Effects of phosphorylation of light chain residues threonine 18 and serine 19 on the properties and conformation of smooth muscle myosin.
Smooth muscle myosin can be phosphorylated by myosin light chain kinase at the serine 19 and threonine 18 residues of the two 20,000-dalton light chains (Ikebe, M., Hartshorne, D. J., and Elizinga, M. (1986) J. Biol. Chem. 261, 36-39). These studies with myosin and heavy meromyosin (HMM) compare the effects induced by phosphorylation of serine 19 (M2P and HMM2P) and serine 19 plus threonine 18 (M4P and HMM4P). Formation of M4P altered the KCl dependence of viscosity and Mg2+-ATPase and higher values were maintained at lower ionic strengths, compared to M2P or dephosphorylated myosin (Mo). This is consistent with the stabilization of the 6 S conformation. The tendency for aggregation, as judged by light scattering, followed the sequence M4P greater than M2P greater than Mo. Filaments formed with M4P were more resistant to dissociation by ATP compared to filaments of M2P. Phosphorylation of HMM2P doubled Vmax of actin-activated ATPase with little effect on the apparent affinity for actin. The Mg2+-ATPase of HMM4P exhibited a higher activity at low ionic strength compared to HMM2P and HMMo. Hydrodynamic differences were detected at low ionic strength in the presence of ATP by sedimentation velocity measurements with HMM4P, HMM2P, and HMMo. Proteolysis by papain indicated an increased susceptibility of the head-neck junction of HMM4P compared to HMM2P. These data suggest that the phosphorylation of threonine 18 in addition to serine 19 change the conformation of myosin and HMM and this is associated with altered biological properties.
Ikebe
M
M
Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106.
Koretz
J
J
Hartshorne
D J
DJ
D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism; D002417:Cattle; D007700:Kinetics; D008854:Microscopy, Electron; D009130:Muscle, Smooth / Q000201:enzymology*; D015879:Myosin Subfragments / Q000378:metabolism; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation; D011189:Potassium Chloride / Q000494:pharmacology; D011487:Protein Conformation; D011817:Rabbits; D012694:Serine / Q000378:metabolism*; D013912:Threonine / Q000378:metabolism*; D014422:Turkeys; D014783:Viscosity
D015879:Myosin Subfragments; D013912:Threonine; D012694:Serine; D011189:Potassium Chloride; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins