Finidori J, Laperche Y, Haguenauer-Tsapis R, Barouki R, Guellaen G, Hanoune J
J Biol Chem. 1984 Apr 25;259(8):4687-90.
gamma-Glutamyl transpeptidase consists of two polypeptide chains anchored to the kidney brush-border membrane only through a short hydrophobic domain near the NH2-terminal end of the heavy subunit. The two subunits were reported to derive from a single polypeptide precursor by tissue labeling experiments. We have investigated the first steps of GGT biosynthesis and processing in a cell-free system. mRNA was prepared from kidney and enriched in specific sequences by a preparative gel electrophoresis. In vitro translation resulted in the synthesis of a single polypeptide (Mr = 63,000) specifically immunoprecipitated by antibodies raised against the mature dimeric enzyme. Incubation with microsomal membranes resulted in the appearance of a glycosylated form of the propeptide (Mr = 78,000). This latter form was cotranslationally segregated into microsomes and was sensitive to endoglycosidase H. Purified Escherichia coli leader peptidase did not process the primary gamma-glutamyl transpeptidase chain. This ectoprotein therefore appears to be inserted in the phospholipid bilayer without cleavage of a signal peptide, similar to most integral membrane proteins so far studied.
γ-谷氨酰转肽酶由两条多肽链组成，仅通过重亚基NH2末端附近的一个短疏水结构域锚定在肾刷状缘膜上。通过组织标记实验报道这两个亚基来源于单一多肽前体。我们已经在无细胞系统中研究了γ-谷氨酰转肽酶生物合成和加工的第一步。从肾脏制备mRNA，并通过制备性凝胶电泳富集特定序列。体外翻译导致合成了一种单一多肽（Mr = 63,000），该多肽被针对成熟二聚体酶产生的抗体特异性免疫沉淀。与微粒体膜一起孵育导致前肽糖基化形式（Mr = 78,000）的出现。后一种形式在翻译过程中被共分离到微粒体中，并且对内切糖苷酶H敏感。纯化的大肠杆菌前导肽酶不能加工初级γ-谷氨酰转肽酶链。因此，这种细胞外蛋白似乎在没有信号肽切割的情况下插入磷脂双层中，这与迄今为止研究的大多数整合膜蛋白相似。
