Expression cloning of PIG-L, a candidate N-acetylglucosaminylphosphatidylinositol deacetylase
Expression cloning of PIG-L, a candidate N-acetylglucosaminylphosphatidylinositol deacetylase
复制标题
DOI：
10.1074/jbc.272.25.15834
复制
发表时间：
1997-06-20
影响因子：
4.8
通讯作者：
Kinoshita, T
中科院分区：
文献类型：
--
作者：
Nakamura, N;Inoue, N;Kinoshita, T
许多真核细胞表面蛋白通过糖基磷脂酰肌醇（GPT）锚结合到细胞膜上。已经使用在其生物合成途径中有缺陷的突变哺乳动物细胞系和酵母的互补来克隆涉及GPT锚生物合成的几个基因。然而，该途径的第二步，其中N-乙酰氨基葡萄糖磷脂酰肌醇（GlcNAc-PI）的N-脱乙酰化形成氨基葡萄糖（GlcN）-PI，参与的基因尚未被克隆。与已知GPI锚突变体细胞的互补分析表明，它属于与CHO细胞突变体G9 PLAP. 85相同的互补组。利用新的突变体，我克隆了一个参与这一步骤的大鼠基因，称为PIG-L（在bar下为磷脂酰肌醇聚糖类（L）），PIG-L编码252个氨基酸的内质网膜蛋白，其中大部分位于细胞质侧。PIG-L蛋白的这种取向与GPI锚生物合成的第二步发生在内质网的胞质侧的观点一致。
Many eukaryotic cell surface proteins are bound to the cell membrane by a glycosylphosphatidylinositol (GPT) anchor. Several genes involved in GPT anchor biosynthesis have been cloned using complementation of mutant mammalian cell lines and yeasts that are defective in its biosynthesis pathway. However, the gene involved in the second step of this pathway, in which N-acetylglucosaminyl-phosphatidylinositol (GlcNAc-PI) is N-deacetylated to form glucosaminyl (GlcN)-PI, has not been cloned, In this study, we established a GPI anchor-deficient mutant of Chinese hamster ovary (CHO) cells defective in the second step. Complementation analysis with the known GPI anchor mutant cells demonstrated that it belonged to the same complementation group as the CHO cell mutant G9PLAP.85. Using the new mutant, me cloned a rat gene termed PIG-L (for phosphatidylinositol glycan class (L) under bar) that is involved in this step, PIG-L encodes a 252-amino acid, endoplasmic reticulum membrane protein, most of which is ill the cytoplasmic side. This orientation of PIG-L protein is consistent with the notion that the second step of GPI anchor biosynthesis occurs on the cytoplasmic side of the endoplasmic reticulum.