EXPRESSION AND CHARACTERIZATION OF HEPATITIS-B VIRUS SURFACE-ANTIGEN POLYPEPTIDES IN INSECT CELLS WITH A BACULOVIRUS EXPRESSION SYSTEM
EXPRESSION AND CHARACTERIZATION OF HEPATITIS-B VIRUS SURFACE-ANTIGEN POLYPEPTIDES IN INSECT CELLS WITH A BACULOVIRUS EXPRESSION SYSTEM
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DOI：
10.1128/jvi.63.4.1549-1557.1989
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发表时间：
1989-04-01
影响因子：
5.4
通讯作者：
SUMMERS, MD
中科院分区：
文献类型：
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作者：
LANFORD, RE;LUCKOW, V;SUMMERS, MD
将杆状病毒苜蓿银纹夜蛾核型多角体病毒用作表达载体以产生具有和不具有前S结构域的B肝炎病毒表面抗原。S基因产物表达为融合和非融合多肽。在融合和非融合多肽的翻译后修饰中没有观察到差异。S蛋白不分泌到培养基中，但插入到内质网，糖基化，并部分挤出到内质网的内腔作为22 nm的脂蛋白颗粒。昆虫细胞来源的S蛋白上的寡糖链是N-连接的高甘露糖形式，与血浆来源的B病毒表面抗原上检测到的复合型寡糖相反。将前S-S多肽插入内质网，糖基化，并通过用肉豆蔻酸进行脂肪酸酰化修饰。本研究采用去污剂抽提和免疫亲和层析法从细胞膜上纯化S蛋白。纯化的S蛋白呈蛋白-去污剂胶束形式，具有高度的抗原性和免疫原性。
The baculovirus Autographa californica nuclear polyhedrosis virus was used as an expression vector to produce hepatitis B virus surface antigen with and without the pre-S domain. The S gene product was expressed as both fusion and nonfusion polypeptides. No difference was observed in the posttranslational modification of the fusion and nonfusion polypeptides. The S proteins were not secreted into the medium but were inserted into the endoplasmic reticulum, glycosylated, and partially extruded into the lumen of the endoplasmic reticulum as 22-nm lipoprotein particles. The oligosaccharide chains on the insect cell-derived S protein were of the N-linked high-mannose form, in contrast to the complex-type oligosaccharides detected on plasma-derived hepatitis B virus surface antigen. The pre-S-S polypeptides were inserted into the endoplasmic reticulum, glycosylated, and modified by fatty acid acylation with myristic acid. A procedure was developed to purify the S protein from cellular membranes by using detergent extraction and immunoaffinity chromatography. The purified S protein was in the form of protein-detergent micelles and was highly antigenic and immunogenic.