The pestivirus glycoprotein Erns is anchored in plane in the membrane via an amphipathic helix
The pestivirus glycoprotein Erns is anchored in plane in the membrane via an amphipathic helix
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DOI：
10.1074/jbc.m706803200
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发表时间：
2007-11-09
影响因子：
4.8
通讯作者：
Meyers, Gregor
中科院分区：
文献类型：
--
作者：
Tews, Birke Andrea;Meyers, Gregor
E-RNS是鼠疫病毒的一种结构糖蛋白，发现它通过COOH末端附着在病毒粒子和感染细胞内的膜上，尽管它缺乏疏水锚定序列。假设COOH末端序列折叠成两亲性的α-螺旋。丙氨酸插入扫描显示，Erns COOH末端与膜结合的能力因在多种位置插入单一氨基酸而显著降低。突变降低了推测的螺旋的非极化面的疏水性，导致膜结合减少。蛋白水解酶K保护分析表明，在微体膜存在的情况下体外翻译的Erns受到保护，而带有人工跨膜区和较短胞质标记的突变体则被蛋白酶处理缩短。在洋地黄素渗透性细胞内，融合到野生型ERNS的COOH末端的标签不能被抗体访问，但在相同的条件下，可以检测到带有标签位于人工跨膜区下游的变体。这些结果与ERNS的COOH-末端膜锚定代表平面嵌入膜中的两亲性螺旋的模型一致。膜的完整性对感染性病毒的恢复很重要。
E-rns is a structural glycoprotein of pestiviruses found to be attached to the virion and to membranes within infected cells via its COOH terminus, although it lacks a hydrophobic anchor sequence. The COOH- terminal sequence was hypothesized to fold into an amphipathic alpha-helix. Alanine insertion scanning revealed that the ability of the Erns COOH terminus to bind membranes is considerably reduced by the insertion of a single amino acid at a wide variety of positions. Mutations decreasing the hydrophobicity of the apolar face of the putative helix led to reduction of membrane association. Proteinase K protection assays showed that Erns translated in vitro in the presence of microsomal membranes was protected, whereas a mutant with an artificial transmembrane region and a short cytosolic tag was shortened by the protease treatment. A tag fused to the COOH terminus of wild type Erns was not accessible for antibodies within digitonin- permeabilized cells, but the variant with the tag located downstream of the artificial transmembrane region was detected under the same conditions. These results are in accordance with the model that the COOH- terminal membrane anchor of Erns represents an amphipathic helix embedded in plane into the membrane. The integrity of the membrane important for recovery of infectious virus.