Cold-sensitive mutants G680V and G691C of Dictyostelium myosin II confer dramatically different biochemical defects.
Cold-sensitive myosin mutants represent powerful tools for dissecting discrete deficiencies in myosin function. Biochemical characterization of two such mutants, G680V and G691C, has allowed us to identify separate facets of myosin motor function perturbed by each alteration. Compared with wild type, the G680V myosin exhibits a substantially enhanced affinity for several nucleotides, decreased ATPase activity, and overoccupancy or creation of a novel strongly actin-binding state. The properties of the novel strong binding state are consistent with a partial arrest or pausing at the onset of the mechanical stroke. The G691C mutant, on the other hand, exhibits an elevated basal ATPase indicative of premature phosphate release. By releasing phosphate without a requirement for actin binding, the G691C can bypass the part of the cycle involving the mechanical stroke. The two mutants, despite having alterations in glycine residues separated by only 11 residues, have dramatically different consequences on the mechanochemical cycle.
Patterson
B
B
Department of Molecular and Cell Biology, University of Arizona, Tucson, Tucson, Arizona 85721, USA. Bruce_Patterson@tikal.biosci.arizona.edu
Ruppel
K M
KM
Wu
Y
Y
Spudich
J A
JA
D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D003080:Cold Temperature; D004023:Dictyostelium / Q000235:genetics*; D005998:Glycine / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009218:Myosins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding
D000199:Actins; D009218:Myosins; D000252:Calcium-Transporting ATPases; D005998:Glycine