Desensitization of substrate inhibition of acto-H-meromyosin ATPase by treatment of H-meromyosin with rho-chloromercuribenzoate. Relation between the extent of desensitization and the amount of bound rho-chloromercuribenzoate1.

H-Meromyosin (CMB leads to betaME-H-meromyosin) was prepared by tryptic digestion of myosin, which had been treated with CMB bound to H-meromyosin and the extent of desensitization of the substrate inhibition of acto-H-meromyosin ATPase [EC 3.6.1.3.] was investigated. Both the dissociation of acto-H-meromyosin induced by ATP and substrate inhibition decreased with increase in the amount of bound CMB to a minimum value at about 1 mole of CMB bound per mole of H-meromyosin. The substrate inhibition of acto-H-meromyosin ATPase was restored to the original level by complete removal of the bound CMB by further treatment of CMB leads to beta ME-H-meromyosin with a large excess of beta-mercaptoethanol. The dissociation constant of acto-H-meromyosin in the presence of ATP decreased markedly on modification with CMB, while the maximum ATPase activity ar a sufficiently high concentration of F-actin remained essentially unchanged. Acto-H-meromyosin was reconstituted from F-actin and CMB LEADS TO beta ME-H-meromyosin, containing less than the stoichiometric amount of bound CMB. Its ATPase activity and the extent of dissociation of acto-H-meromyosin induced by ATP were explained as those of a mixture of unmodified H-meromyosin and CMB leads to beta ME-H-meromyosin containing 1 mole of CMB per mole of H-meromyosin. Half of the light chains (g2), with a molecular weight of 18,000, were removed from myosin by treatment with CMB and beta-mercaptoethanol. After this treatment, on further incubation of the myosin with a large excess of beta-mercaptoethanol, the myosin contained only half of the g2, but the substrate inhibition of acto-H-meromyosin ATPase was restored completely. The initial burst of P1 liberation and the EDTA-ATPase activity decreased to almost zero on specific modification of the SH1-groups with NEM, while the initial burst decreased to some extent and the EDTA-ATPase activity to 50% of the original value on binding of 1 mole CMB per mole of H-meromyosin. The actomyosin-type of ATPase activity was strongly inhibited by modification with CMB. The extent of the dissociation of acto-H-meromyosin induced by ATP was unaffected by modification with NEM, while it decreased on further treatment of NEM-myosin with CMB FOLLOWED BY BETA-MERCAPTOETHANOL.