Inhibition of conformational change in smooth muscle myosin by a monoclonal antibody against the 17-kDa light chain.
Monoclonal antibodies against gizzard smooth muscle myosin were generated and characterized. One of these antibodies, designated MM-2, recognized the 17-kDa light chain and modulated the ATPase activities and hydrodynamic properties of smooth muscle myosin. Rotary shadowing electron microscopy showed that MM-2 binds 51 (+/- 25) A from the head-rod junction. The depression of Ca2+- and Mg2+-ATPase activities of myosin and Ca2+-ATPase activity of heavy meromyosin at low KCl concentration were abolished by MM-2. Viscosity measurement indicated that MM-2 inhibits the transition of 6 S myosin to 10 S myosin. While the rate of the production of subfragment-1 by papain proteolysis of 6 S myosin was inhibited by MM-2, the rate of proteolysis of the heavy chain of 10 S myosin was enhanced by MM-2 and reached the same rate as that of 6 S myosin plus MM-2. These results suggest that MM-2 inhibits the formation of 10 S myosin by binding to the 17-kDa light chain which is localized at the head-neck region of the myosin molecule. MM-2 increased the Vmax of actin-activated Mg2+-ATPase activities of both dephosphorylated myosin and dephosphorylated heavy meromyosin about 10- and 20-fold, respectively. MM-2 also activated the actin-activated Mg2+-ATPase activity of phosphorylated myosin at a low MgCl2 concentration and thus abolished the Mg2+-dependence of acto phosphorylated myosin ATPase activity. These results suggest that MM-2 inhibits the formation of 10 S myosin, and this results in the activation of actin-activated Mg2+-ATPase activity even in the absence of phosphorylation.
Highashihara
M
M
Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106.
Frado
L L
LL
Craig
R
R
Ikebe
M
M
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D000911:Antibodies, Monoclonal / Q000502:physiology*; D001667:Binding, Competitive; D002417:Cattle; D000939:Epitopes / Q000032:analysis; D006868:Hydrolysis; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D009130:Muscle, Smooth / Q000201:enzymology / Q000276:immunology* / Q000648:ultrastructure; D009218:Myosins / Q000276:immunology* / Q000648:ultrastructure; D010206:Papain; D011487:Protein Conformation; D011817:Rabbits; D014422:Turkeys
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010206:Papain; D000251:Adenosine Triphosphatases; D009218:Myosins