Substrate Regulation of Membrane Phosphorylation and of Ca2+ Transport in the Sarcoplasmic Reticulum
Substrate Regulation of Membrane Phosphorylation and of Ca2+ Transport in the Sarcoplasmic Reticulum
复制标题
肌浆网膜磷酸化和 Ca2+ 转运的底物调节
DOI：
10.1016/s0021-9258(19)43982-3
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发表时间：
1973
影响因子：
4.8
通讯作者：
M. C. Mello
中科院分区：
文献类型：
--
作者：
L. Meis;M. C. Mello
用不同底物研究了钙摄取、钙依赖性核苷三磷酸酶活性和钙依赖性核苷三磷酸转移到肌浆网膜蛋白的能力。半最大膜磷酸化所需的Ca 2+浓度随温度、所用底物及其浓度而变化。使用0°下10 µm至60 µm的ITP或GTP浓度，可以测量膜磷酸化的初始速率。在0°时，GTP（0.45 µm）和ITP（0.31 µm）的膜磷酸化的表观Km被测定。通过竞争实验，计算了其它底物的表观Km值。发现的值为：乙酰磷酸，3.8 × 10 ~（-4）m; UTP，1.3 × 10 ~（-4）m; CTP，4.0 × 10 ~（-5）m; ATP，2.4 × 10 ~（-8）m。在0°时，CDP、IDP或GDP逆转该反应的能力比ADP低180倍。ITP、GTP和ATP增加膜去磷酸化的速率。碱离子抑制骨骼肌肌浆网囊泡的Ca 2+转运、底物水解和膜磷酸化。Na+和K+的抑制活性随底物及其浓度、Ca 2+浓度和温度的变化而变化，根据实验数据提出了反应顺序。
Calcium uptake, Ca2+-dependent nucleoside triphosphatase activity, and Ca2+-dependent transfer of the γ-phosphate of nucleoside triphosphate to the membrane protein of isolated sarcoplasmic reticulum fragments were studied using different substrates.Ca2+is a limiting factor for maximal membrane phosphorylation. The Ca2+concentration required for half-maximal membrane phosphorylation varies with the temperature, the substrate used, and its concentration.Using ITP or GTP concentrations ranging from 10 µm to 60 µm at 0°, it is possible to measure the initial rate of membrane phosphorylation. With the use of ATP, this rate is very fast and cannot be measured by the techniques employed.The apparentKmfor membrane phosphorylation at 0° was determined for GTP (0.45 µm) and ITP (0.31 µm). Through competitive experiments the apparentKmof other substrates were calculated. The values found were: acetyl phosphate, 3.8 x 10-4m; UTP, 1.3 x 10-4m; CTP, 4.0 x 10-5m, and ATP, 2.4 x 10-8m.The membrane phosphorylation reaction can be reversed by different nucleoside diphosphates. At 0° the ability of CDP, IDP, or GDP to reverse this reaction is 180 times lower than ADP.ITP, GTP, and ATP increase the rate of membrane dephosphorylation.Alkali ions inhibit the Ca2+transport, substrate hydrolysis, and membrane phosphorylation of skeletal muscle sarcoplasmic reticulum vesicles. The inhibitory activity of Na+and K+varies with the substrate used and its concentration, the Ca2+concentration and the temperature.A reaction sequence is proposed on the basis of the data presented.