The Phosphorylation of the Membranal Protein of the Sarcoplasmic Vesicles during Active Calcium Transport

Abstract

The γ‐phosphate of ATP is transfered rapidly to the membranal protein of the isolated sarcoplasmic vesicles as soon as the calcium pump of the vesicles is activated (2 to 4 moles phosphate per 106 g vesicular protein). The transphosphorylation occurs only if calcium and magnesium are present and the SH‐groups essential for calcium transport are unimpaired. If, in the presence of magnesium, the concentration of the free calcium ions in the solution is reduced from 200 to about 20 nM, the phosphoprotein level declines to insignificant low values. The calcium dependency of the phosphoprotein formation coincides with that of the ATP‐ADP exchange reaction, the extra ATPase activity and the calcium accumulation. The ATP‐ADP exchange reaction as well as the formation of the phosphoprotein are independent of the calcium of the calcium concentration inside the vesicles. Phosphoprotein formation is optimally activated when the magnesium concentration equals the ATP concentration just as in ATP extra‐splitting, phosphate exchange and calcium accumulation. The amount of phosphoprotein is reduced by the addition of 1 mM ADP to about 60%. No further decline of the phosphoprotein level takes place if the concentration of ADP is raised above 1 mM, while increasing concentrations of ADP inhibit progressively ATP extra‐splitting and calcium transport. The phosphoprotein complex which exists in the presence of 1 mM ADP disappears when the calcium and/or magnesium in the solution are chelated. The stability of the isolated phosphoprotein has its optimum at pH 2. Neither molybdate nor mercury ions accelerate the hydrolysis of phosphoprotein, while hydroxylamine catalizes its decay. The activity of the calcium pump is not affected appreciably when the vesicles are treated with hydrazine under optimal conditions for phosphorylation. Copyright © 1969, Wiley Blackwell. All rights reserved