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The folding cooperativity of a protein is controlled by its chain topology

Nature volume 465, pages 637–640 (2010)Cite this article

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Abstract

The three-dimensional structures of proteins often show a modular architecture comprised of discrete structural regions or domains. Cooperative communication between these regions is important for catalysis, regulation and efficient folding; lack of coupling has been implicated in the formation of fibrils and other misfolding pathologies1. How different structural regions of a protein communicate and contribute to a protein’s overall energetics and folding, however, is still poorly understood. Here we use a single-molecule optical tweezers approach to induce the selective unfolding of particular regions of T4 lysozyme and monitor the effect on other regions not directly acted on by force. We investigate how the topological organization of a protein (the order of structural elements along the sequence) affects the coupling and folding cooperativity between its domains. To probe the status of the regions not directly subjected to force, we determine the free energy changes during mechanical unfolding using Crooks’ fluctuation theorem. We pull on topological variants (circular permutants) and find that the topological organization of the polypeptide chain critically determines the folding cooperativity between domains and thus what parts of the folding/unfolding landscape are explored. We speculate that proteins may have evolved to select certain topologies that increase coupling between regions to avoid areas of the landscape that lead to kinetic trapping and misfolding.

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Figure 1: Experimental set-up and structure of *T4L variants.
Figure 2: Unfolding and refolding force-extension curves of WT*T4L and CP13*T4L variants.
Figure 3: The dynamic force spectrum of *T4L variants.
Figure 4: The normalized probability curves of unfolding and refolding work of 16,61 WT*T4L and CP13*T4L.

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Acknowledgements

We would like to thank G. Crooks for assistance in use of Crooks fluctuation analysis, R. Dahlquist and B. Matthews for help in initiating this study, and the entire Bustamante and Marqusee labs for advice and technical help. We would particularly like to thank E. Kwon for her assistance in reagent preparation and data collection. This work was supported in part by NIH grants GM 32543 (C.B.), GM 50945 (S.M.) and a grant from the NSF (S.M.).

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Author notes

  1. Elizabeth A. Shank & Ciro Cecconi

    Present address: Present addresses: Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts 02115, USA (E.A.S.); Department of Physics, University of Modena and Reggio Emilia, Via Campi 213/A 4100 Modena, Italy (C.C.).,

  2. Elizabeth A. Shank, Ciro Cecconi and Jesse W. Dill: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular & Cell Biology, University of California, Berkeley, California 94720, USA,

    Elizabeth A. Shank, Ciro Cecconi, Susan Marqusee & Carlos Bustamante

  2. Jason L. Choy Laboratory of Single Molecule Biophysics, Institute for Quantitative Biosciences, University of California, Berkeley, California 94720, USA ,

    Elizabeth A. Shank, Ciro Cecconi, Jesse W. Dill, Susan Marqusee & Carlos Bustamante

  3. Biophysics Graduate Group, University of California, Berkeley, California 94720, USA ,

    Jesse W. Dill

  4. Howard Hughes Medical Institute, University of California, Berkeley, California 94720, USA ,

    Carlos Bustamante

  5. Department of Physics, University of California, Berkeley, California 94720, USA,

    Carlos Bustamante

  6. Department of Chemistry, University of California, Berkeley, California 94720, USA,

    Carlos Bustamante

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  1. Elizabeth A. Shank
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Contributions

E.A.S., C.C., J.W.D. conducted the experiments and analysed the data. J.W.D. carried out the Crooks fluctuation analysis. E.A.S., J.W.D., S.M. and C.B. wrote the manuscript.

Corresponding authors

Correspondence to Susan Marqusee or Carlos Bustamante.

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The authors declare no competing financial interests.

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This file contains Supplementary Figures 1-6 with legends, Supplementary Tables 1-2 and References. (PDF 836 kb)

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Shank, E., Cecconi, C., Dill, J. et al. The folding cooperativity of a protein is controlled by its chain topology. Nature 465, 637–640 (2010). https://doi.org/10.1038/nature09021

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