Hostname: page-component-745bb68f8f-lrblm Total loading time: 0 Render date: 2025-02-05T03:50:40.488Z Has data issue: false hasContentIssue false
  • English
  • Français

Mutational studies of protein structures and their stabilities

Published online by Cambridge University Press:  17 March 2009

David Shortle
David Shortle
Affiliation:
The Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, Maryland 21205, USA
Get access Rights & Permissions [Opens in a new window]

Extract

The fundamental relationship between structure and function has served to guide investigations into the workings of living systems at all levels - from the whole organism to individual cells on down to individual molecules. When X-ray crystallography began to reveal the three-dimensional structures of proteins like myoglobin, lysozyme and RNase A, protein chemists were well prepared to draw inferences about functional mechanisms from the precise positioning of amino acid residues they could see. The close proximity between an amino acid side chain and a chemical group on a bound ligand strongly suggests a functional role for that side chain in binding affinity and specificity. Likewise, the nearly universal finding of large clusters of hydrophobic side chains buried in the core of proteins strongly supports a major functional role of hydrophobic interactions in protein folding and stability. Even though eminently plausible hypotheses like these, grounded in the most fundamental principles of chemistry and the logic of structure–function relationships, become widely accepted and make their way into textbooks, protein chemists have felt compelled to search for ways to test them and put them on a more quantitative basis.

Type
Research Article
Information
Quarterly Reviews of Biophysics , Volume 25 , Issue 2 , May 1992 , pp. 205 - 250
Copyright
Copyright © Cambridge University Press 1992

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Ackers, G. K. & Smith, F. R. (1985). Effects of site-specific amino acid modification on protein interactions and bioogical function. A. Rev. Biochem. 54, 597–629.CrossRefGoogle Scholar
Ackers, G. K. & Smith, F. R. (1987). The hemoglobin tetramer. A three-state molecular snitch for control of ligand affinity. A. Rev. Biophys. Biophys. Chem. 16, 583–609.CrossRefGoogle Scholar
Ackers, G. K., Doyle, M. L., Myers, D. & Daugherty, M. A. (1992). Molecular code for cooperativity in hemoglobin. Science 255, 54–63.CrossRefGoogle ScholarPubMed
Ahrweiler, P. M. & Frieden, C. (1991). Effects of point mutations in a hinge region on the stability, folding, and enzymatic activity of Escherichia coli dihydrofolate reductase. Biochemistry 30, 7801–7809.CrossRefGoogle Scholar
Akke, M. & Forsen, S. (1990). Protein stability and electrostatic interactions between solvent exposed charged side chains. Proteins: Struct. Fund. Genet. 8, 23–29.CrossRefGoogle ScholarPubMed
Alber, T. (1989). Mutational effects on protein stability. A. Rev. Biochem. 58, 765–798.CrossRefGoogle ScholarPubMed
Alber, T., Dao-Pin, S., Nye, J. A., Muchmore, D. C. & Matthews, B. W. (1987). Temperature-sensitive mutations of bacteriophage T4 lysozyme occur at sites with low mobility and low solvent accessibility in the folded protein. Biochemistry 26, 3754–3758.CrossRefGoogle ScholarPubMed
Alber, T., Bell, J. A., Dao-Pin, S., Nicholson, H., Wozniak, J. A., Cook, S. & Matthews, B. W. (1988). Replacement of pro86 in phage T4 lysozyme extend an αhelix but do not alter protein stability. Science 239, 631–635.CrossRefGoogle Scholar
Alexandrescu, A. T., Ulrich, E. L. & Markley, J. L. (1989). Hydrogen-i nmr evidence for three interconverting forms of staphylococcal nuclease: effects of mutations and solution conditions on their distribution. Biochemistry 28, 204–211.CrossRefGoogle Scholar
Anderson, D. E., Becktel, W. J. & Dahlquist, F. W. (1990). pH-induced denaturation of proteins: a single salt bridge contributes 3–5 kcal/mole to the free energy of folding of T4 lysozyme. Biochemistry 29, 2403–2308.CrossRefGoogle Scholar
Artymiuk, P. J., Blake, C. C. F., Grace, D. E. P., Oatley, S. J., Phillips, D. C. & Sternberg, M. J. E. (1979). Crystallographic studies of the dynamic properties of lysozyme. Nature (Lond.) 280, 563–568.CrossRefGoogle ScholarPubMed
Auld, D. S. & Pielak, G. J. (1991). Constraints on amino acid substitutions in the Nterminal helix of cytochrome c explored by random mutagenesis. Biochemistry 30, 8684–8690.CrossRefGoogle ScholarPubMed
Baker, E. N. & Hubbard, R. E. (1984). Hydrogen bonding in globular proteins. Prog. Biophys. Mol. Biol. 44, 97–141.CrossRefGoogle ScholarPubMed
Baum, J., Dobson, C. M., Evans, P. A. & Hanley, C. (1989). Characterization of a partly folded protein by NMR methods: studies on the molten globule state of guinea pig alpha-lactalbumin. Biochemistry 28, 7–13.CrossRefGoogle ScholarPubMed
