INVIVO PHOSPHORYLATION AND PROTEIN-ANALYSIS OF HEPATITIS-B VIRUS CORE ANTIGEN
INVIVO PHOSPHORYLATION AND PROTEIN-ANALYSIS OF HEPATITIS-B VIRUS CORE ANTIGEN
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DOI：
10.1128/jvi.61.4.955-961.1987
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发表时间：
1987-04-01
影响因子：
5.4
通讯作者：
SIDDIQUI, A
中科院分区：
文献类型：
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作者：
ROOSSINCK, MJ;SIDDIQUI, A
B型肝炎病毒的C开放阅读框含有两个框内ATG密码子，其被前C区分开，并编码两种抗原性不同的主要多肽，其可能由单个mRNA合成。前核心区指导e抗原的分泌，而核心抗原可以在不存在这些序列的情况下表达。本研究应用瞬时表达系统对B型肝炎病毒核心抗原进行了研究。通过使用腺病毒主要晚期启动子-猿猴病毒40增强子序列的嵌合复合物，我们能够在转染的细胞中实现高水平的核心抗原表达，允许通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳表征该蛋白质并进行分析。核心多肽是一个20.9千道尔顿的蛋白质，我们在这项研究中表明，它是在体内磷酸化。细胞分级分离的研究，其结果是由间接免疫荧光支持，定位在胞质溶胶和细胞核中的磷酸核心，并表明它是与转染细胞的膜。Triton X-114增溶研究的结果表明，磷酸化核心是周边与细胞质膜。膜相关磷酸化核心的表达发生在前核心序列的情况下。在没有其他病毒基因产物或完整DNA的情况下，磷酸核心也组装成颗粒。
The C open reading frame of the hepatitis B virus contains two in-frame ATG codons that are separated by the precore region and encodes two major polypeptides that are antigenically distinct and that are probably synthesized from individual mRNAs. The precore region directs the secretion of the e antigen, whereas the core antigen can be expressed in the absence of these sequences. In this report a transient expression system was used to study the hepatitis B virus core antigen. By using a chimeric complex of adenovirus major late promoter-simian virus 40 enhancer sequences, we were able to achieve high levels of core antigen expression in transfected cells, permitting characterization of this protein and analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The core polypeptide is a 20.9-kilodalton protein, and we show in this study that it is phosphorylated in vivo. Cell fractionation studies, the results of which are supported by indirect immunofluorescence, localized the phosphocore in the cytosol and the nucleus and indicated that it is associated with the membrane of transfected cells. Results of Triton X-114 solubilization studies indicated that the phosphocore is peripherally associated with cytoplasmic membranes. Expression of the membrane-associated phosphocore occurred in the absence of the precore sequences. The phosphocore also assembled into particles in the absence of other viral gene products or intact DNA.