Cooperative dependence of the actin-activated Mg2+-ATPase activity of Acanthamoeba myosin II on the extent of filament phosphorylation.
The actin-activated Mg2+-ATPase of myosin II from Acanthamoeba castellanii is regulated by phosphorylation of 3 serine residues at the tip of the tail of each of its two heavy chains; only dephosphorylated myosin II is active, whereas the phosphorylated and dephosphorylated forms have identical Ca2+-ATPase activities and Mg2+-ATPase activities in the absence of F-actin. We have now chemically modified phosphorylated and dephosphorylated myosin II with N-ethylmaleimide (NEM). The modification occurred principally at a single site within the NH2-terminal 73,000 Da of the globular head of the heavy chain. NEM-myosin II bound to F-actin and formed filaments normally, but the Ca2+- and Mg2+-ATPase activities of phosphorylated and dephosphorylated myosin II and the actin-activated Mg2+-ATPase activity of NEM-dephosphorylated myosin II were inhibited. Only filamentous myosin II has actin-activated Mg2+-ATPase activity. Native phosphorylated myosin II acquired actin-activated Mg2+-ATPase activity when it was co-polymerized with NEM-inactivated dephosphorylated myosin II, and the increase in its activity was cooperatively dependent on the fraction of NEM-dephosphorylated myosin II in the filaments. From this result, we conclude that the specific activity of each molecule within a filament is independent of its own state of phosphorylation, but is highly cooperatively dependent upon the state of phosphorylation of the filament as a whole. This enables the actin-activated Mg2+-ATPase activity of myosin II filaments to respond rapidly and extensively to small changes in the level of their phosphorylation.
Atkinson
M A
MA
Department of Biochemistry, University of Texas Health Center, Tyler 75710.
Lambooy
P K
PK
Korn
E D
ED
D000048:Acanthamoeba / Q000201:enzymology / Q000378:metabolism; D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism*; D005033:Ethylmaleimide / Q000494:pharmacology; D066298:In Vitro Techniques; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D009218:Myosins / Q000378:metabolism*; D010446:Peptide Fragments / Q000032:analysis; D010766:Phosphorylation
D000199:Actins; D010446:Peptide Fragments; D009219:Myosin-Light-Chain Kinase; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins; D005033:Ethylmaleimide