Bax, A. (1991). Experimental NMR techniques for studies of biopolymers. Curr. Opin. struc. Biol. I, 1030–1035.CrossRefGoogle Scholar
Becktel, W. J. & Schellman, J. A. (1987). Protein stability curves. Biopolymers 26, 1859–1877.CrossRefGoogle ScholarPubMed
Behe, M. J., Lattman, E. E. & Rose, G. D. (1991). The protein-folding problem: the native fold determines packing, but does packing determine the native fold? Proc. natn. Acad. Sci. USA 88, 4195–4199.CrossRefGoogle ScholarPubMed
Birdsall, B., Feeney, J., Tendler, S. J. B., Hammond, S. J. & Roberts, G. C. K. (1989). Dihydrofolate reductase: multiple conformations and alternative modes of substrate binding. Biochemistry 28, 2297–2305.CrossRefGoogle ScholarPubMed
Bolden, D. W. & Santoro, M. M. (1988). Unfolding free energy changes determined by the linear extrapolation method. 2. Incorporation of values in a thermodynamic cycle. Biochemistry 27, 8069–8074.CrossRefGoogle Scholar
Braiman, M. S. & Rothschild, K. J. (1988). Fourier transform infrared techniques for probing membrane protein structure. A. Rev. Biophys. Biophys. Chem. 17, 541–570.CrossRefGoogle ScholarPubMed
Brünger, A. T. (1991). Crystallographic phasing and refinement of macromolecules. Curr. Opin. Struc. Biol. 1, 1016–1022.CrossRefGoogle Scholar
Carter, P. J., Winter, G., Wilkinson, A. J. & Fersht, A. R. (1984). The use of double mutants to detect structural changes in the active site of the tyrosyl-tRNA synthetase (Bacillus stearothermophilus). Cell 38, 835–840.CrossRefGoogle ScholarPubMed
Connelly, P., Ghosaini, L., Hu, C.-Q., – Kitamura, S., Tanaka, A. & Sturtevant, J. M. (1991). A differential scanning calorimetric study of the thermal unfolding of seven mutant forms of phage T4 lysozyme. Biochemistry 30, 1887–1891.CrossRefGoogle ScholarPubMed
Cornell, W. D., Howard, A. E. & Kollman, P. (1991). Molecular mechanical potential functions and their application to study molecular systems. Curr. Opin. struc. Biol. 1, 201–212.CrossRefGoogle Scholar
Cunningham, B. C. & Wells, J. A. (1989). High-resolution epitope mapping of hGH—receptor interactions by alanine-scanning mutagenesis. Science 244, 1081–1085.CrossRefGoogle ScholarPubMed
Dang, L. X., Merz, K. M. Jr, & Kollman, P. A. (1989). Free energy calculations on protein stability: Thr-157 → Val-157 mutation of T4 lysozyme. J. Am. Chem. Soc. III, 8505–8508.CrossRefGoogle Scholar
Dao-Pin, S., Alber, T., Baase, W. A., Wozniak, J. A. & Matthews, B. W. (1991 a). Structural and thermodynamic analysis of the packing of two α-helices in bacteriophage T4 lysozyme. J. Mol. Biol. 221, 647–667.CrossRefGoogle Scholar
Dao-Pin, S., Anderson, D. E., Baase, W. A., Dahlquist, F. W. & Matthews, B. W. (1991 b). Structural and thermodynamic consequences of burying a charged residue within the hydrophobic core of T4 lysozyme. Biochemistry 30, 11521–11529.CrossRefGoogle ScholarPubMed
Dao-Pin, S., Soderlind, E., Baase, W. A., Wozniak, J. A., Sauer, U. & Matthews, B. W. (1991 c). Cumulative site-directed charge-change replacements in T4 lysozyme suggest that long-range electrostatic interactions contribute little to protein stability. J. Mol. Biol. 221, 873–887.CrossRefGoogle ScholarPubMed
Daugherty, M. A., Shea, M. A., Johnson, J. A., Licata, V. J., Turner, G. J. & Ackers, G. K. (1991). Identification of the intermediate allosteric species in human hemoglobin reveals a molecular code for cooperative switching. Proc. natn. Acad. Sci. USA 88, 1110–1114.CrossRefGoogle ScholarPubMed
Dill, K. A. (1990). Dominant Forces in protein folding. Biochemistry 29, 7133–7155.CrossRefGoogle ScholarPubMed
Ecker, D. J., Butt, T. R., Marsh, J., Sternberg, E., Shatzman, A., Dixon, J. S., Weber, P. L. & Crooke, S. T. (1989). Ubiquitin function studied by disulfide engineering. J. Biol. Chem. 264, 1887–1893.CrossRefGoogle ScholarPubMed
Eigenbrot, C., Randal, M. & Kossiakoff, A. A. (1990). Structural effects induced by removal of a disulfide-bridge: the X-ray structure of the C30A/C51A mutant of basic pancreatic trypsin inhibitor at i-6 Å. Protein Engineering 3, 591–598.CrossRefGoogle Scholar
Eisenstein, E., Markby, D. W. & Schachman, H. K. (1990). Heterotropic effectors promote a global conformational change in aspartate transcarbamoylase. Biochemistry 29, 3724–3731.CrossRefGoogle ScholarPubMed
Ellman, J. A., Mendel, D. & Schultz, P. G. (1992). Site-specific incorporation of novel backbone structures into proteins. Science 255, 197–200.CrossRefGoogle ScholarPubMed
Eriksson, A. E., Baase, W. A., Zhang, X.-J., Heinz, D. W., Blaber, M., Baldwin, E. P. & Matthews, B. W. (1992). Response of a protein structure to cavity-creating mutations and its relation to the hydrophobic effect. Science 255, 178–183.CrossRefGoogle ScholarPubMed
Evans, P. A., Dobson, C. M., Kautz, R. A., Hatfull, G. & Fox, R. O. (1987). Proline isomerism in staphylococcal nuclease characterized by nmr and site-directed mutagenesis. Nature 329, 266–268.CrossRefGoogle ScholarPubMed
