Transmembrane domains of hepatitis C virus envelope glycoproteins:: Residues involved in E1E2 heterodimerization and involvement of these domains in virus entry
Transmembrane domains of hepatitis C virus envelope glycoproteins:: Residues involved in E1E2 heterodimerization and involvement of these domains in virus entry
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DOI：
10.1128/jvi.02198-06
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发表时间：
2007-03-01
影响因子：
5.4
通讯作者：
Dubuisson, Jean
中科院分区：
文献类型：
--
作者：
Ciczora, Yann;Callens, Nathalie;Dubuisson, Jean
丙型肝炎病毒（HCV）包膜糖蛋白E1和E2的跨膜（TM）结构域已被证明在E1 E2异二聚体的生物发生过程中发挥多种作用。通过在HCV包膜糖蛋白的TM结构域内使用丙氨酸扫描插入诱变，我们先前已经表明这些结构域的中心区域以及El的TM结构域的N-末端部分参与异源二聚化。在这里，我们使用这些区域的色氨酸置换扫描来识别参与这些相互作用的单个残基。我们的诱变研究确定了至少四个残基参与异源二聚化：Gly 354，Gly 358，Lys 370和Asp 728。有趣的是，Gly 354和Gly 358属于GXXXG寡聚基序。我们的色氨酸突变体也被用来产生逆转录病毒为基础的，HCV假型颗粒（HCVpp），以分析这些突变对病毒进入的影响。令人惊讶的是，与野生型相比，两种突变体始终显示出更高的感染性。与此相反，HCVpp感染性强烈影响许多突变体，尽管正常的E1 E2异源二聚化和正常水平的HCV糖蛋白纳入HCVpp。在最近开发的体外融合试验中，使用荧光标记的脂质体对这些HCVpp突变体中的一些进行了表征，表明在不改变E1 E2异源二聚化的情况下降低HCVpp感染性的突变影响了HCV包膜糖蛋白的融合特性。总之，该突变分析确定了E1 E2异源二聚化所涉及的残基，并揭示了HCV包膜糖蛋白的TM结构域在这些蛋白的融合特性中起主要作用。
The transmembrane (TM) domains of hepatitis C virus (HCV) envelope glycoproteins El and E2 have been shown to play multiple roles during the biogenesis of the E1E2 heterodimer. By using alanine scanning insertion mutagenesis within the TM domains of HCV envelope glycoproteins, we have previously shown that the central regions of these domains as well as the N-terminal part of the TM domain of El are involved in heterodimerization. Here, we used a tryptophan replacement scan of these regions to identify individual residues that participate in those interactions. Our mutagenesis study identified at least four residues involved in heterodimerization: Gly 354, Gly 358, Lys 370, and Asp 728. Interestingly, Gly 354 and Gly 358 belong to a GXXXG oligomerization motif. Our tryptophan mutants were also used to generate retrovirus-based, HCV-pseudotyped particles (HCVpp) in order to analyze the effects of these mutations on virus entry. Surprisingly, two mutants consistently displayed higher infectivity compared to that of the wild type. In contrast, HCVpp infectivity was strongly affected for many mutants, despite normal E1E2 heterodimerization and normal levels of incorporation of HCV glycoproteins into HCVpp. The characterization of some of these HCVpp mutants in the recently developed in vitro fusion assay using fluorescent-labeled liposomes indicated that mutations reducing HCVpp infectivity without altering E1E2 heterodimerization affected the fusion properties of HCV envelope glycoproteins. In conclusion, this mutational analysis identified residues involved in E1E2 heterodimerization and revealed that the TM domains of HCV envelope glycoproteins play a major role in the fusion properties of these proteins.