The carboxy-terminal sequence of the pestivirus glycoprotein Erns represents an unusual type of membrane anchor
The carboxy-terminal sequence of the pestivirus glycoprotein Erns represents an unusual type of membrane anchor
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DOI：
10.1128/jvi.79.18.11901-11913.2005
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发表时间：
2005-09-01
影响因子：
5.4
通讯作者：
Meyers, G
中科院分区：
文献类型：
--
作者：
Fetzer, C;Tews, BA;Meyers, G
E-rns蛋白是瘟病毒的结构糖蛋白，其缺乏典型的膜锚序列，并且已知从感染的细胞分泌。然而，大量的蛋白质保留在细胞内，并通过迄今未知的机制附着在病毒体上。用cDNA构建体进行的瞬时表达研究表明，在稳态情况下，在转染细胞的上清液中发现16%的蛋白质，而84%的蛋白质以细胞内蛋白质的形式出现。我们在这里表明，E-rns代表一种膜结合蛋白。膜结合通过E-rns的羧基末端区域发生。通过将该序列融合到绿色荧光蛋白（GFP）的羧基末端，报告蛋白的亚细胞定位从胞质转变为膜结合。在用GFP构建体的截短实验中引出膜结合所必需的11个氨基酸的核心序列。然而，该肽不足以赋予膜锚定，而是需要上游或下游辅助序列。不同提取方法的分析表明，E-rns既不像外周膜蛋白那样容易从膜上剥离，也不像跨膜蛋白那样与膜紧密结合。
The E-rns protein is a structural glycoprotein of pestiviruses that lacks a typical membrane anchor sequence and is known to be secreted from the infected cell. However, major amounts of the protein are retained within the cell and attached to the virion by a so far unknown mechanism. Transient-expression studies with cDNA constructs showed that in a steady-state situation, 16% of the protein is found in the supernatant of the transfected cells while 84% appears as intracellular protein. We show here that E-rns represents a membrane-bound protein. Membrane binding occurs via the carboxy-terminal region of E-rns. By fusion of this sequence to the carboxy terminus of green fluorescent protein (GFP), the subcellular localization of the reporter protein switched from cytosolic to membrane bound. A core sequence of 11 amino acids necessary for membrane binding was elicited in truncation experiments with GFP constructs. However, this peptide is not sufficient to confer membrane anchoring but needs either upstream or downstream accessory sequences. Analyses with different extraction procedures showed that E-rns is neither easily stripped from the membrane, like a peripheral membrane protein, nor as tightly membrane bound as a transmembrane protein.