Evans, P. R. (1991). Structural aspects of allostery. Curr. Opin. Struc. Biol. 1, 773–779.CrossRefGoogle Scholar
Faber, H. R. & Matthews, B. W. (1990). A mutant T4 lysozyme displays five different crystal conformations. Nature 348, 263–266.CrossRefGoogle ScholarPubMed
Fersht, A. (1985). Enzyme Structure and Mechanism. San Francisco: W. H. Freeman and Company.Google Scholar
Fersht, A. R. (1988). Relationships between apparent binding energies measured in site-directed mutagenesis experiments and energetics of binding and catalysis. Biochemistry 27, 1577–1580.CrossRefGoogle ScholarPubMed
Fishel, L. A., Villafranca, J. E., Mauro, J. M. & Kraut, J. (1987). Yeast cytochrome c peroxidase: mutagenesis and expression in Escherichia coli show tryptophan-51 is not the radical site in compound I. Biochemistry 26, 351–360.CrossRefGoogle Scholar
Fox, R. O., Evans, P. A. & Dobson, C. M. (1986). Multiple conformations of a protein demonstrated by magnetization transfer nmr spectroscopy. Nature 320, 192–194.CrossRefGoogle ScholarPubMed
Gardell, S. J., Craik, C. S., Hilvert, D., Urdea, M. S. & Rutter, W. J. (1985). Sitedirected mutagenesis shows that tyrosine 248 of carboxypeptidase A does not play a crucial role in catalysis. Nature 317, 551–554.CrossRefGoogle Scholar
Garvey, E. P. & Matthews, C. R. (1989). Effects of multiple replacements at a single position on the folding and stability of dihydrofolate reductase from Escherichia coli. Biochemistry 28, 2083–2093.CrossRefGoogle Scholar
Gibbs, C. S. & Zoller, M. J. (1991). Identification of electrostatic interactions that determine the phosphorylation site specificity of the cAMP-dependent protein kinase. Biochemistry 22, 5329–5334.CrossRefGoogle Scholar
Gibbs, M. R., Moody, P. C. E. & Leslie, A. G. W. (1990). Crystal structure of the aspartic acid-199 → asparagine mutant of chloramphenicol acetyltransferase to 2°35-Å resolution: structural consequences of disruption of a buried salt bridge. Biochemistry 29, 11261–11265.CrossRefGoogle Scholar
Goldenberg, D. P. (1988). Genetic studies of protein stability and mechanisms of folding. A. Rev. Biophys. Biophys. Chem. 17, 481–507.CrossRefGoogle ScholarPubMed
Gray, T. M. & Matthews, B. W. (1987). Structural analysis of the temperaturesensitive mutant of bacteriophage T4 lysozyme, glycine 156 → aspartic acid. J. Biol. Chem. 262, 16858–16864.CrossRefGoogle ScholarPubMed
Green, S. M., Meeker, A. K. & Shortle, D. (1992 a). Contributions of the polar, uncharged amino acids to the stability of staphylococcal nuclease: evidence for mutational effects on the free energy of the denatured state. Biochemistry, in press.CrossRefGoogle Scholar
Green, S. M., Sondek, J. & Shortle, D. (1992b). Manuscript in preparation.Google Scholar
Grütter, M. G., Gray, T. M., Weaver, L. H., Alber, T., Wilson, K. & Matthews, B. W. (1987). Structural studies of mutants of the lysozyme of bacteriophage T4. The temperature-sensitive mutant protein Thr157 → Ile. J. Mol. Biol. 197, 315–329.CrossRefGoogle ScholarPubMed
Harary, F. (1969). Graph Theory. Reading, MA: Addison-Wesley.CrossRefGoogle Scholar
Hawkes, R., Grutter, M. G. & Schellman, J. (1984). Thermodynamic stability and point mutations of bacteriophage T4 lysozyme. Jf. Mol. Biol. 175, 195–212.CrossRefGoogle ScholarPubMed
Hecht, M. H., Sturtevant, J. M. & Sauer, R. T. (1984). Effect of single amino acid replacements on the thermal stability of the NH2-terminal domain of phage lambda repressor. Proc. Natn. Acad. Sci. USA 81, 5685–5689.CrossRefGoogle ScholarPubMed
Hecht, M. H., Sturtevant, J. M. & Sauer, R. T. (1986). Stabilization of λ repressor against thermal denaturation by site-directed gly → ala changes in α-helix 3. Proteins: Struct. Fund. Genet. 1, 43–46.CrossRefGoogle ScholarPubMed
Hibler, D. W., Stolowich, N. J., Reynolds, M. A., Gerlt, J. A., Wilde, J. A. & Bolton, P. H. (1987). Site-directed mutants of staphylococcal nuclease. Detection and localization by 1H-NMR spectroscopy of conformational changes accompanying substitution for glutamic acid 43. Biochemistry 26, 6278–6283.CrossRefGoogle ScholarPubMed
Horovitz, A., Serrano, L., Avron, B., Bycroft, M. & Fersht, A. R. (1990). Strength and co-operativity of contributions of surface salt bridges to protein stability. J. Mol. Biol. 216, 1031–1044.CrossRefGoogle ScholarPubMed
Howell, E. E., Villafranca, J. E., Warren, M. S., Oatley, S. J. & Kraut, J. (1986). Functional role of aspartic acid-27 in dihydrofolate reductase revealed by mutagenesis. Science 231, 1123–1128.CrossRefGoogle ScholarPubMed
Hughson, F. M. & Baldwin, R. L. (1989). Use of site-directed mutagenesis to destabilize native apomyoglobin relative to folding intermediates. Biochemistry 28, 4415–4422.CrossRefGoogle ScholarPubMed
Hughson, F. M., Barrick, D. & Baldwin, R. L. (1991). Probing the stability of a partly folded apomyoglobin intermediate by site-directed mutagenesis. Biochemistry 30, 4113–4118.CrossRefGoogle ScholarPubMed
