Regulation of actin-activated ATP hydrolysis by arterial myosin.
Myosin was isolated from the main pulmonary artery of swine and was phosphorylated or dephosphorylated by utilizing the endogenous kinase or phosphatase, respectively. The myosins, phosphorylated to various degrees, were purified free of kinase and phosphatase activities by gel filtration on Sepharose CL-4B agarose columns. The level of actin-activated ATPase activity was dependent upon the degree of myosin light chain phosphorylation. Fully phosphorylated myosin reconstituted with actin and tropomyosin (actin/tropomyosin = 61:1) had the highest ATPase activity (0.1 mumol of Pi/mg . min). The actin-activated ATPase activity showed maximal (60--65%) Ca2+ sensitivity at 2 mol of Ca2+ bound per mol of myosin. The actin-activated ATPase activity, Ca2+ binding, and Ca2+ sensitivity of arterial myosin were also dependent upon Mg2+ concentration. The ATPase activity was maximal at 2--3 mM Mg2+ and, at low (0.5 mM) Mg2+ concentration, the activity was only one-third of the maximal activity. Increasing the Mg2+ above 3 mM was not associated with a further increase in ATPase activity, but the Ca2+ binding and Ca2+ sensitivity decreased with increasing Mg2+ concentration. The maximal Ca2+ sensitivity was observed at 2--3 mM Mg2+, a concentration at which the myosin bound 2 mol of Ca2+/mol. Both the ATPase activity and the Ca2+ sensitivity were more remarkable when actin that contained tropomyosin was used to activate the ATPase activity. The data indicate that calcium regulates the actin-activated ATP hydrolysis not only by its effects on the phosphorylation system but also by direct binding to the myosin.
D000199:Actins / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D046911:Macromolecular Substances; D008274:Magnesium / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation; D011651:Pulmonary Artery; D013329:Structure-Activity Relationship; D013552:Swine
D000199:Actins; D046911:Macromolecular Substances; D000251:Adenosine Triphosphatases; D009218:Myosins; D008274:Magnesium; D002118:Calcium