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Folding at the speed limit

Nature volume 423, pages 193–197 (2003)Cite this article

Abstract

Many small proteins seem to fold by a simple process explicable by conventional chemical kinetics and transition-state theory. This assumes an instant equilibrium between reactants and a high-energy activated state1. In reality, equilibration occurs on timescales dependent on the molecules involved, below which such analyses break down1. The molecular timescale, normally too short to be seen in experiments, can be of a significant length for proteins. To probe it directly, we studied very rapidly folding mutants of the five-helix bundle protein λ6–85, whose activated state is significantly populated during folding. A time-dependent rate coefficient below 2 µs signals the onset of the molecular timescale, and hence the ultimate speed limit for folding2. A simple model shows that the molecular timescale represents the natural pre-factor for transition state models of folding.

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Figure 1: Spectroscopy of mutant proteins.
Figure 2: Relaxation after temperature jumps.
Figure 3: Rate coefficient determination.
Figure 4: One-dimensional free-energy model used to reproduce the λQ33Y and λD14A data as a function of temperature.

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References

  1. Berne, B. J. in Activated Barrier Crossing: Applications in Physics, Chemistry and Biology (eds Hänggi, P. & Fleming, G. R.) 82–119 (World Scientific, Singapore, 1993)

    Book  Google Scholar 

  2. Hagen, S. J., Hofrichter, J., Szabo, A. & Eaton, W. A. Diffusion-limited contact formation in unfolded cytochrome c: Estimating the maximum rate of protein folding. Proc. Natl Acad. Sci. USA 93, 11615–11617 (1996)

    Article  ADS  CAS  Google Scholar 

  3. Lapidus, L. J., Eaton, W. A. & Hofrichter, J. Measuring the rate of intramolecular contact formation in polypeptides. Proc. Natl Acad. Sci. USA 97, 7220–7225 (2000)

    Article  ADS  CAS  Google Scholar 

  4. Bieri, O. et al. The speed limit for protein folding measured by triplet–triplet energy transfer. Proc. Natl Acad. Sci. USA 96, 9597–9601 (1999)

    Article  ADS  CAS  Google Scholar 

  5. Yeh, I. C. & Hummer, G. Peptide loop-closure kinetics from microsecond molecular dynamics simulations in explicit solvent. J. Am. Chem. Soc. 124, 6563–6568 (2002)

    Article  CAS  Google Scholar 

  6. Thirumalai, D. Time scales for the formation of the most probable tertiary contacts in proteins with applications to cytochrome C. J. Phys. Chem. B 103, 608–610 (1999)

    Article  CAS  Google Scholar 

  7. Portman, J. J., Takada, S. & Wolynes, P. G. Microscopic theory of protein folding rates. II. Local reaction coordinates and chain dynamics. J. Chem. Phys. 114, 5082–5096 (2001)

    Article  ADS  CAS  Google Scholar 

  8. Portman, J. J., Takada, S. & Wolynes, P. G. Microscopic theory of protein folding rates. I. Fine structure of the free energy profile and folding routes from a variational approach. J. Chem. Phys. 114, 5069–5081 (2001)

    Article  ADS  CAS  Google Scholar 

  9. Hagen, S. J., Hofrichter, J. & Eaton, W. A. Rate of intrachain diffusion of unfolded cytochrome c. J. Phys. Chem. B 101, 2352–2365 (1997)

    Article  CAS  Google Scholar 

  10. Schuler, B., Lipman, E. A. & Eaton, W. A. Probing the free-energy surface for protein folding with single-molecule fluorescence spectroscopy. Nature 419, 743–747 (2002)

    Article  ADS  CAS  Google Scholar 

  11. Gruebele, M., Sabelko, J., Ballew, R. & Ervin, J. Laser temperature jump induced protein refolding. Acc. Chem. Res. 31, 699–707 (1998)

