Processing of E1 and E2 glycoproteins of hepatitis C virus expressed in mammalian and insect cells.
Processing of E1 and E2 glycoproteins of hepatitis C virus expressed in mammalian and insect cells.
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哺乳动物和昆虫细胞中表达的丙型肝炎病毒 E1 和 E2 糖蛋白的加工。
DOI：
10.1006/viro.1994.1629
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发表时间：
1994
期刊：
影响因子：
3.7
通讯作者：
Tatsuo Miyamura
中科院分区：
文献类型：
--
作者：
Y. Matsuura;Tetsuro Suzuki;R. Suzuki;Mitsuru Sato;H. Aizaki;Izumu Saito;Tatsuo Miyamura
利用覆盖丙型肝炎病毒（HCV）包膜糖蛋白（E1和E2）结构区和部分非结构区的cDNA克隆，研究了HCV包膜糖蛋白的加工过程。在哺乳动物和昆虫细胞中表达的cDNA克隆通过丙型肝炎患者的血清和针对在昆虫细胞或大肠杆菌中表达的重组蛋白的单克隆和多克隆抗体进行免疫沉淀。在昆虫和哺乳动物细胞中表达的E2蛋白是60 kDa的糖蛋白（gp 60），通过N-聚糖酶去除糖残基得到38-和40-kDa的蛋白质。脉冲追踪实验表明，包膜蛋白的有效表达和加工需要与侧翼核心蛋白和NS 2蛋白共表达。在表达结构蛋白和NS蛋白的重组杆状病毒（NS 2和NS 3）感染的细胞中，不仅E1和E2蛋白被抗E1或抗E2单克隆抗体共沉淀，而且NS 2和NS 3蛋白也被抗E1或抗E2单克隆抗体共沉淀，而NS 3抗体仅沉淀NS 3蛋白。的协会E1和E2蛋白的存在下，还原剂的影响，并仍然观察到在与缺失突变体缺乏内部和C-末端疏水区的每种蛋白质的共感染的细胞。此外，E1和E2蛋白的截短形式被分泌到培养上清液中，并且其中一些仍然彼此缔合。
Processing of the envelope glycoproteins (E1 and E2) of hepatitis C virus (HCV) was investigated by using cDNA clones covering the structural and part of the nonstructural (NS) protein regions. The cDNA clones expressed in mammalian and insect cells were immunoprecipitated by serum of a hepatitis C patient and by monoclonal and polyclonal antibodies raised against the recombinant proteins expressed in insect cells or Escherichia coli. The E2 protein expressed in both insect and mammalian cells was a glycoprotein of 60 kDa (gp60) and removal of the sugar residues by N-glycanase yielded 38- and 40-kDa proteins. Pulse-chase experiments revealed that efficient expression and processing of the envelope proteins required coexpression with the flanking core and NS2 proteins. Not only E1 and E2 proteins but also NS2 and NS3 proteins were coprecipitated by anti-E1 or anti-E2 monoclonal antibody in the cells infected with the recombinant baculovirus expressing structural and NS proteins (NS2 and NS3), while only the NS3 protein was precipitated by anti-NS3 antibody. The association of E1 and E2 proteins was not influenced by the presence of a reducing agent and was still observed in the cells coinfected with the deletion mutants lacking both internal and C-terminal hydrophobic regions of each protein. Furthermore, the truncated forms of the E1 and E2 proteins were secreted into the culture supernatant and some of them were still associated with each other.