Hurle, M. R., Tweedy, N. B. & Matthews, C. R. (1986). Synergism in folding a double mutant of the alpha subunit of tryptophan synthetase. Biochemistry 25, 6356–6360.CrossRefGoogle Scholar
Hurley, J. H., Baase, W. A. & Matthews, B. W. (1992). Design and structural analysis of alternative hydrophobic core packing arrangements in T4 lysozyme. J. Mol. Biol., in press.CrossRefGoogle Scholar
Jacobson, R., Matsumura, M.Faber, H. R. & Matthews, B. W. (1992). Structure of a stabilizing disulfide bridge mutant that closes the active-site cleft of T4 lysozyme. Protein Science 1, 46–57.CrossRefGoogle ScholarPubMed
Johnson, K. A. & Benkovic, S. J. (1990). Analysis of protein function by mutagenesis. The Enzymes 19, 159–211.CrossRefGoogle Scholar
Kaiser, E. M., Lawrence, D. S. & Rokita, S. E. (1985). The chemical modification of enzymatic specificity. A. Rev. Biochem. 54, 565–595.CrossRefGoogle ScholarPubMed
Kantrowitz, E. R. & Lipscomb, W. N. (1990). Escherichia coli aspartate transcarbamoylase: the molecular basis for a concerted allosteric transition. TIBS 15, 53–59.Google ScholarPubMed
Katz, B. & Kossiakoff, A. A. (1986). The crystallographically determined structures of atypical strained disulfides engineered into subtilising. Biol. Chem. 261, 15480–15485.CrossRefGoogle Scholar
Katz, B. & Kossiakoff, A. A. (1990). Crystal structures of subtilisin BPN′ variants containing disulfide bonds and cavities: concerted structural rearrangements induced by mutagenesis. Proteins: Struct. Fund. Genet. 7, 343–357.CrossRefGoogle ScholarPubMed
Kauzmann, W. (1959). Some factors in the interpretation of protein denaturation. Adv. Protein Chem. 14, 1–63.CrossRefGoogle ScholarPubMed
Keefe, L. J., Lattman, E. E., Wolkow, C., Woods, A., Chevrier, M. & Cotter, R. J. (1992 a). Resolution of a protein sequence ambiguity by X-ray crystallographic and mass spectrometric methods. J. Appl. Cryst. 25, (in press).CrossRefGoogle Scholar
Keefe, L. J., Sondek, J., Gittis, A., Shortle, D. & Lattman, E. E. (1992 b). Manuscripts in preparation.Google Scholar
Kellis, J. T. Jr, Nyberg, K. & Fersht, A. R. (1989). Energetics Of Complementary side-chain packing in a protein hydrophobic core. Biochemistry 28, 4914–4922.CrossRefGoogle Scholar
Kellis, J. T. Jr, Nyberg, K., Sali, D. & Fersht, A. R. (1988). Contribution of hydrophobic interactions to protein stability. Nature 333, 784–786.CrossRefGoogle ScholarPubMed
Kitamura, S. & Sturtevant, J. M. (1989). A scanning calorimetric study of the thermal denaturation of the lysozyme of phage T4 and the Arg 96 → His mutant form thereof. Biochemistry 28, 3788–3792.CrossRefGoogle ScholarPubMed
Knowles, J. R. (1987). Tinkering with enzymes: what are we learning? Science 236, 1252–1258.CrossRefGoogle ScholarPubMed
Koshland, D. E., Nemethy, G. & Filmer, D. (1966). Comparison of experimental binding data and theoretical models of proteins containing subunits. Biochemistry 5, 365–385.CrossRefGoogle ScholarPubMed
Kossiakoff, A. A., Randal, M., Guenot, J. & Eigenbrot, C. (1992). Variability of conformations at crystal contacts in BPTI represent true low energy structures: correspondence among lattice packing and molecular dynamics structures. Proteins 13 (in press).CrossRefGoogle Scholar
Kundrot, C. E. & Evans, P. R. (1991). Designing an allosterically locked phosphofructokinase. Biochemistry 30, 1478–1484.CrossRefGoogle ScholarPubMed
Kuntz, I. D. (1975). An approach to the tertiary structure of globular proteins. J. Am. Chem. Soc. 97, 4362–4366.CrossRefGoogle Scholar
Kuriyan, J. & Weis, W. I. (1991). Rigid protein motion as a model for crystallographic temperature factors. Proc. Natl. Acad. Sci. USA 88, 2773–2777.CrossRefGoogle Scholar
Lau, F. T.-K. & Fersht, A. R. (1987). Conversion of allosteric inhibition to activation in phosphofructokinase by protein engineering. Nature 326, 811–812.Google ScholarPubMed
Leatherbarrow, R. J. & Fersht, A. R. (1987). Investigation of transition-state stabilization by residues histidine-45 and threonine-40 in the tyrosyl-tRNA synthetase. Biochemistry 26, 8524–8528.CrossRefGoogle ScholarPubMed
Lee, C. & Levitt, M. (1991). Accurate prediction of the stability and activity effects of site-directed mutagenesis on a protein core. Nature 352, 448–450.CrossRefGoogle ScholarPubMed
Lee, J. C., Gekko, K. & Timasheff, S. N. (1979). Measurements of preferential solvent interactions by densimetric techniques. In Methods in Enzymology, vol. 61 (ed. Hirs, C. H. W. and Timasheff, S. N.), pp. 26–57. San Diego: Academic Press.Google Scholar
Lim, W. A. & Sauer, R. T. (1989). Alternative packing arrangements in the hydrophobic core of lambda repressor. Nature 339, 31–36.CrossRefGoogle ScholarPubMed
Lim, W. A. & Sauer, R. T. (1991). The role of internal packing interactions in determining the structure and stability of a protein. J. Mol. Biol. 219, 359–376.CrossRefGoogle ScholarPubMed
