Chiesi M, Zurini M, Carafoli E
Biochemistry. 1984 Jun 5;23(12):2595-600. doi: 10.1021/bi00307a009.
The Ca2+-transporting ATPase of erythrocytes was isolated by calmodulin affinity chromatography. The backward reaction of the ATPase was investigated. The phosphorylation of the solubilized enzyme by Pi required Mg and was inhibited by Ca and vanadate in the micromolar concentration range. Significant amounts of phosphoenzyme could be obtained only in a medium containing high dimethyl sulfoxide concentrations (greater than 25%) in order to diminish water activity at the phosphorylation site. The phosphoenzyme formed in this way could not phosphorylate ADP. However, upon addition of Ca2+ ions and dilution of dimethyl sulfoxide in the phosphorylated preparation (water activity jump), a highly reactive phosphoenzyme species was obtained which could transfer phosphate in nearly stoichiometric amounts to ADP to form ATP.
通过钙调蛋白亲和层析法分离出红细胞的钙离子转运ATP酶。对该ATP酶的逆向反应进行了研究。Pi对溶解酶的磷酸化作用需要镁离子，并且在微摩尔浓度范围内受到钙离子和钒酸盐的抑制。只有在含有高浓度二甲基亚砜（大于25%）的介质中才能获得大量的磷酸酶，以便降低磷酸化位点的水活性。以这种方式形成的磷酸酶不能使ADP磷酸化。然而，在磷酸化制剂中加入钙离子并稀释二甲基亚砜（水活性跃升）后，得到了一种高反应活性的磷酸酶物种，它可以将几乎化学计量的磷酸盐转移到ADP上以形成ATP。
