Stimulation of cardiac sarcoplasmic reticulum calcium pump by acylphosphatase: relationship to phospholamban phosphorylation.

Ca21 transport by cardiac sarcoplasmic reticulum is tightly coupled with the enzymatic activity of Ca-dependent ATPase, which forms and decomposes an intermediate phosphoenzyme. Heart sarcoplasmic reticulum Ca pump is regulated by cAMP-dependent protein kinase (PKA) phospholamban phosphorylation, which results in a stimulation of the initial rates of Ca21 transport and Ca21 ATPase activity. In the present studies we found that acylphosphatase from heart muscle, used at concentrations within the physiological range, actively hydrolyzes the phosphoenzyme of cardiac sarcoplasmic reticulum Ca pump, suggesting an high affinity of the enzyme for this special substrate. In unphosphorylated vesicles acylphosphatase enhanced the rate of ATP hydrolysis and Ca uptake with a concomitant significant decrease in apparent Km for Ca and ATP. In vesicles whose phospholamban was PKA-phosphorylated, acylphosphatase also stimulated the rate of Ca uptake and ATP hydrolysis but to a lesser extent, and the Km values for Ca and ATP were not significantly different with respect to those found in the absence of acylphosphatase. These findings suggest that acylphosphatase, owing to its hydrolytic effect, accelerates the turnover of the phosphoenzyme intermediate with the consequence of an enhanced activity of Ca pump. It is known that phosphorylation of phospholamban results in an increase of the rate at which the phosphoenzyme is decomposed. Thus, as discussed, a competition between phospholamban and acylphosphatase effect on the phosphoenzyme might be proposed to explain why the stimulation induced by this enzyme is less marked in PKAphosphorylated than in unphosphorylated heart vesicles.