Sato S, Jinbu Y, Nakao M
J Biochem. 1986 Sep;100(3):643-9. doi: 10.1093/oxfordjournals.jbchem.a121756.
Human erythrocyte cytoskeletal ATPase was extracted with 0.2 mM ATP (pH 8.0) from Triton X-100 treated ghosts. The ATPase fraction contained mainly spectrin, actin, and band 4.1. When the ATPase fraction was applied to a Sepharose 4B column, 90% of the ATPase activity was recovered in a spectrin, actin, and band 4.1 complex fraction and none was detected in the spectrin fraction. A specific activity of the complex ATPase was 60-120 nmol/(mg protein X h). No ATPase activity was detected in the presence of EDTA. The presence of magnesium in the incubation medium was essential for the ATPase activity. The activity was activated by KCl and was almost completely inhibited by 10(-5) M free calcium in the presence of 0.2 mM MgCl2. The Ki for Ca2+ was 7 X 10(-7) M. Phalloidin and DNase 1, which affect actin, inhibited this K,Mg-ATPase activity by 95%, but cytochalasin B did not inhibit it. N-Ethylmaleimide activated the ATPase 1.6-fold. The order of affinity for nucleotides was ATP greater than ITP greater than CTP, ADP, AMP-PNP, GTP. A specific ATPase activity of purified actin was 50 nmol/(mg X h). These results suggest that the cytoskeletal ATPase is actin ATPase and the actin ATPase is activated by spectrin and band 4.1.
用0.2 mM ATP（pH 8.0）从经Triton X - 100处理的血影中提取人红细胞细胞骨架ATP酶。该ATP酶组分主要包含血影蛋白、肌动蛋白和4.1带蛋白。当将该ATP酶组分应用于琼脂糖4B柱时，90%的ATP酶活性在血影蛋白、肌动蛋白和4.1带蛋白复合物组分中回收，而在血影蛋白组分中未检测到活性。复合ATP酶的比活性为60 - 120 nmol/（mg蛋白质×小时）。在存在EDTA的情况下未检测到ATP酶活性。孵育介质中镁的存在对ATP酶活性至关重要。该活性被KCl激活，在存在0.2 mM MgCl2的情况下，几乎完全被10^(-5) M游离钙抑制。Ca2+的Ki为7×10^(-7) M。影响肌动蛋白的鬼笔环肽和DNase 1使这种K、Mg - ATP酶活性抑制95%，但细胞松弛素B不抑制它。N - 乙基马来酰亚胺使ATP酶活性激活1.6倍。对核苷酸的亲和力顺序为ATP＞ITP＞CTP、ADP、AMP - PNP、GTP。纯化肌动蛋白的比ATP酶活性为50 nmol/（mg×小时）。这些结果表明细胞骨架ATP酶是肌动蛋白ATP酶，并且肌动蛋白ATP酶被血影蛋白和4.1带蛋白激活。
