INHIBITION OF ACTIN FILAMENT MOVEMENT BY MONOCLONAL-ANTIBODIES AGAINST THE MOTOR DOMAIN OF MYOSIN

Conformational transitions in defined regions of the motor domain of s keletal muscle myosin involved in ATP hydrolysis, actin binding and mo tility were probed with monoclonal antibodies. Competition binding ass ays demonstrate that three different monoclonal antibodies react with spatially related sites on the myosin heavy chain. One recognizes a se quential epitope between residues 65 and 80 and has no effect on actin filament movement in an in vitro motility assay despite tight binding to myosin and acto-S1. The other two monoclonal antibodies react with sequential epitopes between residues 29 and 60 and both inhibit actin filament movement. A fourth monoclonal antibody reacts with the N-ter minus of the heavy chain (residues 1-12) at a spatially distinct site on the myosin head and also inhibits actin filament movement. These fo ur monoclonal antibodies have been mapped by immunoelectron microscopy to the large, actin binding region of the myosin head, however, the a ntibody binding sites remain accessible on rigor complexes of acto-S1. Thus, this group of monoclonal antibodies identify sequential epitope s in a mobile segment of the myosin heavy chain. In addition, two conf ormation-sensitive monoclonal antibodies are described that are affect ed by ATP and actin binding to myosin S1, and display distinct and mar ked inhibitory effects on actin filament movement. In contrast, an ant i-light chain monoclonal antibody that binds near the myosin head-rod junction has little effect on the number and velocity of moving actin filaments. These results identify mobile regions on the myosin head th at are perturbed by antibody binding and that may be linked to force p roduction and motion.