Force-velocity relation of sliding of skeletal muscle myosin, arranged on a paramyosin filament, on actin cables.
To investigate in vitro ATP-dependent sliding of regularly arranged myosin molecules on actin filaments, we prepared thick hybrid filaments in which myosin molecules isolated from rabbit skeletal muscle were arranged around the paramyosin core (length, 10-20 micron; diameter, </=0.2 micron) obtained from a molluscan smooth muscle. A single to a few thick hybrid filaments were attached to a polystyrene bead (diameter, 4.5 micron; specific gravity, 1.5) and made to slide on actin filament arrays (actin cables) in the internodal cell of an alga, mounted on the rotor of a centrifuge microscope. The bead was subjected to centrifugal forces serving as external loads to the ATP-dependent actin-myosin sliding. The maximum unloaded sliding velocity of the thick filament attached-bead (mean, 3.4 micron/s; 20-23 degrees C) was significantly higher than that of the bead coated with randomly oriented myosin molecules reported previously. The steady-state force-velocity (P-V) relations obtained were qualitatively similar to those in intact skeletal muscle fibers. These results indicate that this in vitro motility assay system retains the basic characteristics of contracting skeletal muscle fibers, and that it may be effectively used to study mechanisms underlying the steady-state P-V characteristics of ATP-dependent actin-myosin sliding using various recombinant myosins produced in nonmuscle cells.
Tameyasu
T
T
Department of Physiology, School of Medicine, St. Marianna University, Kawasaki, 216-0015, Japan.
Akimoto
T
T
Hirohata
Y
Y
Shirakawa
I
I
Yamamoto
N
N
Kosuge
S
S
Sugi
H
H
D008841:Actin Cytoskeleton / Q000502:physiology*; D000199:Actins / Q000502:physiology*; D000818:Animals; D049872:Bivalvia; D006706:Homeostasis / Q000502:physiology; D018482:Muscle, Skeletal / Q000378:metabolism*; D009218:Myosins / Q000502:physiology*; D011817:Rabbits; D013997:Time Factors; D014335:Tropomyosin / Q000502:physiology*
D000199:Actins; D014335:Tropomyosin; D009218:Myosins