Fischer I, Sapirstein V S
Department of Anatomy and Neurobiology, Medical College of Pennsylvania, Philadelphia 19129.
J Biol Chem. 1994 Oct 7;269(40):24912-9.
Plasmolipin is an 18-kDa proteolipid protein found in kidney and brain, where it is restricted to the apical surface of tubular epithelial cells and to mammalian myelinated tracts, respectively (Sapirstein, V.S., Nolan, C. E., Stadler, I.I., and Fischer, I. (1992) J. Neurosci. Res. 31, 96-102; Cochary, E. F., Bizzozero, O. A., Sapirstein, V. S., Nolan, C. E., and Fischer, I. (1990) J. Neurochem. 55, 602-610). Addition of plasmolipin to lipid bilayers induces the formation of ion channels, which are voltage-dependent and K(+)-selective (Tosteson, M. T., and Sapirstein, V. S. (1981) J. Membr. Biol 63, 77-84). The present study describes the isolation and cloning of plasmolipin cDNA that includes the sequence of the complete coding region of the protein, the analysis of plasmolipin mRNA expression and a proposed model for its membrane structure. Northern blot analysis 1) shows that plasmolipin is encoded by a 1.7-kilobase mRNA, 2) confirms that the distribution of plasmolipin is restricted to kidney and brain, and 3) indicates that the expression of plasmolipin mRNA in cultured oligodendrocytes increases with cell maturation consistent with changes in the level of the protein. Restriction enzyme digestion of DNA followed by Southern blot analysis indicates that plasmolipin is encoded by a single gene. Sequence analysis of plasmolipin cDNA shows an open reading frame encoding a 157-amino acid protein of 17.4 kDa. The deduced amino acid sequence confirms the hydrophobic nature and high helical content of the protein and predicts a structure with four transmembrane domains similar to several other small hydrophobic proteins implicated in ion movement. The proposed model for membrane topology shows an enrichment of hydroxyl groups within two of the transmembrane domains and places cysteine residues near the extracellular membrane surface. Examination of protein sequence data bases reveals little overall homology with other proteins including proteolipids; however, three of the four transmembrane segments of plasmolipin show strong similarity with known membrane transport proteins. These results indicate that plasmolipin is an unique proteolipid protein that may participate in ion transport events specific to select membrane domains.
质膜脂蛋白是一种18 kDa的蛋白脂质，存在于肾脏和大脑中，分别局限于肾小管上皮细胞的顶端表面和哺乳动物的有髓神经纤维束（萨皮尔斯坦，V.S.，诺兰，C.E.，斯塔德勒，I.I.，和菲舍尔，I.（1992年）《神经科学研究杂志》31卷，96 - 102页；科查里，E.F.，比佐泽罗，O.A.，萨皮尔斯坦，V.S.，诺兰，C.E.，和菲舍尔，I.（1990年）《神经化学杂志》55卷，602 - 610页）。将质膜脂蛋白添加到脂质双层中会诱导形成离子通道，这些通道具有电压依赖性且对K⁺具有选择性（托斯滕森，M.T.，和萨皮尔斯坦，V.S.（1981年）《膜生物学杂志》63卷，77 - 84页）。本研究描述了质膜脂蛋白cDNA的分离和克隆，其中包括该蛋白质完整编码区的序列，质膜脂蛋白mRNA表达的分析以及其膜结构的推测模型。Northern印迹分析表明：1）质膜脂蛋白由一个1.7千碱基的mRNA编码；2）证实质膜脂蛋白的分布局限于肾脏和大脑；3）表明在培养的少突胶质细胞中，质膜脂蛋白mRNA的表达随着细胞成熟而增加，这与该蛋白质水平的变化一致。用限制性内切酶消化DNA，随后进行Southern印迹分析表明质膜脂蛋白由单个基因编码。质膜脂蛋白cDNA的序列分析显示一个开放阅读框，编码一个17.4 kDa、含157个氨基酸的蛋白质。推导的氨基酸序列证实了该蛋白质的疏水性和高螺旋含量，并预测其结构有四个跨膜结构域，类似于其他一些与离子转运有关的小疏水蛋白质。推测的膜拓扑结构模型显示两个跨膜结构域内羟基富集，并且半胱氨酸残基位于细胞外膜表面附近。对蛋白质序列数据库的检查发现，与其他蛋白质（包括蛋白脂质）的总体同源性较低；然而，质膜脂蛋白的四个跨膜片段中的三个与已知的膜转运蛋白有很强的相似性。这些结果表明质膜脂蛋白是一种独特的蛋白脂质，可能参与特定膜结构域特有的离子转运事件。
