title: Load and Pi control flux through the branched kinetic cycle of myosin V.
abstract: Myosin V is a processive actin-based motor protein that takes multiple 36-nm steps to deliver intracellular cargo to its destination. In the laser trap, applied load slows myosin V heavy meromyosin stepping and increases the probability of backsteps. In the presence of 40 mm phosphate (P(i)), both forward and backward steps become less load-dependent. From these data, we infer that P(i) release commits myosin V to undergo a highly load-dependent transition from a state in which ADP is bound to both heads and its lead head trapped in a pre-powerstroke conformation. Increasing the residence time in this state by applying load increases the probability of backstepping or detachment. The kinetics of detachment indicate that myosin V can detach from actin at two distinct points in the cycle, one of which is turned off by the presence of P(i). We propose a branched kinetic model to explain these data. Our model includes P(i) release prior to the most load-dependent step in the cycle, implying that P(i) release and load both act as checkpoints that control the flux through two parallel pathways.
authors.affiliation: Department of Molecular Physiology & Biophysics, University of Vermont, Burlington, Vermont 05405, USA.
authors.forename: Neil M
authors.initials: NM
authors.lastname: Kad
authors.forename: Kathleen M
authors.initials: KM
authors.lastname: Trybus
authors.forename: David M
authors.initials: DM
authors.lastname: Warshaw
mesh_terms: D000199:Actins / Q000737:chemistry; D000244:Adenosine Diphosphate / Q000737:chemistry; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D008954:Models, Biological; D009010:Monte Carlo Method; D018995:Myosin Heavy Chains / Q000737:chemistry; D024701:Myosin Type V / Q000737:chemistry*; D010710:Phosphates / Q000737:chemistry*; D011485:Protein Binding; D015202:Protein Engineering; D013816:Thermodynamics
chemical_list: D000199:Actins; D010710:Phosphates; D000244:Adenosine Diphosphate; D000251:Adenosine Triphosphatases; D024701:Myosin Type V; D018995:Myosin Heavy Chains