Abe M, Shirakata Y, Sato D, Konishi K, Watanabe T, Nakamura J
Biological Institute, Faculty of Science, Tohoku University, Miyagi.
J Biochem. 1992 Dec;112(6):822-7. doi: 10.1093/oxfordjournals.jbchem.a123983.
Sarcoplasmic reticulum with calcium transport activity has been isolated from the cross-striated adductor muscle of the scallop, which lives in cold (< or = 20 degrees C) sea water, by using pH 7.0 buffer solution both to homogenize the tissue and to sediment the membrane fraction. The yield of the preparation was 60-100 mg protein from 100 g of the scallop muscle. Ca(2+)-activated ATPase protein of about 100 kDa accounted for 40-50% of the protein preparation. The maximum activities of ATP-dependent, oxalate-facilitated calcium accumulation and Ca(2+)-ATPase were observed at a pH of about 7.0 and temperature of 20-30 degrees C, and their values were about 2 mumol Ca2+/mg of protein/min and about 3 mumol ATP hydrolysis/mg of protein/min, respectively. At 0 degree C, 10-20% of these activities was maintained, while at 37 degrees C, the activities were irreversibly lost. The Ca(2+)-ATPase activity was half-maximally activated at about 0.3 microM [Ca2+]. The ATPase activity exhibited non-Michaelian behavior with respect to ATP, with two different Km values of approximately 10 microM and 0.1-0.3 mM. GTP, CTP, and ITP were also hydrolyzed by the preparation at a rate of 10-30% of that of ATP. The preparation was stored at -80 degrees C with retention of function for about a year.
利用pH 7.0的缓冲溶液匀浆组织并沉淀膜组分，从生活在寒冷（≤20℃）海水中的扇贝横纹收肌中分离出具有钙转运活性的肌浆网。制备物的产量为每100克扇贝肌肉中含60 - 100毫克蛋白质。约100 kDa的Ca(2 + ) - 激活ATP酶蛋白占蛋白质制备物的40 - 50%。在pH约为7.0、温度为20 - 30℃时，观察到ATP依赖性、草酸盐促进的钙积累和Ca(2 + ) - ATP酶的最大活性，其值分别约为2 μmol Ca2 + /毫克蛋白质/分钟和约3 μmol ATP水解/毫克蛋白质/分钟。在0℃时，这些活性可维持10 - 20%，而在37℃时，活性不可逆丧失。Ca(2 + ) - ATP酶活性在约0.3 μM [Ca2 + ]时被半最大激活。ATP酶活性对ATP表现出非米氏行为，有两个不同的Km值，分别约为10 μM和0.1 - 0.3 mM。GTP、CTP和ITP也被该制备物以ATP水解速率的10 - 30%水解。该制备物保存在 - 80℃，功能可保留约一年。
