Cavieres J D
Biochim Biophys Acta. 1987 May 12;899(1):83-92. doi: 10.1016/0005-2736(87)90242-2.
Calmodulin-depleted red cell membranes catalyse a Ca2+, Mg2+-dependent ATP-[3H]ADP exchange at 37 degrees C. The Ca2+, Mg2+-dependent exchange, measured at 20 microM CaCl2, 1.5 mM MgCl2, 1.5 mM ADP and 1.5 mM ATP, is comparable to the (Ca2+ + Mg2+)-ATPase activity, between 0.3 and 0.8 mmol/litre original cells per h. EDTA-washed membranes present a Ca2+-dependent ATP-ADP exchange whose rate is not more than 7% of that found in a Mg2+-containing medium, while their Ca2+-dependent ATPase is essentially zero. Addition of 1.5 mM MgCl2 to the medium restores both activities to the levels found with membranes not treated with EDTA. Calmodulin (16 micrograms/ml) produces an eight-fold stimulation of the Ca2+-dependent ATP-ADP exchange, slightly less than it stimulates the Ca2+-dependent ATP hydrolysis. The effect of 1.5 mM MgCl2 on the exchange is greater in the presence than in the absence of calmodulin. It is proposed that the reversal of the initial phosphorylation of the Ca2+ pump, occurring at a fast rate at 37 degrees C, involves a conformational change in the phosphoenzyme. Thus, it would be an ADP-liganded phosphoenzyme of the form EP(ADP) that would experience the fast conformational transition at 37 degrees C. The great difficulty in producing an overall reversal of the Ca2+ pump should then be due to one or more reaction steps later than and including Ca2+ release and dephosphorylation.
钙调蛋白缺失的红细胞膜在37℃催化Ca2 +、Mg2 +依赖的ATP-[3H]ADP交换。在20μM CaCl2、1.5 mM MgCl2、1.5 mM ADP和1.5 mM ATP条件下测得的Ca2 +、Mg2 +依赖的交换，与（Ca2 + + Mg2 +）-ATP酶活性相当，为每小时每升原始细胞0.3至0.8 mmol。用EDTA洗涤的膜呈现Ca2 +依赖的ATP-ADP交换，其速率不超过在含Mg2 +培养基中发现的速率的7%，而它们的Ca2 +依赖的ATP酶基本为零。向培养基中添加1.5 mM MgCl2可使两种活性恢复到未用EDTA处理的膜所发现的水平。钙调蛋白（16μg/ml）使Ca2 +依赖的ATP-ADP交换产生八倍的刺激，略低于它对Ca2 +依赖的ATP水解的刺激。1.5 mM MgCl2对交换的影响在有钙调蛋白存在时比不存在时更大。有人提出，Ca2 +泵初始磷酸化的逆转在37℃快速发生，涉及磷酸酶的构象变化。因此，在37℃经历快速构象转变的将是形式为EP(ADP)的ADP结合磷酸酶。那么，使Ca2 +泵整体逆转的巨大困难应该归因于Ca2 +释放和去磷酸化之后的一个或多个反应步骤，包括这两个步骤。