Lim, W. A., Farruggio, D. C. & Sauer, R. T. (1992). The structural and energetic consequences of disruptive mutations in a protein core. Biochemistry, in press.CrossRefGoogle Scholar
Lin, T.-Y. & Kim, P. S. (1989). Urea dependence of thiol-disulfide equilibria in thioredoxin: confirmation of the linkage relationship and a sensitive assay for structure. Biochemistry 28, 5282–5287.CrossRefGoogle Scholar
Loll, P. J. & Lattman, E. E. (1990). Active site mutant glu-43 → asp in staphylococcal nuclease displays nonlocal structural changes. Biochemistry 29, 6866–6873.CrossRefGoogle ScholarPubMed
Lumry, R. & Rajender, S. (1970). Enthalpy-entropy compensation phenomena in water solutions of proteins and small molecules: a ubiquitous property of water. Biopolymers 9, 1125–1227.CrossRefGoogle ScholarPubMed
McCamon, J. A. & Harvey, S. C. (1987). Dynamics of Proteins and Nucleic Acids. New York: Cambridge University Press.CrossRefGoogle Scholar
McGrath, M. E., Wilke, M. E., Higaki, J. N., Craik, C. S. & Fletterick, R. J. (1989). Crystal structures of two engineered thiol trypsins. Biochemistry 28, 9264–9270.CrossRefGoogle ScholarPubMed
McRee, D. E., Redford, S. M., Getzoff, E. E., Lepock, J. R., Hallewell, R. A. & Tainer, J. A. (1990). Changes in crystallographic structure and thermostability of a Cu, Zn superoxide dismutase mutant resulting from the removal of a buried cysteine. J. Biol. Chem. 265, 14234–14241.CrossRefGoogle ScholarPubMed
Markley, J. L., Williams, M. N. & Jardetzky, O. (1970). Nuclear magnetic resonance studies of the structure and binding sites of enzymes. XII. A conformational equilibrium in staphylococcal nuclease involving a histidine residue. Proc. Natl. Acad. Sci. USA 65, 645–651.CrossRefGoogle ScholarPubMed
Matsumura, M., Becktel, W. J. & Matthews, B. W. (1988). Hydrophobic stabilization in T4 lysozyme determined directly by multiple substitutions of He 3. Nature 334 406–410.CrossRefGoogle Scholar
Matsumura, M., Yahanda, S., Yasumura, S., Yutani, K. & Aiba, S. (1988). Role of tyrosine-80 in the stability of kanamycin nucleotidyltransferase analyzed by sitedirected mutagenesis. Eur.J. Biochem. 171, 715–720.CrossRefGoogle ScholarPubMed
Matsumura, M., Wozniak, J. A., Dao-Pin, S. & Matthews, B. W. (1989). Structural studies of mutants of T4 lysozyme that alter hydrophobic stabilization. J. Biol. Chem. 264, 16059–16066.CrossRefGoogle ScholarPubMed
Matthews, B. W. (1987). Genetic and structural analysis of the protein stability problem. Biochemistry 26, 6885–6888.CrossRefGoogle ScholarPubMed
Matthews, B. W. (1991). Mutational analysis of protein stability. Curr. Opin. Struc. Biol. 1, 17–21.CrossRefGoogle Scholar
Matthews, B. W., Nicholson, H. & Becktel, W. J. (1987). Enhanced protein thermostability from site-directed mutations that decrease the entropy of unfolding. Proc. Natl. Acad. Sci. USA 84, 6663–6667.CrossRefGoogle ScholarPubMed
Matthews, C. R. (1991). The mechanism of protein folding. Curr. Opin. Struc. Biol. I, 28–35.CrossRefGoogle Scholar
Mildvan, A. S., Weber, D. J. & Kuliopulis, A. (1992). Quantitative interpretations of double mutations of enzymes. Arch. Biochem. Biophys. 294, 327–340.CrossRefGoogle ScholarPubMed
Miller, W. T., Hou, Y. M. & Schimmel, P. (1991). Mutant aminoacyl-tRNA synthetase that compensates for a mutation in the major identity determinant of its tRNA. Biochemistry 30, 2635–2641.CrossRefGoogle ScholarPubMed
Monod, J., Wyman, J. & Changeux, J. P. (1965). On the nature of allosteric transitions: A plausible model. J. Mol. Biol. 12, 88–118.CrossRefGoogle ScholarPubMed
Murphy, D. J. & Benkovic, S. J. (1989). Hydrophobic interactions via mutants of Escherichia coli dihydrofolate reductase: separation of binding and catalysis. Biochemistry 28, 3025–3031.CrossRefGoogle ScholarPubMed
Nagai, K., Luisi, B., Shih, D., Miyazaki, G., Imai, K., Poyart, C., De Young, A., Kwiatkowsky, L., Noble, R. W., Lin, S.-H. & Yu, N. T. (1987). Distal residues in the oxygen binding site of haemoglobin studied by protein engineering. Nature 329 858–860.CrossRefGoogle ScholarPubMed
Nathans, J. (1990). Determinants of visual pigment absorbance: identification of the retinylidene Schiff's base counterion in bovine rhodopsin. Biochemistry 29, 9746–9752.CrossRefGoogle ScholarPubMed
Newton, C. J. & Kantrowitz, E. R. (1990). The regulatory subunit of Escherichia coli aspartate carbamoyltransferase may influence homotropic cooperativity and heterotropic interactions by a direct interaction with the loop containing residues 230–245 of the catalytic chain. Proc. Natl. Acad. Sci. USA 87, 2309–2313.CrossRefGoogle ScholarPubMed
Nicholson, H., Anderson, D. E., Dao-Pin, S. & Matthews, B. W. (1991). Analysis of the interaction between charged side chains and the α-helix dipole using designed thermostable mutants of phage T4 lysozyme. Biochemistry 30, 9816–9828.CrossRefGoogle ScholarPubMed
