Mas-Oliva J
Departamento de Bioenergética, Universidad Nacional Autónoma de México.
Cardioscience. 1992 Jun;3(2):107-15.
A Ca(2+)-dependent ATPase, purified from cardiac microsomal membranes by solubilization and chromatography, is identified as cardiac sarcoplasmic reticulum ATPase on the basis of its electrophoretic mobility and its trypsin digestion pattern. The ATPase (both in membranous and purified form) is stimulated by calmodulin, while the skeletal muscle ATPase is not. Rapid kinetic experiments demonstrate that the calmodulin stimulation is already present within the first enzyme cycle following the addition of ATP, and consists of an increased turnover of the phosphorylated enzyme intermediate. The calmodulin effect does not involve the phosphorylation of any protein other than the ATPase. Following the incubation of ATPase with [gamma-32P]ATP, even in conditions of calmodulin stimulation, radioactive phosphorus is found only on the ATPase electrophoretic band, corresponding to the phosphorylated enzyme intermediate. These observations, together with the results obtained for [125I]calmodulin binding to the ATPase, suggest that the stimulation in turnover produced by calmodulin on the ATPase is due to a direct effect on the enzyme. This may provide an independent regulation of the cardiac sarcoplasmic reticulum Ca(2+)-ATPase, in addition to the known regulation mediated by other accessory proteins.
一种通过溶解和色谱法从心脏微粒体膜中纯化得到的钙依赖性ATP酶，根据其电泳迁移率和胰蛋白酶消化模式被鉴定为心脏肌浆网ATP酶。该ATP酶（无论是膜结合形式还是纯化形式）都受到钙调蛋白的刺激，而骨骼肌ATP酶则不受刺激。快速动力学实验表明，在添加ATP后的第一个酶循环中就已经存在钙调蛋白刺激，其表现为磷酸化酶中间体的周转增加。钙调蛋白的作用并不涉及除ATP酶以外的任何蛋白质的磷酸化。即使在钙调蛋白刺激的条件下，将ATP酶与[γ-32P]ATP一起孵育后，放射性磷也仅在与磷酸化酶中间体相对应的ATP酶电泳条带上被发现。这些观察结果，连同[125I]钙调蛋白与ATP酶结合的结果表明，钙调蛋白对ATP酶周转的刺激是由于对该酶的直接作用。除了由其他辅助蛋白介导的已知调节外，这可能为心脏肌浆网钙ATP酶提供一种独立的调节机制。
