Two New Modes of Smooth Muscle Myosin Regulation by the Interaction between the Two Regulatory Light Chains, and by the

Previous studies indicated that single-headed smooth muscle myosin and SI (a single head fragment) are not regulated through phosphorylation of the regulatory light chain (RLC). To investigate the importance of the double-headedness of myosin and of the S2 region for the phosphorylation-dependent regulation, we made three types of recombi-nant mutant smooth muscle HMMs with one intact head and an N-terminally truncated head. The truncated head of AMD lacked the motor domain, that of A(MD+ELC) lacked the motor and essential light chain binding domains, and single-headed HMM had one intact head alone. The basal ATPase activities of the three mutants decreased as the KC1 concentration became less than 0.1 M. Such a decrease was not observed for SI, which had no S2 region, suggesting that S2 is necessary for this myosin behavior. This activity decrease also disappeared when RLCs of AMD and A(MD+ELC), but that of single-headed HMM, were phosphorylated. When their RLCs were unphosphorylated, the three mutants exhibited similar actin-activated ATPase levels. However, when they were phosphorylated, the actin-activated ATPase activities of AMD and A(MD+ELC) in-creased to the SI level, while that of single-headed HMM remained unchanged. Even i