Phosphorylation kinetics of skeletal muscle myosin and the effect of phosphorylation on actomyosin adenosinetriphosphatase activity.
Purified rabbit skeletal muscle myosin is phosphorylated on one type of light-chain subunit (P-light chain) by calmodulin-dependent myosin light chain kinase and dephosphorylated by phosphoprotein phosphatase C. Analyses of the time courses of both phosphorylation and dephosphorylation of skeletal muscle myosin indicated that both reactions, involving at least 90% of the P-light chain, were kinetically homogeneous. These results suggest that phosphorylation and dephosphorylation of rabbit skeletal muscle myosin heads are simple random processes in contrast to the sequential phosphorylation mechanism proposed for myosin from gizzard smooth muscle. We also examined the effect of phosphorylation of rabbit skeletal muscle myosin on the actin-activated ATPase activity. We observed an apparent 2-fold decrease in the Km for actin, from about 6 microM to about 2.5 microM, with no significant effect on the Vmax (1.8s-1) in response to P-light-chain phosphorylation. There was no significant effect of phosphorylation on the ATPase activity of myosin alone (0.045 s-1). ATPase activation could be fully reversed by addition of phosphatase catalytic subunit. The relationship between the extents of P-light-chain phosphorylation and ATPase activation (at 3.5 microM actin and 0.6 microM myosin) was essentially linear. Thus, in contrast to results obtained with myosin from gizzard smooth muscle, these results suggest that cooperative interactions between the myosin heads do not play an important role in the activation process in skeletal muscle. Since the effect of P-light-chain phosphorylation is upon the Km for actin, it would appear to be associated with a significant activation of ATPase activity only at appropriate concentrations of actin and salt.(ABSTRACT TRUNCATED AT 250 WORDS)
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D007700:Kinetics; D009132:Muscles / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase; D009218:Myosins / Q000378:metabolism*; D009994:Osmolar Concentration; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D011817:Rabbits
D011494:Protein Kinases; D009219:Myosin-Light-Chain Kinase; D000251:Adenosine Triphosphatases; D009218:Myosins