Zhou Q, Zhao J, Stout J G, Luhm R A, Wiedmer T, Sims P J
Blood Research Institute, The Blood Center of Southeastern Wisconsin, Milwaukee, Wisconsin 53201-2178, USA.
J Biol Chem. 1997 Jul 18;272(29):18240-4. doi: 10.1074/jbc.272.29.18240.
The rapid movement of phospholipids (PL) between plasma membrane leaflets in response to increased intracellular Ca2+ is thought to play a key role in expression of platelet procoagulant activity and in clearance of injured or apoptotic cells. We recently reported isolation of a approximately 37-kDa protein in erythrocyte membrane that mediates Ca2+-dependent movement of PL between membrane leaflets, similar to that observed upon elevation of Ca2+ in the cytosol (Bassé, F., Stout, J. G., Sims, P. J., and Wiedmer, T. (1996) J. Biol. Chem. 271, 17205-17210). Based on internal peptide sequence obtained from this protein, a 1,445-base pair cDNA was cloned from a K-562 cDNA library. The deduced "PL scramblase" protein is a proline-rich, type II plasma membrane protein with a single transmembrane segment near the C terminus. Antibody against the deduced C-terminal peptide was found to precipitate the approximately 37-kDa red blood cell protein and absorb PL scramblase activity, confirming the identity of the cloned cDNA to erythrocyte PL scramblase. Ca2+-dependent PL scramblase activity was also demonstrated in recombinant protein expressed from plasmid containing the cDNA. Quantitative immunoblotting revealed an approximately 10-fold higher abundance of PL scramblase in platelet ( approximately 10(4) molecules/cell) than in erythrocyte ( approximately 10(3) molecules/cell), consistent with apparent increased PL scramblase activity of the platelet plasma membrane. PL scramblase mRNA was found in a variety of hematologic and nonhematologic cells and tissues, suggesting that this protein functions in all cells.
磷脂（PL）响应细胞内Ca2+增加而在质膜小叶间的快速移动，被认为在血小板促凝血活性的表达以及受损或凋亡细胞的清除中起关键作用。我们最近报道了在红细胞膜中分离出一种约37 kDa的蛋白质，它介导PL在膜小叶间的Ca2+依赖性移动，类似于胞质溶胶中Ca2+升高时所观察到的情况（Bassé, F., Stout, J. G., Sims, P. J., and Wiedmer, T. (1996) J. Biol. Chem. 271, 17205 - 17210）。基于从该蛋白质获得的内部肽序列，从K - 562 cDNA文库中克隆出一个1445碱基对的cDNA。推导的“PL翻转酶”蛋白是一种富含脯氨酸的II型质膜蛋白，在C末端附近有一个单一的跨膜区段。发现针对推导的C末端肽的抗体能沉淀约37 kDa的红细胞蛋白并吸收PL翻转酶活性，证实克隆的cDNA与红细胞PL翻转酶的一致性。在含有该cDNA的质粒表达的重组蛋白中也证明了Ca2+依赖性PL翻转酶活性。定量免疫印迹显示血小板中PL翻转酶的丰度（约10(4)个分子/细胞）比红细胞中（约10(3)个分子/细胞）高约10倍，这与血小板质膜中PL翻转酶活性明显增加一致。在多种血液学和非血液学细胞及组织中发现了PL翻转酶mRNA，表明该蛋白在所有细胞中都发挥作用。
