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Characterization of a protein candidate for the ion channel of influenza C virus and its expression.

丙型流感病毒离子通道候选蛋白的表征及其表达。

基本信息

批准号：
07670337
负责人：
HONGO Seiji
金额：
$ 1.41万
依托单位：
YAMAGATA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07670337/
关键词：
Influenza C virus M gene CM2 ion channel

项目摘要

The biochemical properties of a second protein (CM2) encoded by RNA segment 6 of influenza C virus were investigated.1.Three forms of CM2 with different electrophoretic mobilities (CM2_0, CM2a and CM2b) were detected in infected cells by immunoprecipitation with antiserum to GST-CM2 fusion protein. 2.A mannose-rich oligosaccharide core is added to unglycosylated CM2_0 (Mr-16,000) to form CM2a (Mr-18,000) and the processing of the carbohydrate chain from high mannose type to complex one converts CM2a into CM2b heterogeneous in electrophoretic mobility (Mr-22,000 to 30,000). 3.Labeling of infected cells with [^3H] palmitic acid showed that CM2 is fatty acylated. The fatty acid bond was sensitive to treatment with hydroxylamine and mercaptoethanol, which indicates a labile thioester-type linkage. 4.The CM2 protein was also found to form disulfide-linked dimers and tetramers on SDS-PAGE under nonreducing conditions. 5.Trypsin treatment of infected cell surface as well as that of microsome vesicles from infected cells followed by immunoprecipitation with antiserum to the GST fusion protein containing the 56 C-terminal amino acid residues of CM2 suggested that this domain is intracellular and exposed on the cytoplasmic side of microsomes. 6.A small amount of CM2 is incorporated into progeny virus particles.These results, altogether, suggest that CM2 is an integral membrane protein with biochemical properties similar to those of influenza A virus M2 and influenza B virus NB proteins.

本研究对丙型流感病毒第6段RNA编码的第二蛋白（CM 2）的生化特性进行了研究：1.用抗GST-CM 2融合蛋白的抗血清免疫沉淀法，在感染细胞中检测到CM2_0、CM 2a和CM 2b三种不同电泳迁移率的CM 2。2.将富含甘露糖的寡糖核心添加到未糖基化的CM2_0（Mr-16，000）中以形成CM 2a（Mr-18，000），并且将糖链从高甘露糖型加工成复合物1将CM 2a转化成电泳迁移率异质的CM 2b（Mr-22，000至30，000）。3.用[^3H]棕榈酸标记感染的细胞表明CM 2是脂肪酰化的。脂肪酸键是敏感的羟胺和巯基乙醇，这表明一个不稳定的硫酯型连接处理。4.还发现CM 2蛋白在非还原条件下在SDS-PAGE上形成二硫键连接的二聚体和四聚体。5.胰蛋白酶处理感染的细胞表面以及微粒体囊泡从感染的细胞，然后用抗血清免疫沉淀的GST融合蛋白含有56个C-末端氨基酸残基的CM 2表明，这个域是细胞内和暴露在细胞质侧的微粒体。6.CM2在子代病毒颗粒中有少量的掺入，这些结果表明CM 2是一种完整的膜蛋白，其生化性质与甲型流感病毒M2和B病毒NB蛋白相似。