Conformation, filament assembly, and activity of single-headed smooth muscle myosin.
Single-headed myosin was prepared by digestion of porcine aorta smooth muscle myosin with Staphylococcus aureus V8 protease in the presence of actin. The single-headed myosin preparation contained intact light chains, a rod fragment of a heavy chain, and a heavy chain of which only a minor fraction contained a nick in the head segment. Below 0.2 M NaCl, the single-headed myosin showed a decrease in Ca2+-ATPase activity and an increase in the elution time on gel filtration HPLC in a phosphorylation-dependent manner, indicating a phosphorylation-dependent conformational transition between the extended and folded forms. These conformations were confirmed by electron microscopic observation of rotary-shadowed samples of single-headed myosin. However, the conformational transition of single-headed myosin occurred in a narrower range with lower salt concentrations than that of double-headed myosin. The filament assembly of single-headed myosin was thus facilitated and phosphorylation-independent. The single-headed myosin also showed high actin-activated ATPase activity independent of phosphorylation. These results indicate that the two-headed structure of smooth muscle myosin is not essential for the conformational transition, but is required for the phosphorylation-dependent regulation of enzymatic activity and filament assembly.
Konishi
K
K
Division of Chemistry, Graduate School of Science, Hokkaido University, Kita-ku, Sapporo, 060-0810, Japan.
Katoh
T
T
Morita
F
F
Yazawa
M
M
D008841:Actin Cytoskeleton / Q000378:metabolism / Q000648:ultrastructure; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000737:chemistry*; D009218:Myosins / Q000737:chemistry* / Q000378:metabolism; D010766:Phosphorylation; D011487:Protein Conformation; D013552:Swine
D000251:Adenosine Triphosphatases; D009218:Myosins