Dalla Serra M, Pederzolli C, Antolini R, Cusinato F, Luciani S, Menestrina G
Dipartimento di Fisica, Università di Trento, Povo, Italy.
Cardioscience. 1991 Sep;2(3):193-7.
Sarcolemmal membrane vesicles isolated from bovine ventricular tissue accumulate Ca2+ through the Na+/Ca2+ exchanger when exposed to an outwardly directed Na+ gradient. This Ca2+ is then released by the same mechanism if the vesicles are transferred to a Ca(2+)-depleted Na+ buffer. Using the Ca+ indicator, arsenazo III, and a stopped-flow spectrophotometer, we can directly follow the kinetics of Ca2+ extrusion. We can thus measure the activity of the Na+/Ca2+ exchanger by the initial rate of Ca2+ release. We found that it depends upon the external Na+ concentration in a cooperative way, with a Hill coefficient of 2. By studying the temperature dependence of Na+/Ca2+ exchange, we found that it can be described by a single activation energy: Ea = 8.3 +/- 0.4 Kcal/mol. When the Na+/Ca2+ exchanger is reconstituted into lipid vesicles of defined composition, we observe a higher activity if cholesterol is among the lipids. The activation energy becomes 6.1 +/- 0.1 Kcal/mol in this system, but the Arrhenius plot shows a decreased slope for temperatures above 33 degrees C.
从牛心室组织分离出的肌膜囊泡，当暴露于外向性的钠离子梯度时，会通过钠钙交换体积累钙离子。如果将这些囊泡转移到钙离子耗尽的钠缓冲液中，钙离子会通过相同的机制释放出来。使用钙离子指示剂偶氮胂III和停流分光光度计，我们可以直接跟踪钙离子外流的动力学过程。因此，我们可以通过钙离子释放的初始速率来测量钠钙交换体的活性。我们发现，它以协同方式依赖于外部钠离子浓度，希尔系数为2。通过研究钠钙交换的温度依赖性，我们发现它可以用单一的活化能来描述：Ea = 8.3 +/- 0.4千卡/摩尔。当钠钙交换体重组到特定组成的脂质囊泡中时，如果脂质中有胆固醇，我们会观察到更高的活性。在这个系统中，活化能变为6.1 +/- 0.1千卡/摩尔，但阿累尼乌斯图显示，在温度高于33摄氏度时斜率降低。
