A Folding Insulator Defines Cryptic Domains in Tropomyosin

Multidomain proteins are the product of evolutionary selection for diversity of function through concatenation and repurposing of existing modular units of structures. In structures of proteins with multiple domains, components are often globular units stitched together with flexible linkers. Multidomain proteins often fold as multiple distinct order–disorder transitions. However, the relationship between structure and folding is not always straightforward. Tropomyosin binds to actin in muscle and cytoskeletal filaments. The structure is that of a continuous ɑ-helix lacking domain boundaries, but unfolding shows distinct transitions suggesting at least three possible domains do exist. To explore how domains might occur in a continuous structure, we used Lifson-Roig helix-coil models with sequence domains of varying helical nucleation propensities. Of these models, ones with a central folding insulator, separating folding of N- and C-terminal domains, are most consistent with experimental folding studies. The positions of domain boundaries are identified by hydrogen–deuterium exchange mass spectrometry. The presence of structurally cryptic folding domains in tropomyosin could relate to its evolution and explain the uneven distribution of deleterious mutations that lead to various cardiomyopathies.