No abstract

A large number of tropomyosin (Tm) isoforms function as gatekeepers of the actin filament, controlling the spatiotemporal access of actin-binding proteins to specialized actin networks. Residues ~40-80 vary significantly among Tm isoforms, but the impact of sequence variation on Tm structure and interactions with actin is poorly understood, since structural studies have focused on skeletal muscle Tma. We describe structures of N-terminal fragments of smooth muscle Tm a and b (sm-Tma and sm-Tmb). The 2.0-A ˚structure of sm-Tma81 (81-aa) resembles that of skeletal Tma, displaying a similar super-helical twist matching the contours of the actin filament. The 1.8-A structure of sm-Tma98 (98-aa) unexpectedly reveals an anti-parallel coiledcoil, with the two chains staggered by only 4-aa and displaying hydrophobic core interactions similar to those of the parallel dimer. In contrast, the 2.5-A structure of sm-Tmb98, containing Gly-Ala-Ser at the N-terminus to mimic acetylation, reveals a parallel coiled-coil. None of the structures contains coiled-coil stabilizing elements, favoring the formation of head-to-tail overlap complexes in four out of five crystallographically independent parallel dimers. These complexes show similarly arranged 4-helix bundles stabilized by hydrophobic interactions, but the extent of the overlap varies between sm-Tmb98 and sm-Tma81 from 2 to 3 helical turns. The formation of overlap complexes thus appears to be an intrinsic property of the Tm coiled-coil, with the specific nature of hydrophobic contacts determining the extent of the overlap. Overall, the results suggest that sequence variation among Tm isoforms has a limited effect on actin binding, but could determine its gatekeeper function.