title: Kinetic characterization of the ATPase and actin-activated ATPase activities of Acanthamoeba castellanii myosin-2.
abstract: Phosphorylation of Ser-639 in loop-2 of the catalytic motor domain of the heavy chain of Acanthamoeba castellanii myosin-2 and the phosphomimetic mutation S639D have been shown previously to down-regulate the actin-activated ATPase activity of both the full-length myosin and single-headed subfragment-1 (Liu, X., Lee, D. Y., Cai, S., Yu, S., Shu, S., Levine, R. L., and Korn, E. D. (2013) Proc. Natl. Acad. Sci. U.S.A. 110, E23-E32). In the present study we determined the kinetic constants for each step in the myosin and actomyosin ATPase cycles of recombinant wild-type S1 and S1-S639D. The kinetic parameter predominantly affected by the S639D mutation is the actin-activated release of inorganic phosphate from the acto myosin·ADP·Pi complex, which is the rate-limiting step in the steady-state actomyosin ATPase cycle. As consequence of this change, the duty ratio of this conventional myosin decreases. We speculate on the effect of Ser-639 phosphorylation on the processive behavior of myosin-2 filaments.
authors.affiliation: From the Laboratory of Molecular Physiology and the Laboratory of Cell Biology, NHLBI, National Institutes of Health, Bethesda, Maryland 20892-8015.
authors.forename: Sarah M
authors.initials: SM
authors.lastname: Heissler
authors.forename: Xiong
authors.initials: X
authors.lastname: Liu
authors.forename: Edward D
authors.initials: ED
authors.lastname: Korn
authors.forename: James R
authors.initials: JR
authors.lastname: Sellers
mesh_terms: D048248:Acanthamoeba castellanii / Q000201:enzymology*; D000199:Actins / Q000737:chemistry*; D000205:Actomyosin / Q000737:chemistry; D000244:Adenosine Diphosphate / Q000737:chemistry; D000251:Adenosine Triphosphatases / Q000737:chemistry*; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry
chemical_list: D000199:Actins; D011994:Recombinant Proteins; D012694:Serine; D000244:Adenosine Diphosphate; D000205:Actomyosin; D000251:Adenosine Triphosphatases; D009218:Myosins
keywords: ATPases; Actomyosin; Kinetics; Molecular Motors; Myosin; Phosphorylation
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