Full-length myosin Va exhibits altered gating during processive movement on actin.
Myosin Va (myoV) is a processive molecular motor that transports intracellular cargo along actin tracks with each head taking multiple 72-nm hand-over-hand steps. This stepping behavior was observed with a constitutively active, truncated myoV, in which the autoinhibitory interactions between the globular tail and motor domains (i.e., heads) that regulate the full-length molecule no longer exist. Without cargo at near physiologic ionic strength (100 mM KCl), full-length myoV adopts a folded (approximately 15 S), enzymatically-inhibited state that unfolds to an extended (approximately 11 S), active conformation at higher salt (250 mM). Under conditions favoring the folded, inhibited state, we show that Quantum-dot-labeled myoV exhibits two types of interaction with actin in the presence of MgATP. Most motors bind to actin and remain stationary, but surprisingly, approximately 20% are processive. The moving motors transition between a strictly gated and hand-over-hand stepping pattern typical of a constitutively active motor, and a new mode with a highly variable stepping pattern suggestive of altered gating. Each head of this partially inhibited motor takes longer-lived, short forward (35 nm) and backward (28 nm) steps, presumably due to globular tail-head interactions that modify the gating of the individual heads. This unique mechanical state may be an intermediate in the pathway between the inhibited and active states of the motor.
Department of Molecular Physiology and Biophysics, University of Vermont, Burlington, VT 05405, USA.
Jessica M
JM
Armstrong
Elena
E
Krementsova
Arthur J
AJ
Michalek
Aoife T
AT
Heaslip
Shane R
SR
Nelson
Kathleen M
KM
Trybus
David M
DM
Warshaw
D000199:Actins / Q000502:physiology*; D000818:Animals; D051379:Mice; D018995:Myosin Heavy Chains / Q000502:physiology*; D024701:Myosin Type V / Q000502:physiology*; D009994:Osmolar Concentration; D014461:Ultracentrifugation
D000199:Actins; C513082:Myo5a protein, mouse; D024701:Myosin Type V; D018995:Myosin Heavy Chains