Purification and characterization of a third isoform of myosin I from Acanthamoeba castellanii.
A third isoform of myosin I has been isolated from Acanthamoeba and designated myosin IC. Peptide maps and immunoassays indicate that myosin IC is not a modified form of myosin IA, IB, or II. However, myosin IC has most of the distinctive properties of a myosin I. It is a globular protein of native Mr approximately 162,000, apparently composed of a single 130-kDa heavy chain and a pair of 14-kDa light chains. It is soluble in MgATP at low ionic strength, conditions favoring filament assembly by myosin II. Myosin IC has high Ca2+- and (K+,EDTA)-ATPase activities. Its low Mg2+-ATPase activity is stimulated to a maximum rate of 20 s-1 by the addition of F-actin if its heavy chain has been phosphorylated by myosin I heavy chain kinase. The dependence of the Mg2+-ATPase activity of myosin IC on F-actin concentration is triphasic; and, at fixed concentrations of F-action, this activity increases cooperatively as the concentration of myosin IC is increased. These unusual kinetics were first demonstrated for myosins IA and IB and shown to be due to the presence of two actin-binding sites on each heavy chain which enable those myosins I to cross-link actin filaments. Myosin IC is also capable of cross-linking F-actin, which, together with the kinetics of its actin-activated Mg2+-ATPase activity, suggests that it, like myosins IA and IB, possesses two independent actin-binding domains.
Lynch
T J
TJ
Laboratory of Cell Biology, National Heart, Lung, and Blood Institute, Bethesda, Maryland 20892.
Brzeska
H
H
Miyata
H
H
Korn
E D
ED
D000048:Acanthamoeba / Q000201:enzymology*; D000199:Actins / Q000378:metabolism; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000302:isolation & purification* / Q000378:metabolism; D006836:Hydro-Lyases / Q000235:genetics*; D007527:Isoenzymes / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D009218:Myosins / Q000302:isolation & purification* / Q000378:metabolism; D010446:Peptide Fragments / Q000302:isolation & purification; D010766:Phosphorylation; D011485:Protein Binding; D014357:Trypsin
D000199:Actins; D007527:Isoenzymes; D010446:Peptide Fragments; D014357:Trypsin; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins; D006836:Hydro-Lyases; C061663:agropine synthase