold)-like calculations. Biophys J, 2007, 93 (10): 3382-3391
[72] Calandrini, V., Onori, G., and Santucci, A., Examination by dynamic light scattering of lysozyme in water/alcohol mixtures Journal of Molecular Structure, 2001, 565: 183-188
[73] Westh, P. and Koga, Y., Intermolecular Interactions of Lysozyme and Small Alcohols: A Calorimetric Investigation. J Phys Chem B, 1997, 101: 5755-5758
[74] Lerbret, A., et al., How do trehalose, maltose, and sucrose influence some structural and dynamical properties of lysozyme? Insight from molecular dynamics simulations. J Phys Chem B, 2007, 111 (31): 9410-9420
[2] Levintal, C., How to fold graciously In Mossbauer Spectroscopy in Biological Systems. In: Monticello I L eds. Proceedings of a Meeting held at Allerton House, University of Illinois Press, 1969, 22-24
[3] Haber, E. and Anfinsen, C.B., Side-chain interactions governing the pairing of half-cystine residues in ribonuclease. J Biol Chem, 1962, 237: 1839-1844
[4] Anfinsen, C.B., Principles that govern the folding of protein chains. Science, 1973, 181 (96): 223-230
[5] Kim, P.S. and Baldwin, R.L., Intermediates in the folding reactions of small proteins. Annu Rev Biochem, 1990, 59: 631-660
[6] Fersht, A.R., Optimization of rates of protein folding: the nucleation-condensation mechanism and its implications. Proc Natl Acad Sci U S A, 1995, 92 (24): 10869-10873
[7] Schellman, J.A., The stability of hydrogen-bonded peptide structures in aqueous solution. C R Trav Lab Carlsberg [Chim], 1955, 29 (14-15): 230-259
[8] Vendruscolo, M. and Dobson, C.M., Towards complete descriptions of the free-energy landscapes of proteins. Philos Transact A Math Phys Eng Sci, 2005, 363 (1827): 433-50; discussion 450-452
[9] Yancey, P.H., Water Stress, Osmolytes, and Proteins. AMER. ZOOL, 2001, 41: 699-709
[10] Crowe, J.H., Hoekstra, F.A., and Crowe, L.M., Anhydrobiosis. Annu Rev Physiol, 1992, 54: 579-599
[11] Story, K.B., Organic Solutes in freezing tolerance. Comp. Biochem. Physiol, 1997, 117 (3): 319-326
[12] Siebenaller, J.F. and Somero, G.N., Biochemical adaptation to the deep sea. CRC Crit. Rev. Aq. Sci., 1989, 1: 1-25
[13] Hardy, J. and Selkoe, D.J., The amyloid hypothesis of Alzheimer's disease: progress and problems on the road to therapeutics. Science, 2002, 297 (5580): 353-356
[14] Dobson, C.M., Protein misfolding, evolution and disease. Trends Biochem Sci, 1999, 24 (9): 329-332.
[15] Goldberg, M.S. and Lansbury, P.T., Jr., Is there a cause-and-effect relationship between alpha-synuclein fibrillization and Parkinson's disease? Nat Cell Biol, 2000, 2 (7): E115-119
[16] Yancey, P.H. and Somero, G.N., Counteraction of urea destabilization of protein structure by methylamine osmoregulatory compounds of elasmobranch fishes. Biochem J, 1979, 183 (2): 317-323
[17] Yancey, P.H., et al., Living with water stress: evolution of osmolyte systems. Science, 1982, 217 (4566): 1214-1222
[18] Timasheff, S.N., A physicochemical basis for the selection of osmolytes by nature. In G. N. Somero, C. B. Osmond, and C. L. Bolis (eds.). Water and life: A comparative analysis of water relationships at the organismic, cellular, and molecular levels, Springer-Verlag, Berlin, 1992, 70-84
[19] Santoro, M.M., et al., Increased thermal stability of proteins in the presence of naturally occurring osmolytes. Biochemistry, 1992, 31 (23): 5278-5283
[20] Timasheff, S.N., The control of protein stability and association by weak interactions with water: how do solvents affect these processes? Annu Rev Biophys Biomol Struct, 1993, 22: 67-97
[21] De Virgilio, C., et al., Acquisition of thermotolerance in Saccharomyces cerevisiae without heat shock protein hsp 104 and in the absence of protein synthesis. FEBS Lett, 1991, 288 (1-2): 86-90
[22] Salvucci, M.E., Hendrix, D.L., and Wolfe, G.R., A thermoprotective role for sorbitol in the silverleaf whitefly, Bemisia argentifolii. J Insect Physiol, 1998, 44 (7-8): 597-603
[23] Yancey, P.H. and Walsh, L.P., Hyperthermic and hypertonic shock induce HSP70 accumulation and thermotolerance but not osmotic tolerance in cultured mammalian renal cells. The Physiologist, 1994, 37: A98
