Myosin lever arm orientation in muscle determined with high angular resolution using bifunctional spin labels.

ABSTRACT: Despite advances in x-ray crystallography, cryo-electron microscopy (cryo-EM), and fluorescence polarization, none of these techniques provide high-resolution structural information about the myosin light chain domain (LCD; lever arm) under ambient conditions in vertebrate muscle. Here, we measure the orientation of LCD elements in demembranated muscle fibers by electron paramagnetic resonance (EPR) using a bifunctional spin label (BSL) with an angular resolution of 4°. To achieve stereoselective site-directed labeling with BSL, we engineered a pair of cysteines in the myosin regulatory light chain (RLC), either on helix E or helix B, which are roughly parallel or perpendicular to the myosin lever arm, respectively. By exchanging BSL-labeled RLC onto oriented muscle fibers, we obtain EPR s

SUBMITTER: Savich Y 

PROVIDER: S-EPMC6683674 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature