ROLE OF SEQUENCE-18-29 ON ACTIN IN ACTOMYOSIN INTERACTIONS

Affinity-purified polyclonal antibodies prepared against a synthetic p eptide corresponding to sequence 18-29 from the N-terminus of rabbit a lpha-skeletal actin reacted with G- and F-actin. Epitope mapping exper iments with thrombin and hydroxylamine cleaved actin, and immunochemic al assays verified the specificity of antibodies for the 18-29 sequenc e on actin. The binding of up to 0.5 mol of IgG per mole of actin did not affect the rigor binding of myosin subfragment 1 (S-1) to actin. S imilarly, the binding of IgG to actin was not changed by a complete sa turation of actin by S-1. In contrast to this, the weak acto-S-1 inter actions in the presence of ATP were strongly inhibited by the 18-29 an tibodies. At 25-degrees-C, the acto-S-1 ATPase activity was inhibited by IgG stronger than the binding of S-1-ATPgammaS to actin. Thus, at t his temperature, a catalytic inhibition of the acto-S-1 system appears to account at least in part for the antibody effect. Acto-S-1 ATPase activities at 25-degrees-C were inhibited also by Fab(18-29). At 5-deg rees-C, the acto-S-1 ATPase activity and the binding of S-1.ATP to act in were inhibited approximately to the same extent by IgG(18-29). Thes e results are discussed in terms of S-1 binding sites on actin and the possible role of sequence 18-29 in actomyosin interactions.