Cohen C M, Kramer R M, Branton D
Biochim Biophys Acta. 1980 Mar 27;597(1):29-40. doi: 10.1016/0005-2736(80)90147-9.
Erythrocyte and HeLa cell plasma membranes were isolated on polylysine-coated polyacrylamide beads and the transbilayer disposition of their proteins was investigated. When membranes of intact erythrocytes were isolated on beads and then labelled by lactoperoxidase-catalysed iodination, their labelling pattern was similar to that of inside-out vesicles in solution. When the membranes of intact HeLa cells were isolated on beads and then labelled by galactose oxidase-[3H]borohydride treatment, no glycoprotein or glycolipid sugars were accessible. On the other hand, when the HeLa cell membranes were isolated on beads and then labelled by the lactoperoxidase-catalysed iodination, all of the major membrane proteins were iodinated. These experiments confirmed for HeLa cell membranes what had previously been shown for erythrocyte membranes: when the membranes of intact cells are isolated on beads, the accessibility of their surfaces to enzymatic probes is the same as would be expected of inside-out vesicles in suspension. Double-label experiments, in which the HeLa cell membranes were labelled first on the intact HeLa cells and again after isolation on beads, identified several proteins which may span the membrane.
红细胞和HeLa细胞质膜在聚赖氨酸包被的聚丙烯酰胺珠上分离，并研究其蛋白质的跨膜分布。当完整红细胞的膜在珠子上分离，然后通过乳过氧化物酶催化碘化进行标记时，其标记模式与溶液中外翻囊泡的标记模式相似。当完整HeLa细胞的膜在珠子上分离，然后通过半乳糖氧化酶-[3H]硼氢化钠处理进行标记时，没有糖蛋白或糖脂糖可供标记。另一方面，当HeLa细胞膜在珠子上分离，然后通过乳过氧化物酶催化碘化进行标记时，所有主要膜蛋白都被碘化。这些实验证实了HeLa细胞膜与先前红细胞膜的情况相同：当完整细胞的膜在珠子上分离时，其表面对酶探针的可及性与悬浮中外翻囊泡的预期相同。双标记实验中，HeLa细胞膜首先在完整的HeLa细胞上标记，然后在珠子上分离后再次标记，鉴定出了几种可能跨膜的蛋白质。
