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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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.).
The authors declare no competing financial interests.
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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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