Proteolytic processing of the open reading frame 1b-encoded part of arterivirus replicase is mediated by nsp4 serine protease and is essential for virus replication
Proteolytic processing of the open reading frame 1b-encoded part of arterivirus replicase is mediated by nsp4 serine protease and is essential for virus replication
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DOI：
10.1128/jvi.73.3.2027-2037.1999
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发表时间：
1999-03-01
影响因子：
5.4
通讯作者：
Snijder, EJ
中科院分区：
文献类型：
--
作者：
van Dinten, LC;Rensen, S;Snijder, EJ
马动脉炎病毒（EAV）复制酶的开放阅读框（ORF）1b编码部分在基因组翻译过程中通过核糖体移码表达，从而产生ORF 1ab融合蛋白（345 kDa）。四种ORF 1b编码的加工产物，nsp 9（p80）、nsp 10（p50）、nsp 11（p26）和nsp 12（p12），先前已在EAV感染的细胞中鉴定（L. C.货车Dinten，A. L. M. Wassenaar，A. E. Gorbalenya，W. J. M. Spaan和E. J. Snijder，J. Virol. 70：6625-6633，1996）。在本研究中，这四种非结构蛋白的产生被证明是由nsp 4丝氨酸蛋白酶介导的，该蛋白酶是主要的病毒蛋白酶（E. J. Snijder，A. L. M.瓦塞纳尔湖C.货车丁滕J. M. Spaan和A. E. Gorbalenya，J.Biol.Chem.271：4864-4871，1996）。对候选切割位点的诱变显示Glu-2370/Ser、Gln-2837/Ser和Glu-3056/Gly分别是可能的nsp 9/10、nsp 10/11和nsp 11/12连接。将消除ORF 1b蛋白加工的突变引入最近开发的感染性cDNA克隆（L. C.货车丁滕den布恩，A. L. M. Wassenaar，W. J. M. Spaan和E. J. Snijder，Proc. Natl. Acad. Sci. USA 94：991-997，1997）。对这些突变体的分析表明，ORF 1b加工的选择性阻断影响了EAV繁殖的不同阶段。特别是，具有nsp 10/11切割位点突变Gln-2837-->Pro的突变体显示出不寻常的表型，因为它仍然能够合成RNA，但不能产生感染性病毒。
The open reading frame (ORF) 1b-encoded part of the equine arteritis virus (EAV) replicase is expressed by ribosomal frameshifting during genome translation, which results in the production of an ORF1ab fusion protein (345 kDa). Four ORF1b-encoded processing products, nsp9 (p80), nsp10 (p50), nsp11 (p26), and nsp12 (p12), have previously been identified in EAV-infected cells (L. C. van Dinten, A. L. M. Wassenaar, A. E. Gorbalenya, W. J. M. Spaan, and E. J. Snijder, J. Virol. 70:6625-6633, 1996). In the present study, the generation of these four nonstructural proteins was shown to be mediated by the nsp4 serine protease, which is the main viral protease (E. J. Snijder, A. L. M. Wassenaar, L. C. van Dinten, W. J. M. Spaan, and A. E. Gorbalenya, J. Biol. Chem. 271:4864-4871, 1996). Mutagenesis of candidate cleavage sites revealed that Glu-2370/Ser, Gln-2837/Ser, and Glu-3056/Gly are the probable nsp9/10, nsp10/11, and nsp11/12 junctions, respectively. Mutations which abolished ORF1b protein processing were introduced into a recently developed infectious cDNA clone (L. C. van Dinten, J. A. den Boon, A. L. M. Wassenaar, W. J. M. Spaan, and E. J. Snijder, Proc. Natl. Acad. Sci. USA 94:991-997, 1997). An analysis of these mutants showed that the selective blockage of ORF1b processing affected different stages of EAV reproduction. In particular, the mutant with the nsp10/11 cleavage site mutation Gln-2837-->Pro displayed an unusual phenotype, since it was still capable of RNA synthesis but was incapable of producing infectious virus.