Purification of myosin light chain kinase from bovine cardiac muscle.

Abstract

Myosin light chain kinase was purified > 100,000-fold to apparent homogeneity, with a yield of 10% from bovine cardiac muscle. Sodium dodecyl sulfate gels of the purified kinase showed 1 stained band corresponding to a MW of 94,000. The enzyme was activated > 10-fold in the presence of Ca2+ (apparent Ka = 0.6-1.2 .mu.M) and calmodulin (apparent Ka = 3-5 nM). The purified enzyme had a specific activity of 20-30 .mu.mol of phosphate transferred per min per mg from ATP to cardiac myosin light chain 2. One mole of phosphate was incorporated per 94,000 g of the kinase in the presence of Ca2+ and calmodulin, or of cyclic AMP-dependent protein kinase or of both additions. In addition to myosin light chain kinase, a calmodulin-binding protein of unknown function was purified from bovine cardiac muscle. This protein had a MW of 85,000, was composed of 2 dissimilar subunits (MW of 61,000 and 15,000) and competed with myosin light chain kinase for calmodulin. The protein appears to be closely related to the calmodulin-binding protein I purified from brain.