[24] Gillett, M.B., et al., Elevated levels of trimethylamine oxide in muscles of deep-sea gadiform teleosts: A high-pressure adaptation? J. Exper. Zool., 1997, 279: 386-391
[25] Yancey, P.H., Compatible and counteracting solutes. In:Strange K eds. Cellular and molecular physilology of cell volume regulation. CRC Press, Boca Raton, 1994, 81-109
[26] Dong, X.Y., Huang, Y., and Sun, Y., Refolding kinetics of denatured-reduced lysozyme in the presence of folding aids. J Biotechnol, 2004, 114 (1-2): 135-142
[27] Caldas, T., et al., Thermoprotection by glycine betaine and choline. Microbiology, 1999, 145 ( Pt 9) 2543-2548
[28] Singer, M.A. and Lindquist, S., Multiple effects of trehalose on protein folding in vitro and in vivo. Mol Cell, 1998, 1 (5): 639-648
[29] Liu, R., et al., Trehalose differentially inhibits aggregation and neurotoxicity of beta-amyloid 40 and 42. Neurobiol Dis, 2005, 20 (1): 74-81
[30] Brown, A.D. and Simpson, J.R., Water relations of sugar-tolerant yeasts: the role of intracellular polyols. J Gen Microbiol, 1972, 72 (3): 589-591
[31] Pollard, A. and Wyn Jones, R.G., Enzyme activities in concentrated solutions of glycinebetaine and other solutes. Planta, 1979, 144 291-298
[32] Low, P.S., Molecular basis of the biological compatibility of nature's osmolytes. In R. Gilles and M. Gilles-Ballien (eds.). Transport processes, iono- and osmoregulation, Springer-Verlag, Berlin, 1985, 469-477
[33] Roccatano, D., Computer simulations study of biomolecules in non-aqueous or cosolvent/water mixture solutions. Curr Protein Pept Sci, 2008, 9 (4): 407-426
[34] Collins, K.D. and Washabaugh, M.W., The Hofmeister effect and the behaviour of water at interfaces. Q Rev Biophys, 1985, 18 (4): 323-422
[35] Washabaugh, M.W. and Collins, K.D., The systematic characterization by aqueous column chromatography of solutes which affect protein stability. J Biol Chem, 1986, 261 (27): 12477-12485
[36] Clark, M.E., The osmotic role of amino acids: Discovery and function. In R. Gilles and M. Gilles-Ballien (eds.). Transport processes, iono- and osmoregulation, Springer-Verlag, Berlin, 1985, 412-423
[37] Wu, J.W. and Wang, Z.X., New evidence for the denaturant binding model. Protein Sci, 1999, 8 (10): 2090-2097
[38] Cottone, G., Cordone, L., and Ciccotti, G., Molecular dynamics simulation of carboxy-myoglobin embedded in a trehalose-water matrix. Biophys J, 2001, 80 (2): 931-938
[39] Cottone, G., et al., Molecular dynamics simulation of sucrose- and trehalose-coated carboxy-myoglobin. Proteins, 2005, 59 (2): 291-302
[40] Arakawa, T. and Timasheff, S.N., The stabilization of proteins by osmolytes. Biophys J, 1985, 47 (3): 411-414
[41] Timasheff, S.N., Protein hydration, thermodynamic binding, and preferential hydration. Biochemistry, 2002, 41 (46): 13473-13482
[42] Timasheff, S.N., Protein-solvent preferential interactions, protein hydration, and the modulation of biochemical reactions by solvent components. Proc Natl Acad Sci U S A, 2002, 99 (15): 9721-9726
[43] Bolen, D.W. and Baskakov, I.V., The osmophobic effect: natural selection of a thermodynamic force in protein folding. J Mol Biol, 2001, 310 (5): 955-963
[44] Liu, Y. and Bolen, D.W., The peptide backbone plays a dominant role in protein stabilization by naturally occurring osmolytes. Biochemistry, 1995, 34 (39): 12884-91
[45] Street, T.O., Bolen, D.W., and Rose, G.D., A molecular mechanism for osmolyte-induced protein stability. Proc Natl Acad Sci U S A, 2006, 103 (38): 13997-14002
[46] Plaza del Pino, I.M. and Sanchez-Ruiz, J.M., An osmolyte effect on the heat capacity change for protein folding. Biochemistry, 1995, 34 (27): 8621-8630
[47] Berg, O.G., The influence of macromolecular crowding on thermodynamic activity: solubility and dimerization constants for spherical and dumbbell-shaped molecules in a hard-sphere mixture. Biopolymers, 1990, 30 (11-12): 1027-1037
[48] Parsegian, V.A., Rand, R.P., and Rau, D.C., Osmotic stress, crowding, preferential hydration, and binding: A comparison of perspectives. Proc Natl Acad Sci U S A, 2000, 97 (8): 3987-3992