    Article  CAS  Google Scholar 

  12. Metzler, R., Klafter, J. & Jortner, J. Hierarchies and logarithmic oscillations in the temporal relaxation patterns of proteins and other complex systems. Proc. Natl Acad. Sci. USA 96, 11085–11089 (1999)

    Article  ADS  CAS  Google Scholar 

  13. 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 Ala → Gly substitutions. Nature Struct. Biol. 4, 305–309 (1997)

    Article  CAS  Google Scholar 

  14. Ghaemmaghami, S., Word, J. M., Burton, R. E., Richardson, J. S. & Oas, T. G. Folding kinetics of a fluorescent variant of monomeric lambda represssor. Biochemistry 37, 9179–9185 (1998)

    Article  CAS  Google Scholar 

  15. Myers, J. M. & Oas, T. G. Contribution of a buried hydrogen bond to lambda repressor folding kinetics. Biochemistry 38, 6761–6768 (1999)

    Article  CAS  Google Scholar 

  16. Chang, I. J., Lee, J. C., Winkler, J. R. & Gray, H. B. The protein-folding speed limit: intrachein diffusion times set by electron transfer rates in denatured Ru(NH3)5(His-33)-Zn-cytochrome c. Proc. Natl Acad. Sci. USA 100, 3838–3840 (2003)

    Article  ADS  CAS  Google Scholar 

  17. Silow, M. & Oliveberg, M. Transient aggregates in protein folding are easily mistaken for folding intermediates. Proc. Natl Acad. Sci. USA 94, 6084–6086 (1997)

    Article  ADS  CAS  Google Scholar 

  18. Sabelko, J., Ervin, J. & Gruebele, M. Observation of strange kinetics in protein folding. Proc. Natl Acad. Sci. USA 96, 6031–6036 (1999)

    Article  ADS  CAS  Google Scholar 

  19. Garcia-Mira, M. M., Sadqi, M., Fischer, N., Sanchez-Ruiz, J. M. & Muñoz, V. Experimental identification of downhill protein folding. Science 298, 2191–2195 (2002)

    Article  ADS  CAS  Google Scholar 

  20. Silow, M. & Oliveberg, M. High-energy channeling in protein folding. Biochemistry 36, 7633–7637 (1997)

    Article  CAS  Google Scholar 

  21. Pappenberger, G., Saudan, C., Becker, M., Merbach, A. E. & Kiefhaber, T. Denaturant-induced movement of the transition state of protein folding revealed by high-pressure stopped-flow measurements. Proc. Natl Acad. Sci. USA 97, 17–22 (2002)

    Article  ADS  Google Scholar 

  22. Snow, C. D., Nguyen, H., Pande, V. S. & Gruebele, M. Absolute comparison of simulated and experimental protein-folding dynamics. Nature 420, 102–106 (2002)

    Article  ADS  CAS  Google Scholar 

  23. Mayor, U. et al. The complete folding pathway of a protein from nanoseconds to microseconds. Nature 421, 863–867 (2003)

    Article  ADS  CAS  Google Scholar 

  24. Ervin, J., Sabelko, J. & Gruebele, M. Submicrosecond real-time fluorescence detection: application to protein folding. J. Photochem. Photobiol. B 54, 1–15 (2000)

    Article  CAS  Google Scholar 

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Acknowledgements

We thank T. Oas for suggesting many helpful lambda repressor mutations. This work was supported by an Alumni Scholarship and a Camille Dreyfus Teacher–Scholar Award to M.G.

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  1. Center for Biophysics and Computational Biology, University of Illinois, Urbana, Illinois, 61801, USA

    Wei Yuan Yang & Martin Gruebele

  2. Departments of Chemistry and Physics, University of Illinois, Urbana, Illinois, 61801, USA

    Martin Gruebele

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  1. Wei Yuan Yang
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Correspondence to Martin Gruebele.

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Yang, W., Gruebele, M. Folding at the speed limit. Nature 423, 193–197 (2003). https://doi.org/10.1038/nature01609

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