Association of spectrin with its membrane attachment site restricts lateral mobility of human erythrocyte integral membrane proteins
Association of spectrin with its membrane attachment site restricts lateral mobility of human erythrocyte integral membrane proteins
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血影蛋白与其膜附着位点的结合限制了人红细胞整合膜蛋白的横向移动性
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发表时间：
1978
期刊：
影响因子：
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通讯作者：
V. Bennett
中科院分区：
文献类型：
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作者：
V. Fowler;V. Bennett
血影蛋白和人红细胞膜内表面之间的相互作用已被牵连在控制的整体膜蛋白的横向流动性。我们在这里报告说，孵育的“渗漏”的红细胞与水溶性蛋白水解片段含有膜附着位点的血影蛋白实现了选择性和受控的解离血影蛋白从膜，并增加了荧光素异硫氰酸酯标记的整体膜蛋白（> 70%的标记带3和PAS-1）的横向流动率。膜蛋白的迁移率测量为仙台病毒将荧光红细胞与非荧光红细胞融合后，均匀荧光细胞百分比随时间的增加。由于病毒融合的红细胞失去了大部分血红蛋白，因此细胞对大分子是可渗透的。血影蛋白的膜附着位点已经通过由内而外的红细胞囊泡的有限蛋白水解而溶解，并且已经被纯化（V）。班尼特，生物化学杂志253：2292（1978）。这个72，000-道尔顿的片段与溶液中的血影蛋白结合，竞争性抑制32 P-血影蛋白与由内而外的囊泡结合，Ki为10− 7 M，并导致32 P-血影蛋白从囊泡中快速解离。经酸处理的72，000-道尔顿片段和条带3的45，000道尔顿-细胞质部分（也从蛋白水解消化物中分离）对血影蛋白结合、释放或膜蛋白迁移率均无影响。通过抑制血影蛋白与倒置囊泡结合并从这些囊泡中置换血影蛋白的相同多肽对膜蛋白横向流动性的增强提供了直接证据，表明血影蛋白与膜中蛋白组分的相互作用限制了红细胞中完整膜蛋白的横向流动性。
Interactions between spectrin and the inner surface of the human erythrocyte membrane have been implicated in the control of lateral mobility of the integral membrane proteins. We report here that incubation of “leaky” erythrocytes with a water-soluble proteolytic fragment containing the membrane attachment site for spectrin achieves a selective and controlled dissociation of spectrin from the membrane, and increases the rate of lateral mobility of fluorescein isothiocyanate-labeled integral membrane proteins (> 70% of label in band 3 and PAS-1). Mobility of membrane proteins is measured as an increase in the percentage of uniformly fluorescent cells with time after fusion of fluorescent with nonfluorescent erythrocytes by Sendai virus. The cells are permeable to macromolecules since virus-fused erythrocytes lose most of their hemoglobin. The membrane attachment site for spectrin has been solubilized by limited proteolysis of inside-out erythrocyte vesicles and has been purified (V). Bennett, J Biol Chem 253:2292 (1978). This 72,000-dalton fragment binds to spectrin in solution, competitively inhibits association of 32P-spectrin with inside-out vesicles with a Ki of 10−7M, and causes rapid dissociation of 32P-spectrin from vesicles. Both acid-treated 72,000-dalton fragment and the 45,000 dalton-cytoplasmic portion of band 3, which also was isolated from the proteolytic digest, have no effect on spectrin binding, release, or membrane protein mobility. The enhancement of membrane protein lateral mobility by the same polypeptide that inhibits binding of spectrin to inverted vesicles and displaces spectrin from these vesicles provides direct evidence that the interaction of spectrin with protein components in the membrane restricts the lateral mobility of integral membrane proteins in the erythrocyte.