[49] Davis-Searles, P.R., et al., Interpreting the effects of small uncharged solutes on protein-folding equilibria. Annu Rev Biophys Biomol Struct, 2001, 30: 271-306
[50] RCSB Protein Data Bank, An Information Portal to Biological Macromolecular Structures [EB/OL]. http://www.rcsb.org/pdb/home/home.do, 2006, 02: 1411
[51] Duan, Y. and Kollman, P.A., Pathways to a protein folding intermediate observed in a 1-microsecond simulation in aqueous solution. Science, 1998, 282 (5389): 740-744
[52] Cheatham, T.E., 3rd, Simulation and modeling of nucleic acid structure, dynamics and interactions. Curr Opin Struct Biol, 2004, 14 (3): 360-367
[53] Callen, H.B., Thermodynamics and an Introduction to Thermostatistics. Wiley:New York, 1985
[54] Alder, B.J. and Wainwright, T.E., Journal of Chemistry Physics, 1957, 27: 1208-1209
[55] McCammon, J.A., Molecular Dynamics study of the bovine pancreatic trypsin inhibitor. In Models for Protein Dynamics. CECAM: Orsay, France, 1976, 137
[56] McPhalen, C.A., et al., Crystal and molecular structure of chymotrypsin inhibitor 2 from barley seeds in complex with subtilisin Novo. Proc Natl Acad Sci U S A, 1985, 82 (21): 7242-7246
[57] Otzen, D.E., et al., Structure of the transition state for the folding/unfolding of the barley chymotrypsin inhibitor 2 and its implications for mechanisms of protein folding. Proc Natl Acad Sci U S A, 1994, 91 (22): 10422-10425
[58] Li, L. and Shakhnovich, E.I., Constructing, verifying, and dissecting the folding transition state of chymotrypsin inhibitor 2 with all-atom simulations. Proc Natl Acad Sci U S A, 2001,98 (23): 13014-13018
[59] Day, R., et al., Increasing temperature accelerates protein unfolding without changing the pathway of unfolding. J Mol Biol, 2002, 322 (1): 189-203
[60] Day, R. and Daggett, V., Sensitivity of the folding/unfolding transition state ensemble of chymotrypsin inhibitor 2 to changes in temperature and solvent. Protein Sci, 2005, 14 (5): 1242-1252
[61] Bennion, B.J. and Daggett, V., The molecular basis for the chemical denaturation of proteins by urea. Proc Natl Acad Sci U S A, 2003, 100 (9): 5142-5147
[62] Bennion, B.J. and Daggett, V., Counteraction of urea-induced protein denaturation by trimethylamine N-oxide: a chemical chaperone at atomic resolution. Proc Natl Acad Sci U S A, 2004, 101 (17): 6433-6438
[63] Van Der Spoel, D., et al., GROMACS: fast, flexible, and free. J Comput Chem, 2005, 26 (16): 1701-1718
[64] Van Gunsteren, W.F., et al., Biomolecular simulation: The GROMOS96 manual and user guide. 1996
[65] Berendsen, H.J.C., et al., Intermolecular Forces. 1981
[66] Verlet, L., Computer experiments on classical fluids. I. Thermodynamical properties of Lennard-Jones molecules. Phys. Rev., 1967, 159: 98-103
[67] van Gunsteren, W.F. and Berendsen, H.J.C., Algorithms for macromolecular dynamics and constraint dynamics. Mol. Phys., 1977, 34: 1311-1327
[68] Humphrey, W., Dalke, A., and Schulten, K., VMD: visual molecular dynamics. J Mol Graph, 1996, 14 (1): 33-38, 27-28
[69] Kabsch, W. and Sander, C., Dictionary of protein secondary structure: pattern recognition of hydrogen-bonded and geometrical features. Biopolymers, 1983, 22 (12): 2577-2637
[70] Cohen, F.E. and Sternberg, M.J., On the prediction of protein structure: The significance of the root-mean-square deviation. J Mol Biol, 1980, 138 (2): 321-333
[71] Beck, D.A. and Daggett, V., A one-dimensional reaction coordinate for identification of transition states from explicit solvent P(fold)-like calculations. Biophys J, 2007, 93 (10): 3382-3391
[72] Calandrini, V., Onori, G., and Santucci, A., Examination by dynamic light scattering of lysozyme in water/alcohol mixtures Journal of Molecular Structure, 2001, 565: 183-188
[73] Westh, P. and Koga, Y., Intermolecular Interactions of Lysozyme and Small Alcohols: A Calorimetric Investigation. J Phys Chem B, 1997, 101: 5755-5758
[74] Lerbret, A., et al., How do trehalose, maltose, and sucrose influence some structural and dynamical properties of lysozyme? Insight from molecular dynamics simulations. J Phys Chem B, 2007, 111 (31): 9410-9420