RAPID TURNOVER OF MYOSIN LIGHT-CHAIN PHOSPHATE DURING CROSS-BRIDGE CYCLING IN SMOOTH-MUSCLE

The rate of phosphatase-mediated dephosphorylation of the regulatory l ight chain of smooth muscle myosin was determined under nearly steady- state conditions in permeabilized muscles, from the time course of inc orporation of P-33-labeled phosphate into the light chain after the ph otolytic release of [gamma-P-33]ATP from high specific activity caged [gamma-P-33]ATP. The extent of myosin light chain phosphorylation is u nchanged, and, if the kinase and phosphatase reactions are irreversibl e, the rate constant for the exponential increase in P-33 in the light chain is equal to the rate constant for the phosphatase reaction. Und er activated conditions (pCa 4.5) at 20 degrees C, the incorporation o f P-33 into similar to 80% of the phosphorylated light chain is fit by a single exponential with a rate constant of 0.37 s(-1). ATP usage du e to phosphorylation and dephosphorylation of the light chain is about one-third of the suprabasal energy requirement. The high phosphatase rate constant suggests that dephosphorylation of the light chain is ra pid enough to interact with and potentially modify the completion of t he cross-bridge cycle.