A simple and rapid preparation of fully phosphorylated and fully dephosphorylated skeletal muscle myosin. Application to the preparation of a phosphorylated LC2-modified artificial isozyme

Fast skeletal myosin LC2 is phosphorylated on ser-15 by a specific myosin light chain kinase (MLCK) in the presence of Ca2+ and calmodulin, and dephosphorylated by a muscle phosphatase in the presence of Mg2+. Fully dephosphorylated myosin is obtained by dialysis of muscle crude extract (0.06m NaCl, 0.01m Tris-HCl, pH 7.5, 50 μm EGTA); fully phosphorylated myosin is obtained by addition of Ca2+ (0.2mm), Mg2+ (10mm) and ATP (3mm) and 5 min incubation at 28° C.