Ortega A, Lepock J R
Department of Biology, University of Waterloo, Ontario, Canada.
Biochim Biophys Acta. 1995 Jan 26;1233(1):7-13. doi: 10.1016/0005-2736(94)00243-i.
The presence of calcium stimulated adenosine triphosphatase (Ca2+,Mg(2+)-ATPase) activity in isolated transverse tubule (t-tubule) membranes is distinguished from magnesium adenosine triphosphatase (Mg(2+)-ATPase) activity on the basis of differing thermal stabilities. The Mg(2+)-ATPase is the major protein component of the t-tubule membrane, and it can be difficult to discriminate between the low levels of Ca2+ stimulated ATPase activity found in isolates of t-tubules compared to the much higher Mg(2+)-ATPase activity. Thermal analysis reveals different inactivation temperatures (Ti) for the proteins responsible for ATP dependent calcium transport (Ti = 49 degrees C) and Mg(2+)-ATPase activity (Ti = 57 degrees C) in isolated t-tubule membranes. The differential scanning calorimetry profile of t-tubule membranes consists of three major components with transition temperatures (Tm) of 51 degrees C, 57 degrees C and 63 degrees C. Denaturation of the component with Tm = 57 degrees C correlates with inactivation of Mg(2+)-ATPase activity, and denaturation of the Tm = 51 degrees C component correlates with the inactivation of Ca2+,Mg(2+)-ATPase activity and calcium transport. The functions of the t-tubule membrane component or components that denature with Tm = 63 degrees C have yet to be identified. The lack of stimulation of calcium transport in isolated t-tubules by oxalate, the impermeability of isolated t-tubules to oxalate, and experiments performed on t-tubules with defined amounts of sarcoplasmic reticulum (SR) added suggest that contamination of the isolated t-tubules by SR is unlikely to account for the level of Ca2+,Mg(2+)-ATPase activity detected. The presence of a Ca2+,Mg(2+)-ATPase in the t-tubule membrane would provide a mechanism that may be involved in the partial removal of calcium that is accumulated in the junctional space during muscle relaxation or calcium that is released from the terminal cisternae of sarcoplasmic reticulum during excitation-contraction coupling.
在分离的横管（T管）膜中，钙刺激的三磷酸腺苷酶（Ca2 +，Mg(2 +)-ATP酶）活性的存在与镁三磷酸腺苷酶（Mg(2 +)-ATP酶）活性在热稳定性方面存在差异。Mg(2 +)-ATP酶是T管膜的主要蛋白质成分，与高得多的Mg(2 +)-ATP酶活性相比，很难区分T管分离物中低水平的钙刺激ATP酶活性。热分析揭示了分离的T管膜中负责ATP依赖性钙转运的蛋白质（Ti = 49℃）和Mg(2 +)-ATP酶活性（Ti = 57℃）的不同失活温度（Ti）。T管膜的差示扫描量热曲线由三个主要成分组成，转变温度（Tm）分别为51℃、57℃和63℃。Tm = 57℃成分的变性与Mg(2 +)-ATP酶活性的失活相关，而Tm = 51℃成分的变性与Ca2 +，Mg(2 +)-ATP酶活性和钙转运的失活相关。Tm = 63℃变性的T管膜成分或成分的功能尚未确定。草酸盐对分离的T管中的钙转运缺乏刺激作用、分离的T管对草酸盐的不渗透性以及对添加了确定量肌浆网（SR）的T管进行的实验表明，分离的T管被SR污染不太可能解释检测到的Ca2 +，Mg(2 +)-ATP酶活性水平。T管膜中存在Ca2 +，Mg(2 +)-ATP酶将提供一种机制，该机制可能参与在肌肉松弛期间在连接间隙中积累的钙或在兴奋 - 收缩偶联期间从肌浆网终末池释放的钙的部分清除。
