COUPLING OF ATPASE ACTIVITY AND MOTILITY IN SMOOTH-MUSCLE MYOSIN IS MEDIATED BY THE REGULATORY LIGHT-CHAIN

Smooth muscle myosin acts as a molecular motor only if the regulatory light chain (RLC) is phosphorylated. This subunit can be removed from myosin by a novel method involving the use of trifluoperazine. The mot ility of RLC-deficient myosin is very slow, but native properties are restored when RLC is rebound. Truncating 6 residues from the COOH term inus of the RLC had no effect on phosphorylated myosin's motor propert ies, while removal of the last 12 residues reduced velocity by approxi mate to 30%. Very slow movement was observed once 26 residues were del eted, or with myosin containing only the COOH-terminal RLC domain. The se two mutants thus mimicked the behavior of RLC-deficient myosin, wit h the important difference that the mutant myosins were monodisperse w hen assayed by sedimentation velocity and electron microscopy. The dec reased motility therefore cannot be caused by aggregation. A common fe ature of RLC-deficient myosin and the mutant myosins that moved actin slowly was an increased myosin ATPase compared with dephosphorylated m yosin, and a lower actin-activated ATPase than obtained with phosphory lated myosin. These results suggest that the COOH-terminal portion of an intact RLC is involved in interactions that regulate myosin's ''on- off' switch, both in terms of completely inhibiting and completely act ivating the molecule.