Evidence for an interaction between the SH3 domain and the N-terminal extension of the essential light chain in class II myosins

J Mol Biol. 2007 Aug 24;371(4):902-13. doi: 10.1016/j.jmb.2007.05.080. Epub 2007 Jun 2.

Abstract

The function of the src-homology 3 (SH3) domain in class II myosins, a distinct beta-barrel structure, remains unknown. Here, we provide evidence, using electron cryomicroscopy, in conjunction with light-scattering, fluorescence and kinetic analyses, that the SH3 domain facilitates the binding of the N-terminal extension of the essential light chain isoform (ELC-1) to actin. The 41 residue extension contains four conserved lysine residues followed by a repeating sequence of seven Pro/Ala residues. It is widely believed that the highly charged region interacts with actin, while the Pro/Ala-rich sequence forms a rigid tether that bridges the approximately 9 nm distance between the myosin lever arm and the thin filament. In order to localize the N terminus of ELC in the actomyosin complex, an engineered Cys was reacted with undecagold-maleimide, and the labeled ELC was exchanged into myosin subfragment-1 (S1). Electron cryomicroscopy of S1-bound actin filaments, together with computer-based docking of the skeletal S1 crystal structure into 3D reconstructions, showed a well-defined peak for the gold cluster near the SH3 domain. Given that SH3 domains are known to bind proline-rich ligands, we suggest that the N-terminal extension of ELC interacts with actin and modulates myosin kinetics by binding to the SH3 domain during the ATPase cycle.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Chickens
  • Cryoelectron Microscopy
  • Fluorescence Resonance Energy Transfer
  • Models, Molecular
  • Molecular Sequence Data
  • Myosins / chemistry*
  • Myosins / classification
  • Myosins / genetics
  • Myosins / metabolism*
  • Protein Isoforms / chemistry
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • Sequence Alignment
  • src Homology Domains

Substances

  • Protein Isoforms
  • Myosins