Abstract
The folding and unfolding kinetics of the B-domain of staphylococcal protein A, a small three-helix bundle protein, were probed by NMR. The lineshape of a single histidine resonance was fit as a function of denaturant to give folding and unfolding rate constants. The B-domain folds extremely rapidly in a two-state manner, with a folding rate constant of 120,000 sā1, making it one of the fastest-folding proteins known. Diffusion-collision theory predicts folding and unfolding rate constants that are in good agreement with the experimental values. The apparent rate constant as a function of denaturant ('chevron plot') is predicted within an order of magnitude. Our results are consistent with a model whereby fast-folding proteins utilize a diffusion-collision mechanism, with the preorganization of one or more elements of secondary structure in the unfolded protein.
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References
Baldwin, R.L. Protein folding from 1961ā1982. Nature Struct. Biol. 6, 814ā 817 (1999).
Jackson, S.E. How do small, single-domain proteins fold? Folding Des. 3, R81ā> R91 (1998).
Dobson, C.M. & Karplus, M. The fundamentals of protein folding: bringing together theory and experiment. Curr. Opin. Struct. Biol. 9, 92ā 101 (1999).
Alm, E. & Baker, D. Matching theory and experiment in protein folding. Curr. Opin. Struct. Biol. 9, 189ā 196 (1999).
Daggett, V. Long timescale simulations. Curr. Opin. Struct. Biol. 10, 160ā 164 (2000).
Kolinski, A. & Skolnick, J. Monte Carlo simulations of protein folding. II. Application to protein A, ROP, and crambin. Proteins 18, 353ā 366 (1994).
Alonso, D.O.V. & Daggett, V. Staphylococcal protein A: unfolding pathways, unfolded states, and differences between the B and E domains. Proc. Natl. Acad. Sci. USA 97, 133ā 138 (2000).
Guo, Z., Brooks, C.L., 3rd & Boczko, E.M. Exploring the folding free energy surface of a three-helix bundle protein. Proc. Natl. Acad. Sci. USA 94, 10161ā 10166 (1997).
Shea, J.E., Onuchic, J.N. & Brooks, C.L., III. Exploring the origins of topological frustration: design of a minimally frustrated model of fragment B of protein A. Proc. Natl. Acad. Sci. USA 96, 12512ā 12517 (1999).
Zhou, Y. & Karplus, M. Interpreting the folding kinetics of helical proteins. Nature 401, 400ā 403 (1999).
Bai, Y., Karimi, A., Dyson, H.J. & Wright, P.E. Absence of a stable intermediate on the folding pathway of protein A. Protein Sci. 6, 1449ā 1457 (1997).
Ropson, I.J. & Frieden, C. Dynamic NMR spectral analysis and protein folding: identification of a highly populated folding intermediate of rat intestinal fatty acid-binding protein by 19F NMR. Proc. Natl. Acad. Sci. USA 89, 7222ā 7226 (1992).
Huang, G.S. & Oas, T.G. Submillisecond folding of monomeric Ī» repressor. Proc. Natl. Acad. Sci. USA 92, 6878ā 6882 (1995).
Burton, R.E., Huang, G.S., Daugherty, M.A., Fullbright, P.W. & Oas, T.G. Microsecond protein folding through a compact transition state. J. Mol. Biol. 263, 311ā 322 (1996).
Burton, R.E., Huang, G.S., Daugherty, M.A., Calderone, T.L. & Oas, T.G. The energy landscape of a fast-folding protein mapped by AlaāGly substitutions. Nature Struct. Biol. 4, 305ā 310 (1997).
Myers, J.K. & Oas, T.G. Contribution of a buried hydrogen bond to Ī» repressor folding kinetics. Biochemistry 38, 6761ā 6768 (1999).
Kuhlman, B., Boice, J.A., Fairman, R. & Raleigh, D.P. Structure and stability of the N-terminal domain of the ribosomal protein L9: evidence for rapid two-state folding. Biochemistry 37, 1025ā 1032 (1998).
Spector, S. & Raleigh, D.P. Submillisecond folding of the peripheral subunit-binding domain. J. Mol. Biol. 293, 763ā 768 (1999).
Burton, R.E., Myers, J.K. & Oas, T.G. Protein folding dynamics: quantitative comparison between theory and experiment. Biochemistry 37, 5337ā 5343 (1998).
Myers, J.K. & Oas, T.G. Reinterpretation of GCN4-p1 folding kinetics: partial helix formation precedes dimerization in coiled coil folding. J. Mol. Biol. 289, 205ā 209 (1999).
Pappu, R.V. & Weaver, D.L. The early folding kinetics of apomyoglobin. Protein Sci. 7, 480ā 490 (1998).
Myers, J.K., Pace, C.N. & Scholtz, J.M. Denaturant m values and heat capacity changes: relation to changes in accessible surface areas of protein unfolding. Protein Sci. 4, 2138ā 2148 (1995).
Burton, R.E., Busby, R.S. & Oas, T.G. ALASKA: a mathematical package for two-state kinetic analysis of protein folding reactions. J. Biomol. NMR 11, 355ā 360 (1998).
Carr, H.Y. & Purcell, E.M. Effects of diffusion on free precession in nuclear magnetic resonance experiments. Physics Review 94, 630ā 638 (1954).
Meiboom, S. & Gill, D. Modified spin-echo method for measuring nuclear relaxation times. Rev. Sci. Instrum. 29, 688ā 691 (1958).
Orekhov, V., Pervushin, K.V. & Arseniev, A.S. Backbone dynamics of (1-71)bacterioopsin studied by two-dimensional 1H- 15N NMR spectroscopy. Eur. J. Biochem. 219, 887ā 896 (1994).
Wittung-Stafshede, P., Lee, J.C., Winkler, J.R. & Gray, H.B. Cytochrome b562 folding triggered by electron transfer: approaching the speed limit for formation of a four-helix-bundle protein. Proc. Natl. Acad. Sci. USA 96, 6587ā 6590 (1999).
Chen, B.-L., Baase, W.A., Nicholson, H. & Schellman, J.A. Folding kinetics of T4 lysozyme and nine mutants at 12 Ā°C. Biochemistry 31, 1464ā 1476 (1992).
Tanford, C. Protein denaturation, Part C. Adv. Protein Chem. 24, 1ā 95 (1970).
Reimer, U. et al. Side-chain effects on peptidyl-prolyl cis/trans isomerisation. J. Mol. Biol. 279, 449ā 460 (1998).
Karplus, M. & Weaver, D.L. Protein folding dynamics: the diffusion-collision model and experimental data. Protein Sci. 3, 650ā 668 (1994).
Bashford, D., Cohen, F.E., Karplus, M., Kuntz, I.D. & Weaver, D.L. Diffusion-collision model for the folding kinetics of myoglobin. Proteins 4, 211ā 227 (1988).
Duan, Y. & Kollman, P.A. Pathways to a protein folding intermediate observed in a 1-microsecond simulation in aqueous solution. Science 282, 740ā 744 (1998).
Jacob, M., Geeves, M., Holtermann, G. & Schmid, F.X. Diffusional barrier crossing in a two-state protein folding reaction. Nature Struct. Biol. 6, 923ā 926 (1999).
Plaxco, K.W. & Baker, D. Limited internal friction in the rate-limiting step of a two-state protein folding reaction. Proc. Natl. Acad. Sci. USA 95, 13591ā 13596 (1998).
Goldberg, J.M. & Baldwin, R.L. A specific transition state for S-peptide combining with folded S-protein and then refolding. Proc. Natl. Acad. Sci. USA 96, 2019ā 2024 (1999).
Zitzewitz, J.A., Ibarra-Molero, B., Fishel, D.R., Terry, K.L. & Matthews, C.R. Preformed secondary structure drives the association reaction of GCN4-p1, a model coiled coil system. J. Mol. Biol. 296, 1105ā 1116 (2000).
Moran, L.B., Schneider, J.P., Kentsis, A., Reddy, G.A. & Sosnick, T.R. Transition state heterogeneity in GCN4 coiled coil folding studied by using multisite mutations and crosslinking. Proc. Natl. Acad. Sci. USA 96, 10699ā 10704 (1999).
Fersht, A.R. Optimization of rates of protein folding: the nucleation-condensation mechanism and its implications. Proc. Natl. Acad. Sci. USA 92, 10869ā 10873 (1995).
Fersht, A.R. Transition state structure as a unifying basis in protein-folding mechanisms: contact order, chain topology, stability, and the extended nucleus mechanism. Proc. Natl. Acad. Sci. USA 97, 1525ā 1529 (2000).
Mirny, L.A., Abkevich, V.I. & Shakhnovich, E.I. How evolution makes proteins fold quickly. Proc. Natl. Acad. Sci. USA 95, 4976ā 4981 (1998).
