摘要
Coronaviruses are enveloped RNA viruses with single positive stranded genomic RNA of approximately 32 kilobases which encodes at least 3 structural proteins; a nucleocapsid protein (N), a matrix glycoprotein (M) and a spike glycoprotein (S), as well as several nonstructural proteins (1). The S protein forms the projecting spikes or peplomers on the surface of the virus particle (2). This protein is considered to be involved in the attachment of virus to susceptible cells, the induction of cell-to-cell fusion, and the induction of neutralizing antibodies (3,4). It is also believed that the S protein is important for determining the pathogenic potential of murine coronaviruses (5,6). For more detailed structural and functional analyses of S protein, we need to have large quantities of S protein uncontaminated with other viral proteins. For this purpose, we have sought to express the S gene product in a baculovirus expression vector. The Autographa californica nuclear polyhedrosis virus (AcNPV) insect baculovirus vector was chosen because of the high levels of foreign gene expression obtained using the AcNPV polyhedrin promoter (7). Also, translational modifications such as glycosylation, phosphorylation, and cleavage of the signal peptide are known to occur in this system. In this paper, we describe the expression of the MHV strain JHM (JHMV) S protein by recombinant baculovirus in insect cells, characterization of the protein and analysis of epitopes existing on the S protein.
摘要译文
冠状病毒是具有约32千碱基的单个正链基因组RNA的包膜RNA病毒，其编码至少3种结构蛋白;核衣壳蛋白（N），基质糖蛋白（M）和刺突糖蛋白（S），以及几种非结构蛋白（1）。 S蛋白在病毒颗粒表面形成突出的突起或突起（2）。这种蛋白质被认为与病毒对易感细胞的附着，细胞与细胞融合的诱导以及中和抗体的诱导有关（3,4）。也认为S蛋白对确定鼠冠状病毒的致病潜力很重要（5,6）。对于S蛋白的更详细的结构和功能分析，我们需要大量的S蛋白未被其他病毒蛋白污染。为此，我们试图在杆状病毒表达载体中表达S基因产物。他选择了Autographa californica核型多角体病毒（AcNPV）昆虫杆状病毒载体，因为使用AcNPV多角体蛋白启动子（7）获得高水平的外源基因表达。此外，已知在该系统中发生翻译修饰，例如糖基化，磷酸化和信号肽的切割。在本文中，我们描述了重组杆状病毒在昆虫细胞中MHV毒株JHM（JHMV）S蛋白的表达，蛋白质的表征和S蛋白上存在的表位的分析。
Fumihiro Taguchi1;Sayaka Yoden1;Stuart Siddell2;Tateki Kikuchi1. Expression of the Spike Protein of Murine Coronavirus JHM Using a Baculovirus Vector. Coronaviruses and their Diseases[M].DE: Springer, 1990: 211-216