Studies on the effect of phosphorylation of the 20,000 Mr light chain of vertebrate smooth muscle myosin.
Myosin was rapidly prepared from turkey gizzard muscle to a high level of purity, in high yield and in a non-phosphorylated state. It was consistently observed that the actin-activated Mg2+ ATPase activity of this myosin was dependent on the level of phosphorylation of the 20,000 Mr light chain, for example, in the non-phosphorylated state, the myosin Mg2+ ATPase activity was not activated by actin whereas, when the light chains were phosphorylated, the Mg2+ ATPase activity of the myosin was activated approximately ninefold by actin. Using the "desensitized" scallop myosin test system (Kendrick-Jones et al., 1976; Sellers et al., 1980) it was further demonstrated that phosphorylation of the 20,000 Mr gizzard light chain has a regulatory role. These results also suggest that the regulatory mechanisms mediated by smooth muscle myosin light chains and molluscan myosin regulatory light chains are similar, i.e. in the absence of Ca2+, both types of light chain inhibit myosin interaction with actin and this inhibition is relieved by either phosphorylation in smooth muscle or by direct calcium binding in molluscan myosins. The basis of regulation exerted by these light chains is therefore repression derepression. Using a variety of techniques, i.e. turbidity measurements, quantitative high speed centrifugation, electron microscopy and dark field light microscopy, it was observed that the stability of gizzard myosin filaments at approximately physiological conditions (0.15 M-NaCl, 1 mM-MgATP, pH 7.0) was dependent on the level of light chain phosphorylation. Using purified calmodulin-dependent light chain kinase and phosphatase, it was further shown that these gizzard myosin filaments can be reversibly assembled and disassembled as a result of phosphorylation-dephosphorylation of the 20,000 Mr light chain.
D000199:Actins / Q000494:pharmacology; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D004789:Enzyme Activation / Q000187:drug effects; D005895:Gizzard, Avian / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D046911:Macromolecular Substances; D008274:Magnesium / Q000494:pharmacology; D008854:Microscopy, Electron; D009130:Muscle, Smooth / Q000032:analysis; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation; D011487:Protein Conformation; D014422:Turkeys
D000199:Actins; D046911:Macromolecular Substances; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins; D008274:Magnesium; D002118:Calcium