Plaxco, K.W., Simons, K.T. & Baker, D. Contact order, transition state placement, and the refolding rates of single domain proteins. J. Mol. Biol. 277, 985ā 994 (1998).
Alm, E. & Baker, D. Prediction of protein folding mechanisms from free-energy landscapes derived from native structures. Proc. Natl. Acad. Sci. USA 96, 11305ā 11310 (1999).
Debe, D.A. & Goddard, W.A. First priciples prediction of protein folding rates. J. Mol. Biol. 294, 619ā 625 (1999).
Galzitkaya, O. & Finkelstein, A. A theoretical search for folding/unfolding nuclei in three-dimensional protein structures. Proc. Natl. Acad. Sci. USA 96, 11299ā 11304 (1999).
Munoz, V. & Eaton, W.A. A simple model for the calculating the kinetics of protein folding from three-dimensional structures. Proc. Natl. Acad. Sci. USA 96, 11311ā 11316 (1999).
Clementi, C., Nymeyer, H. & Onuchic, J.N. Topological and energetic factors: what determines the structural details of the transition state ensemble and 'en-route' intermediates for protein folding? An investigation for small globular proteins. J. Mol. Biol. 298, 937ā 953 (2000).
Dalessio, P.M. & Ropson, I.J. Ī²-sheet proteins with nearly identical structures have different folding intermediates. Biochemistry 39, 860ā 871 (2000).
Llinas, M., Gillespie, B., Dahlquist, F.W. & Marqusee, S. The energetics of T4 lysozyme reveal a hierarchy of conformations. Nature Struct. Biol. 6, 1072ā 1078 (1999).
Baldwin, R.L. & Rose, G.D. Is protein folding hierarchic? I. Local structure and peptide folding. Trends Biochem. Sci. 24, 26ā 33 (1999).
Baldwin, R.L. & Rose, G.D. Is protein folding hierarchic? II. Folding intermediates and transition states. Trends Biochem. Sci. 24, 77ā 83 (1999).
Srinivasan, R. & Rose, G.D. A physical basis for protein secondary structure. Proc. Natl. Acad. Sci. USA 96, 14258ā 14263 (1999).
Przytycka, T., Aurora, R. & Rose, G.D. A protein taxonomy based on secondary structure. Nature Struct. Biol. 6, 672ā 682 (1999).
Pace, C.N., Vajdos, F., Fee, L., Grimsley, G. & Gray, T. How to measure and predict the molar absorption coefficient of a protein. Protein Sci. 4, 2411ā 2423 (1995).
Pace, C.N. & Scholtz, J.M. In Protein structure: a practical approach (ed. Crieghton, T.E.) 299ā 321 (IRL Press, Oxford; 1997).
Tashiro, M. et al. High-resolution solution NMR structure of the Z domain of staphylococcal protein A. J. Mol. Biol. 272, 573ā 590 (1997).
Rohl, C.A., Chakrabartty, A. & Baldwin, R.L. Helix propagation and N-cap propensities of the amino acids measured in alanine-based peptides in 40% volume trifluoroethanol. Protein Sci. 5, 2623ā 2637 (1996).
Zimm, B.H. & Bragg, J.K. Theory of the phase transitions between helix and random coil in polypeptide chains. J. Chem. Phys. 31, 526ā 535 (1959).
Scholtz, J.M. et al. Calorimetric determination of the enthalpy change for the Ī±-helix to coil transition of an alanine peptide in water. Proc. Natl. Acad. Sci. USA 88, 2854ā 2858 (1991).
Smith, J.S. & Scholtz, J.M. Guanidine hydrochloride unfolding of peptide helices: separation of denaturant and salt effects. Biochemistry 35, 7292ā 7297 (1996).
Kraulis, P.J. MOLSCRIPT: A program to produce both detailed and schematic plots of protein structures. J. Appl. Crystall. 24, 946ā 950 (1991).
Acknowledgements
We are grateful to P. Wright for the gift of the plasmid expressing BdpA and L. Tennant for technical assistance. We also wish to thank R. Burton for the original suggestion of studying BdpA, and D. Weaver, R. Pappu, M. Karplus, G. Rose, F. Schmid, G. Hammes and the Oas lab group for helpful discussions. We thank the NIH for financial support.
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Myers, J., Oas, T. Preorganized secondary structure as an important determinant of fast protein folding. Nat Struct Mol Biol 8, 552ā558 (2001). https://doi.org/10.1038/88626
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DOI: https://doi.org/10.1038/88626
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