Partial purification of Na+-Ca2+ antiporter from plasma membrane of chick heart.
Partial purification of Na+-Ca2+ antiporter from plasma membrane of chick heart.
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从鸡心脏质膜中部分纯化 Na -Ca2 逆向转运蛋白。
DOI：
10.1016/0005-2736(82)90478-3
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发表时间：
1982
期刊：
影响因子：
--
通讯作者：
K. Goshima
中科院分区：
文献类型：
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作者：
S. Wakabayashi;K. Goshima
摘要为了研究Na+− Ca 2+交换，在磷脂存在下，用胆酸盐溶解鸡心脏膜囊泡的蛋白质，并用链霉蛋白酶处理胆酸盐提取物。这些纯化的脂蛋白体，通过随后的稀释和离心以除去胆酸盐而重构，催化依赖于脂蛋白体内Na+（Na i+）浓度的Ca 2+摄取。在25° C下，脂质体中积累的最大Ca 2+量为140 nmol/mg蛋白质，Na i+-依赖性Ca 2+摄取的初始速率通常为20至40 nmol/mg/3 s，但对于来自未用链霉蛋白酶处理的胆酸盐提取物的粗脂蛋白体，仅为2至4 nmol/mg/3 s。因此，链霉蛋白酶处理导致10倍纯化。Na i+−依赖性的Ca 2+的摄取纯化的蛋白脂质体是30- 50倍高于初始膜囊泡。纯化的蛋白脂质体中Na i+−依赖性Ca 2+摄取的基本性质，如Ca 2+的Krmm，Na+和pH依赖性的敏感性，几乎与膜囊泡和粗蛋白脂质体中的那些相同。因此，链霉蛋白酶处理对于获得含有高度富集的Na+-Ca 2+反向转运蛋白的功能完整的重构脂质体非常有用。
Abstract To study Na+− Ca 2+ exchange, proteins of membrane vesicles from chick hearts were solubilized with cholate in the presence of phospholipids and the cholate extract was treated with pronase. These purified proteoliposomes, reconstituted by subsequent dilution and centrifugation to eliminate the cholate, catalyzed Ca 2+ uptake depending on the intraliposomal Na+(Na i+) concentration. The maximal amount of Ca 2+ accumulating in the liposomes was 140 nmol/mg protein and the initial rates of Na i+− dependent Ca 2+ uptake were routinely 20 to 40 nmol/mg per 3 s at 25° C, but only 2 to 4 nmol/mg per 3 s for the crude proteoliposomes from the cholate extract not treated with pronase. Thus the pronase treatment resulted in 10-fold purification. Na i+− dependent Ca 2+ uptake by purified proteoliposomes was 30-to 50-fold higher than that by the initial membrane vesicles. The fundamental properties of Na i+− dependent Ca 2+ uptake in purified proteoliposomes such as K rmm for Ca 2+, the sensitivity for Na+ and pH dependency, were nearly equal to those in membrane vesicles and crude proteoliposomes. Thus, pronase treatment was very useful for obtaining reconstituted liposomes containing highly enriched Na+-Ca 2+ antiporters which were functionally intact.