Myosin step size. Estimation from slow sliding movement of actin over low densities of heavy meromyosin.
We have estimated the step size of the myosin cross-bridge (d, displacement of an actin filament per one ATP hydrolysis) in an in vitro motility assay system by measuring the velocity of slowly moving actin filaments over low densities of heavy meromyosin on a nitrocellulose surface. In previous studies, only filaments greater than a minimum length were observed to undergo continuous sliding movement. These filaments moved at the maximum speed (Vo), while shorter filaments dissociated from the surface. We have now modified the assay system by including 0.8% methylcellulose in the ATP solution. Under these conditions, filaments shorter than the previous minimum length move, but significantly slower than Vo, as they are propelled by a limited number of myosin heads. These data are consistent with a model that predicts that the sliding velocity (v) of slowly moving filaments is determined by the product of vo and the fraction of time when at least one myosin head is propelling the filament, that is, v = vo [1-(1-ts/tc)N], where ts is the time the head is strongly bound to actin, tc is the cycle time of ATP hydrolysis, and N is the average number of myosin heads that can interact with the filament. Using this equation, the optimum value of ts/tc to fit the measured relationship between v and N was calculated to be 0.050. Assuming d = vots, the step size was then calculated to be between 10nm and 28 nm per ATP hydrolyzed, the latter value representing the upper limit. This range is within that of geometric constraint for conformational change imposed by the size of the myosin head, and therefore is not inconsistent with the swinging cross-bridge model tightly coupled with ATP hydrolysis.
Uyeda
T Q
TQ
Department of Cell Biology, Stanford University School of Medicine, CA 94305.
Kron
S J
SJ
Spudich
J A
JA
D000199:Actins / Q000378:metabolism / Q000502:physiology* / Q000648:ultrastructure; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism; D007700:Kinetics; D008747:Methylcellulose; D009068:Movement; D015879:Myosin Subfragments / Q000378:metabolism / Q000502:physiology* / Q000648:ultrastructure; D009218:Myosins / Q000378:metabolism / Q000502:physiology* / Q000648:ultrastructure; D011817:Rabbits; D012996:Solutions
D000199:Actins; D015879:Myosin Subfragments; D012996:Solutions; D000255:Adenosine Triphosphate; D008747:Methylcellulose; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins