Hirowatari Y, Hijikata M, Tanji Y, Shimotohno K
Virology Division, National Cancer Center Research Institute, Tokyo, Japan.
Virus Res. 1995 Jan;35(1):43-61. doi: 10.1016/0168-1702(94)00078-q.
Processing of the putative nonstructural (NS) proteins, p70(NS3), p4(NS4A), p27(NS4B), p58/56(NS5A), and p66(NS5B), of Japanese type hepatitis C virus (HCV) in insect cells was analyzed by using a baculovirus expression system. Products processed by the HCV serine proteinase (Cpro-2) were essentially identical to those found in mammalian cultured cells transiently producing the NS region of the HCV precursor polyprotein. A series of internal and carboxy (C)-terminal deletion experiments coupled with epitope scanning analysis showed that efficient cleavage at the Cpro-2-dependent processing sites, except at the p4(NS4A)/p27(NS4B) site, is not significantly influenced by those mutations. Efficient cleavage at p4(NS4A)/p27(NS4B) required about 40% of the NS5A N-terminal region. Estimation of the processing sites by determination of the N-terminal amino acid sequences of the processed products revealed that all the Cpro-2-dependent cleavages occurred at essentially identical sites to those reported for another HCV genotype, suggesting that Cpro-2 is a possible target for the development of a strain-independent anti-HCV agent.
利用杆状病毒表达系统分析了日本丙型肝炎病毒（HCV）推定的非结构（NS）蛋白p70（NS3）、p4（NS4A）、p27（NS4B）、p58/56（NS5A）和p66（NS5B）在昆虫细胞中的加工过程。由HCV丝氨酸蛋白酶（Cpro-2）加工的产物与在瞬时产生HCV前体多蛋白NS区的哺乳动物培养细胞中发现的产物基本相同。一系列内部和羧基（C）末端缺失实验以及表位扫描分析表明，除了在p4（NS4A）/p27（NS4B）位点外，在Cpro-2依赖性加工位点的有效切割不受这些突变的显著影响。在p4（NS4A）/p27（NS4B）处的有效切割需要约40%的NS5A N末端区域。通过测定加工产物的N末端氨基酸序列来估计加工位点，结果表明，所有Cpro-2依赖性切割都发生在与另一种HCV基因型报道的位点基本相同的位置，这表明Cpro-2可能是开发不依赖毒株的抗HCV药物的靶点。
