title: Switch 1 mutation S217A converts myosin V into a low duty ratio motor.
abstract: We have determined the kinetic mechanism and motile properties of the switch 1 mutant S217A of myosin Va. Phosphate dissociation from myosin V-ADP-Pi (inorganic phosphate) and actomyosin V-ADP-Pi and the rate of the hydrolysis step (myosin V-ATP-->myosin V-ADP-Pi) were all approximately 10-fold slower in the S217A mutant than in wild type (WT) myosin V, resulting in a slower steady-state rate of basal and filamentous actin (actin)-activated ATP hydrolysis. Substrate binding and ADP dissociation kinetics were all similar to or slightly faster in S217A than in WT myosin V and mechanochemical gating of the rates of dissociation of ADP between trail and lead heads is maintained. The reduction in the rate constants of the hydrolysis and phosphate dissociation steps reduces the duty ratio from approximately 0.85 in WT myosin V to approximately 0.25 in S217A and produces a motor in which the average run length on actin at physiological concentrations of ATP is reduced 10-fold. Thus we demonstrate that, by mutational perturbation of the switch 1 structure, myosin V can be converted into a low duty ratio motor that is processive only at low substrate concentrations.
authors.affiliation: Department of Physiological Sciences, Eastern Virginia Medical School, Norfolk, Virginia 23507, USA.
authors.forename: Eva
authors.initials: E
authors.lastname: Forgacs
authors.forename: Takeshi
authors.initials: T
authors.lastname: Sakamoto
authors.forename: Suzanne
authors.initials: S
authors.lastname: Cartwright
authors.forename: Betty
authors.initials: B
authors.lastname: Belknap
authors.forename: Mihály
authors.initials: M
authors.lastname: Kovács
authors.forename: Judit
authors.initials: J
authors.lastname: Tóth
authors.forename: Martin R
authors.initials: MR
authors.lastname: Webb
authors.forename: James R
authors.initials: JR
authors.lastname: Sellers
authors.forename: Howard D
authors.initials: HD
authors.lastname: White
mesh_terms: D000199:Actins / Q000378:metabolism; D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D006868:Hydrolysis; D007700:Kinetics; D051379:Mice; D009154:Mutation; D015879:Myosin Subfragments / Q000378:metabolism; D024701:Myosin Type V / Q000235:genetics / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism; D011485:Protein Binding; D011817:Rabbits; D012694:Serine / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera
chemical_list: D000199:Actins; D015879:Myosin Subfragments; D010710:Phosphates; D012694:Serine; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D024701:Myosin Type V