The Structural Dynamics of Alpha-Tropomyosin on F-Actin Shape the Overlap Complex between Adjacent Tropomyosin Molecules

Abstract

Coiled-coil tropomyosin, localized on actin filaments in virtually all eukaryotic cells, serves as a gatekeeper regulating access of the motor protein myosin and other actin-binding proteins onto the thin filament surface. In order to form a continuous cable on thin filaments that is free of gaps, adjacent tropomyosin molecules polymerize head-to-tail by means of a short (∼9-10 residue) overlap. Thus to characterize the overlap structure, several laboratories have engineered peptides to mimic the N/C-terminal tropomyosin association. The overlapping domains formed show a compact N-terminal coiled-coil inserting into a partially opened C-terminal partner, where the opposing coiled-coils at the overlap junction face each other at up to ∼90° twist angles. Here, Molecular Dynamics simulations were carried out to determine constraints on the formation of the tropomyosin overlap complex and to assess the amount of twisting exhibited by full-length tropomyosin when bound to actin. With the exception of the last 20 to 40 C- and N-terminal residues, we find that the average tropomyosin structure closely resembles a model proposed in the classic work of McLachlan and Stewart, displaying perfectly symmetrical supercoil geometry matching theF-actin helix with an integral number of coiled-coil turns, a coiled-coil helical pitch of 137 A, a superhelical pitch of 770 A, and no localized pseudo-rotation. Over the middle 70% of tropomyosin, the average twisting of the coiled-coil deviates only by 10° from the canonical model and the torsional freedom is very small (std. dev. of 7°). This small degree of twisting cannot yield the orthogonal N- and C-termini configuration observed experimentally. However, in marked contrast, considerable coiled-coil unfolding, splaying and twisting at N- and C-terminal ends is observed, providing the conformational plasticity needed for head-to-tail nexus formation.