Lanford R E, Notvall L, Chavez D, White R, Frenzel G, Simonsen C, Kim J
Department of Virology and Immunology, Southwest Foundation for Biomedical Research, San Antonio, Texas 78228.
Virology. 1993 Nov;197(1):225-35. doi: 10.1006/viro.1993.1583.
The baculovirus/insect cell expression system was used to express the capsid protein and glycoproteins (e1 and e2/NS1) of the hepatitis C virus (HCV). Each polypeptide domain was expressed individually using two different constructs varying at their carboxy termini in order to retain or delete hydrophobic domains that may be involved in membrane association. The capsid proteins were transported to the nucleus where they formed a single large crystal-like inclusion. The capsid proteins were phosphorylated in insect cells. The e1 and e2 polypeptides were present in both the soluble and insoluble cellular fractions. Deletion of a hydrophobic domain in the carboxy terminus of e2 resulted in the polypeptide becoming soluble but not secreted. Deletion of the carboxy terminus of e1 had no effect on solubility. Both e1 and e2 were glycosylated, with variable glycosylation of e1 giving rise to a series of polypeptides varying in apparent molecular weight. Co-infection of insect cells with viruses expressing e1 and e2 resulted in a complex that permitted the coimmunoprecipitation of e1 with antibodies to e2 and vice versa. Immunofluorescence staining of insect cells expressing e1 and e2 indicated that reactivity to e2 was more prevalent in anti-HCV positive human sera.
杆状病毒/昆虫细胞表达系统被用于表达丙型肝炎病毒（HCV）的衣壳蛋白和糖蛋白（E1和E2/NS1）。每个多肽结构域通过使用两个在羧基末端不同的构建体单独表达，以保留或删除可能参与膜结合的疏水结构域。衣壳蛋白被转运到细胞核，在那里它们形成单个大的晶体样包涵体。衣壳蛋白在昆虫细胞中被磷酸化。E1和E2多肽存在于可溶性和不溶性细胞组分中。E2羧基末端疏水结构域的缺失导致该多肽变得可溶但不分泌。E1羧基末端的缺失对溶解性没有影响。E1和E2都被糖基化，E1的可变糖基化产生了一系列表观分子量不同的多肽。用表达E1和E2的病毒共感染昆虫细胞产生了一种复合物，该复合物允许用抗E2抗体共免疫沉淀E1，反之亦然。对表达E1和E2的昆虫细胞进行免疫荧光染色表明，抗HCV阳性人血清中对E2的反应性更普遍。
