Meyers G, Wirblich C, Thiel H J, Thumfart J O
Institute of Immunology, Federal Research Centre for Virus Diseases of Animals, Tübingen, D-72001, Germany.
Virology. 2000 Oct 25;276(2):349-63. doi: 10.1006/viro.2000.0545.
Rabbit hemorrhagic disease virus (RHDV) belongs to the family Caliciviridae. Studies on this virus are hampered by the lack of a convenient cell culture system. To study viral protein expression a cDNA construct containing the entire protein-coding region of the virus was established and used for transient expression studies. After metabolic labeling of transfected cells and immunoprecipitation with a set of RHDV-specific antisera a variety of polypeptides were identified and assigned to defined regions of the viral genome. The consensus sequences of already identified or putative proteolytic cleavage sites in the viral polyprotein were changed by the introduction of mutations into the expression construct. Expression of these mutated constructs and analysis of the protein patterns allowed us to identify novel cleavage sites in the polyprotein and revealed the first details regarding the order of polyprotein processing.
兔出血症病毒（RHDV）属于杯状病毒科。由于缺乏便捷的细胞培养系统，对该病毒的研究受到阻碍。为了研究病毒蛋白表达，构建了一个包含病毒完整蛋白质编码区的cDNA构建体，并用于瞬时表达研究。在用一组RHDV特异性抗血清对转染细胞进行代谢标记和免疫沉淀后，鉴定出了多种多肽，并将其定位到病毒基因组的特定区域。通过在表达构建体中引入突变，改变了病毒多聚蛋白中已鉴定或推测的蛋白水解切割位点的共有序列。这些突变构建体的表达和蛋白质模式分析使我们能够鉴定多聚蛋白中的新切割位点，并揭示了多聚蛋白加工顺序的首批细节。
