Light Chain Kinase Specificity in Cardiac Myosin

Human ventricular cardiac myosin regulatory light chain (MYL2) phosphorylation modifies Ser15. This modification affects MYL2 secondary structure and modulates the Ca2+ sensitivity of contraction in cardiac tissue. Smooth muscle myosin light chain kinase (smMLCK) is prevalent in uterus and present in other contracting tissues including cardiac muscle. The recombinant 130 kDa (short) smMLCK phosphorylated Ser15 in MYL2 in vitro. Specific modification of Ser15 was verified by direct detection of the phospho group on Ser15 with mass spectrometry. SmMLCK also specifically phosphorylated myosin regulatory light chain Ser15 in porcine ventricular myosin and chicken gizzard smooth muscle myosin (Ser20 in smooth muscle) but failed to phosphorylate the myosin regulatory light chain in rabbit skeletal myosin. Michaelis-Menten Vm and KM constants for Ser15 phosphorylation in MYL2, porcine ventricular myosin, and chicken gizzard myosin are similar. These data demonstrate that smMLCK is a specific and efficient kinase for the in vitro phosphorylation of MYL2, cardiac, and smooth muscle myosin. Whether smMLCK plays a role in cardiac muscle regulation or response to a disease causing stimulus is unclear but it should be considered a potentially significant kinase in cardiac tissue on the basis of its specificity, kinetics, and tissue expression. Supported by NIH NIAMS and NHLBI grants R01AR049277 and R01HL095572.