Nickbarg, E. B., Davenport, R. C., Petsko, G. A. & Knowles, J. R. (1988). Triosephosphate isomerase: removal of a putatively electrophilic histidine residue results in a subtle change in catalytic mechanism. Biochemistry 27, 5948–5960.CrossRefGoogle Scholar
Noren, C. J., Anthony-Cahill, S. J., Griffith, M. C. & Schultz, P. G. (1989). A general method for site-specific incorporation of unnatural amino acids into proteins. Science 244, 182–188.CrossRefGoogle ScholarPubMed
Oas, T. G. & Kim, P. S. (1988). A peptide model of a protein folding intermediate. Nature 336, 42–48.CrossRefGoogle ScholarPubMed
Otting, G., Liepinsh, E. & Wüthrich, K. (1991). Protein hydration in aqueous solution. Science 254, 974–980.CrossRefGoogle ScholarPubMed
Pace, C. N. (1990). Conformational stability of globular proteins. TIBS 15, 14–17.Google ScholarPubMed
Pace, C. N., Grimsley, G. R., Thomson, J. A. & Barnett, B. J. (1988). Conformational stability and activity of ribonuclease T, with zero, one, and two intact disulfide bonds. J. Biol. Chem. 263, 11820–11825.CrossRefGoogle Scholar
Pakula, A. A. & Sauer, R. T. (1989). Genetic analysis of protein stability and function. Ann. Rev. Genet. 23, 289–310.CrossRefGoogle ScholarPubMed
Pakula, A. A. & Sauer, R. T. (1990). Reverse hydrophobic effects relieved by aminoacid substitutions at a protein surface. Nature 344, 363–364.CrossRefGoogle Scholar
Pantoliano, M. W., Ladner, R. C., Bryan, P. N., Rollence, M. L., Wood, J. F. & Poulos, T. L. (1987). Protein engineering of subtilisin BPN: enhanced stabilization through the introduction of two cysteines to form a disulfide bond. Biochemistry 26 2077–2082.CrossRefGoogle Scholar
Perry, L. J. & Wetzel, R. (1984). Disulfide bond engineered into T4 lysozyme: stabilization of the protein toward thermal inactivation. Science 226, 555–557.CrossRefGoogle ScholarPubMed
Perry, K. M., Onuffer, J. J., Touchette, N. A., Herndon, C. S., Gittelman, M. S., Matthews, C. R., Chen, J.-T., Mayer, R. J., Taira, K., Benkovic, S. J., Howell, E. E. & Kraut, J. (1987). Effect of single amino acid replacements on the folding and stability of dihydrofolate reductase from Escherichia coli. Biochemistry 26, 2674–2682.CrossRefGoogle ScholarPubMed
Perry, K. M., Onuffer, J. J., Gittelman, M. S., Barmat, L. & Matthews, C. R. (1989). Long-range electrostatic interactions can influence the folding, stability, and cooperativity of dihydrofolate reductase. Biochemistry 28, 7961–7968.CrossRefGoogle ScholarPubMed
Perutz, M. F. (1989). Mechanisms of cooperativity and allosteric regulation in proteins. Q. Rev. Biophys. 22, 139–236.CrossRefGoogle ScholarPubMed
Petsko, G. A. & Ringe, D. (1984). Fluctuations in protein structure from X-ray diffraction. A. Rev. Biophys. Bioenerg. 13, 331–371.CrossRefGoogle ScholarPubMed
Ponder, J. W. & Richards, F. M. (1987). Tertiary templates for proteins. Use of packing criteria in the enumeration of allowed sequences for different structural classes. J. Mol. Biol. 193, 775–791.CrossRefGoogle Scholar
Prevost, M., Wodak, S. J., Tidor, B. & Karplus, M. (1991). Contribution of the hydrophobic effect to protein stability: analysis based on simulations of the Ile-96 → Ala mutation in barnase. Proc. Natl. Acad. Sci. USA 88, 10880–10884.CrossRefGoogle ScholarPubMed
Privalov, P. L. (1979). Stability of proteins. Small globular proteins. Adv. Protein Chem. 33, 167–241.CrossRefGoogle ScholarPubMed
Rao, S. N., Singh, U. C., Bash, P. A. & Kollman, P. A. (1987). Free energy perturbation calculations on binding and catalysis after mutating Asn 155 in subtilisin. Nature 328, 551–554.CrossRefGoogle ScholarPubMed
Reidhaar-Olson, J. F. & Sauer, R. T. (1988). Combinatorial cassette mutagenesis as a probe of the informational content of protein sequences. Science 241, 53–57.CrossRefGoogle ScholarPubMed
Richards, F. (1986). Protein design: are we ready? In Protein Structure, Folding, and Design (ed. Oxender, D. L.), pp. 171–196. New York: Alan R. Liss.Google Scholar
Richards, F. M. (1977). Areas, volumes, packing, and protein structure. A. Rev. Biophys. Bioeng. 6, 151–176.CrossRefGoogle ScholarPubMed
Riggs, A. F. (1991). Hemoglobins. Curr. Opin. Struc. Biol. I, 915–921.CrossRefGoogle Scholar
Roder, H., Elöve, G. & Englander, S. W. (1988). Structural characterization of folding intermediates in cytochrome c by H-exchange labelling and proton NMR. Nature 335 700–704.CrossRefGoogle ScholarPubMed
Sali, D., Bycroft, M. & Fersht, A. R. (1988). Stabilization of protein structure by interaction of α-helix dipole with a charged side chain. Nature 335, 740–743.Google ScholarPubMed
Sandberg, W. S. & Terwilliger, T. C. (1989). Influence of interior packing and hydrophobicity on the stability of a protein. Science 245, 54–57.CrossRefGoogle ScholarPubMed
Sandberg, W. S. & Terwilliger, T. C. (1991 a). Repacking protein interiors. TIBTECH 9, 59–63.CrossRefGoogle ScholarPubMed
Sandberg, W. S. & Terwilliger, T. C. (1991 b). Energetics of repacking a protein interior. Proc. Natl. Acad. Sci. USA 88, 1706–1710.CrossRefGoogle ScholarPubMed
Santoro, M. M. & Bolen, D. W. (1988). Unfolding free energy changes determined by the linear extrapolation method. 1. Unfolding of phenylmethanesulfonyl α-chymotrypsin using different denaturants. Biochemistry 27, 8063–8068.CrossRefGoogle ScholarPubMed
Sauer, R. T., Hehir, K., Stearman, R. S., Weiss, M. A., Jeitler-Nilsson, A., Suchanek, E. G. & Pabo, C. O. (1986). An engineered intersubunit disulfide enhances the stability and DNA binding of the N-terminal domain of lambda repressor. Biochemistry 25, 5992–5998.CrossRefGoogle ScholarPubMed
Sauer, U. H., Dao-Pin, S. & Matthews, B. W. (1992). Tolerance of T4 lysozyme to proline substitutions within the longer interdomain α-helix illustrates the adaptability of proteins to potentially destabilizing lesions. J. Biol. Chem. 267, 2393–2399.CrossRefGoogle Scholar
Schellman, J. A. (1987). The thermodynamic stability of proteins. A. Rev. Biophys. Biophys. Chem. 16, 115–137.CrossRefGoogle ScholarPubMed
Schimmel, P. (1990). Hazards and their exploitation in the applications of molecular biology to structure-function relationships. Biochemistry 29, 9495–9502.CrossRefGoogle ScholarPubMed
Schirmer, T. & Evans, P. R. (1990). Nature (Lond.) 343, 140–145.CrossRefGoogle Scholar
Serpersu, E. H., Shortle, D. & Mildvan, A. S. (1986). Kinetic and magnetic resonance studies of effects of genetic substitution of a Ca2+-liganding amino acid in staphylococcal nuclease. Biochemistry 25, 68–77.CrossRefGoogle ScholarPubMed
Serrano, L., Bycroft, M. & Fersht, A. R. (1991). Aromatic-aromatic interactions and protein stability investigation by double-mutant cycles. J. Mol. Biol. 218, 465–475.CrossRefGoogle ScholarPubMed
Serrano, L. & Fersht, A. R. (1989). Capping and α-helix stability. Nature 342 296–299.CrossRefGoogle ScholarPubMed
Serre, M. C., Teschner, W. & Garel, J. R. (1990). Specific suppression of heterotropic interactions in phosphofructokinase by the mutation of leucine 178 into tryptophan. J. Biol. Chem. 265, 12146–12148.CrossRefGoogle ScholarPubMed
Shih, H. H. L., Brady, J. & Karplus, M. (1985). Structure of proteins with single-site mutations: A minimum perturbation approach. Proc. Natl. Acad. Sci. USA 82 1697–1700.CrossRefGoogle ScholarPubMed
Shirley, B. A., Stanssens, P., Steyaert, J. & Pace, C. N. (1989). Conformational stability and activity of ribonuclease Ti and mutants. J. Biol. Chetn. 264, 11621–11625.CrossRefGoogle Scholar
Shortle, D. (1989). Probing The determinants of protein folding and stability with amino acid substitutions. J. Biol. Chem. 264, 5315–5318.CrossRefGoogle ScholarPubMed
Shortle, D. & Meeker, A. K. (1986). Mutant forms of staphylococcal nuclease with altered patterns of guanidine hydrochloride and urea denaturation. Proteins: Struct. Fund. Genet. I, 81–89.CrossRefGoogle Scholar
Shortle, D. & Meeker, A. K. (1989). Residual structure in large fragments of staphylococcal nuclease: effects of amino acid substitutions. Biochemistry 28, 936–944.CrossRefGoogle ScholarPubMed
Shortle, D., Meeker, A. K. & Freire, E. (1988). Stability mutants of staphylococcal nuclease: large compensating enthalpy-entropy changes for the reversible denaturation reaction. Biochemistry 27, 4761–4768.CrossRefGoogle ScholarPubMed
Shortle, D., Meeker, A. K. & Gerring, S. L. (1989). Effects of denaturants at low concentrations on the reversible denaturation of staphylococcal nuclease. Arch. Biochem. Biophys. 272, 103–113.CrossRefGoogle ScholarPubMed
Shortle, D., Stites, W. E. & Meeker, A. K. (1990). Contributions of the large hydrophobic amino acids to the stability of staphylococcal nuclease. Biochemistry 29, 8033–8041.CrossRefGoogle Scholar
Shortle, D., Chan, H. S. & Dill, K. A. (1992). Modeling the effects of mutations on the denatured states of proteins. Protein Science I, 201–215.CrossRefGoogle Scholar
Sondek, J. & Shortle, D. (1990). Accommodation of single amino acid insertions by the native state of staphylococcal nuclease. Proteins: Struct. Fund. Genet. 7, 299–305.CrossRefGoogle ScholarPubMed
Sondek, J. & Shortle, D. (1992). Structural and energetic differences between insertions and substitutions in staphylococcal nuclease. Proteins : Struct. Fund. Genet. 13 (still in press).CrossRefGoogle ScholarPubMed
Starzyk, R. M., Burbaum, J. J. & Schimmel, P. (1989). Insertion of new sequences into the catalytic domain of an enzyme. Biochemistry 28, 8479–8484.CrossRefGoogle ScholarPubMed
Stevens, R. C., Chook, Y. M., Cho, C. Y., Lipscomb, W. N. & Kantrowitz, E. R. (1991). Escherichia coli aspartate carbamoyltransferase : the probing of crystal structure analysis via site-specific mutagenesis. Protein Engineering 4, 391–408.CrossRefGoogle ScholarPubMed
Stites, W. E., Gittis, A. G., Lattman, E. E. & Shortle, D. (1991). In a staphylococcal nuclease mutant the side-chain of a lysine replacing valine 66 is fully buried in the hydrophobic core. J. Mol. Biol. 221, 7–14.Google Scholar
Strop, P., Sedlacek, J., Stys, J., Kaderabkova, Z., Blaha, I., Pavlickova, L., Pohl, J., Fabry, M., Kostka, V., Newman, M., Frazao, C., Shearer, A., Tickle, I. J. & Blundell, T. L. (1990). Engineering enzyme subsite specificity: preparation, kinetic characterization, and X-ray analysis at 2°o-Å resolution of ValIIIPhe site-mutated calf chymosin. Biochemistry 29, 9863–9871.CrossRefGoogle Scholar
Sturtevant, J. M. (1992). Effects of mutations on the thermodynamic properties of proteins. Am. Chem. Soc. Symposia series (still in press).Google Scholar
Sturtevant, J. M., Yu, M-H., Haase-Pettingell, C. & King, J. (1989). Thermostability of temperature-sensitive folding mutants of the P22 tailspike protein. J. Biol. Chem. 264, 10693–10698.CrossRefGoogle ScholarPubMed
Tanford, C. (1968). Protein denaturation. Adv. Protein Chem. 23, 121–282.CrossRefGoogle ScholarPubMed
Tanford, C. (1970). Protein denaturation. Adv. Protein Chem. 24, 1–95.CrossRefGoogle ScholarPubMed
Tidor, B. & Karplus, M. (1991). Simulation analysis of the stability mutant R96H of T4 lysozyme. Biochemistry 30, 3217–3228.CrossRefGoogle ScholarPubMed
Udgaonkar, J. B. & Baldwin, R. L. (1988). NMR evidence for an early framework intermediate on the folding pathway of ribonuclease A. Nature 335, 694–699.CrossRefGoogle ScholarPubMed
Van Iwaarden, P. R., Pastore, J. C., Konings, W. N. & Kaback, H. R. (1991). Construction of a functional lactose permease devoid of cysteine residues. Biochemistry 30, 9595–9600.CrossRefGoogle ScholarPubMed
Villafranca, J. E., Howell, E. E., Oatley, S. J., Xuong, N-H. & Kraut, J. (1987). An engineered disulfide bond in dihydrofolate reductase. Biochemistry 26, 2182–2189.CrossRefGoogle ScholarPubMed
Wang, J., Mauro, J. M., Edwards, S. L., Oatley, S. J., Fishel, L. A., Ashford, V. A., Xuong, N-H. & Kraut, J. (1990). X-ray structures of recombinant yeast cytochrome c peroxidase and three heme-cleft mutants prepared by site-directed mutagenesis. Biochemistry 29, 7160–7173.CrossRefGoogle ScholarPubMed
Warren, M. S., Brown, K. A., Farnum, M. F., Howell, E. E. & Kraut, J. (1991). Investigation of the functional role of tryptophan-22 in Escherichia coli dihydrofolate reductase by site-directed mutagenesis. Biochemistry 30, 11092–11103.CrossRefGoogle ScholarPubMed
Weaver, L. H., Gray, T. M., Grutter, M. G., Anderson, D. E., Wozniak, J. A., Dahlquist, F. W. & Matthews, B. W. (1989). High-resolution structure of the temperature-sensitive mutant of phage lysozyme, Arg 96 → His. Biochemistry 28, 3793–3797.CrossRefGoogle ScholarPubMed
Weiner, H., White, W. N., Hoare, D. G. & Koshland, D. E. Jr., (1966). The formation of anhydrochymotrypsin by removing the elements of water from the serine at the active site. J. Am. Chem. Soc. 88, 3851–3859.CrossRefGoogle ScholarPubMed
Wells, J. A. (1990). Additivity of mutational effects in proteins. Biochemistry 29 8509–8517.CrossRefGoogle ScholarPubMed
Wetzel, R., Perry, L. J., Baase, W. A. & Becktel, W. J. (1988). Disulfide bonds and thermal stability in T4 lysozyme. Proc. Natl. Acad. Sci. USA 85, 401–405.CrossRefGoogle ScholarPubMed
Wilke, M. E., Higaki, J. N., Craik, C. S. & Fletterick, R. J. (1991a). Crystal structure of rat trypsin-S195C at — 150 °C. Analysis of low activity of recombinant and semisynthetic thiol proteases. J. Mol. Biol. 219, 511–523.CrossRefGoogle ScholarPubMed
Wilke, M. E., Higaki, J. N., Craik, C. S. & Fletterick, R. J. (1991 b). Crystallographic analysis of trypsin-G226A. A specificity pocket mutant of rat trypsin with altered binding and catalysis. J. Mol. Biol. 219, 525–532.CrossRefGoogle ScholarPubMed
Wüthrich, K. (1990). Protein structure determination in solution by NMR spectroscopy. J. Biol. Chem. 265, 22055–22062.CrossRefGoogle ScholarPubMed
Xi, X. G., Van Vliet, F., Ladjimi, M. M., De Wannemaeker, B., De Staercke, C., Pierard, A., Glansdorff, N., Herve, G. & Cunin, R. (1990). Co-operative interactions between the catalytic sites in Escherichia coli aspartate transcarbamylase. J. Mol. Biol. 216, 375–384.Google ScholarPubMed
Yutani, K., Ogasahara, K., Tsujita, T. & Sugino, Y. (1987). Dependence of conformational stability on hydrophobicity of the amino acid residue in a series of variant proteins substituted at a unique position of tryptophan synthase a subunit. Proc. Natl. Acad. Sci. USA 84, 4441–4444.CrossRefGoogle Scholar
Zebala, J. & Barany, F. (1991). Mapping catalytically important regions of an enzyme using two-codon insertion mutagenesis: a case study correlating β-lactamase mutants with the three-dimensional structure. Gene 100, 51–57.CrossRefGoogle ScholarPubMed
Zhang, X-J., Baase, W. A. & Matthews, B. W. (1991). Toward a simplification of the protein folding problem: a stabilizing polyalanine α-helix engineered in T4 lysozyme. Biochemistry 30, 2012–2017.CrossRefGoogle Scholar
Zoller, M. J. (1991). New molecular biology methods for protein engineering. Curr. Opin. Struc. Biol. 1, 605–610.CrossRefGoogle Scholar