A novel mutation in murine hepatitis virus nsp5, the viral 3C-like proteinase, causes temperature-sensitive defects in viral growth and protein processing.
A novel mutation in murine hepatitis virus nsp5, the viral 3C-like proteinase, causes temperature-sensitive defects in viral growth and protein processing.
复制标题
鼠肝炎病毒 nsp5（病毒 3C 样蛋白酶）的一种新突变会导致病毒生长和蛋白质加工出现温度敏感缺陷。
DOI：
10.1128/jvi.00203-08
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发表时间：
2008
影响因子：
5.4
通讯作者：
Denison,MarkR
中科院分区：
文献类型：
--
作者：
Sparks,JenniferS;Donaldson,EricF;Lu,Xiaotao;Baric,RalphS;Denison,MarkR
Sequencing and reversion analysis of murine hepatitis virus (MHV) temperature-sensitive (ts) viruses has identified putativetsmutations in the replicase nonstructural proteins (nsp's) of these coronaviruses. In this study, reverse transcriptase PCR sequencing of the RNA genome of an isolate of the MHVtsvirus Albts6, referred to as Alb/ts/nsp5/V148A, identified a putativetsmutation in nsp5 (T10651C, Val148Ala), the viral 3C-like proteinase (3CLpro). The introduction of the T10651C mutation into the infectious MHV clone resulted in the recovery of a mutant virus, the nsp5/V148A virus, that demonstrated reduced growth and nsp5 proteinase activity identical to that of Alb/ts/nsp5/V148A at the nonpermissive temperature. Sequence analysis of 40°C revertants of Alb/ts/nsp5/V148A identified primary reversion to Ala148Val in nsp5, as well as two independent second-site mutations resulting in Ser133Asn and His134Tyr substitutions in nsp5. The introduction of the Ser133Asn or His134Tyr substitution into the cloned nsp5/V148A mutant virus background resulted in the recovery of viruses with increased growth fitness and the partial restoration of nsp5 activity at the nonpermissive temperature. Modeling of the nsp5 structure of Alb/ts/nsp5/V148A predicted that the Val148Ala mutation alters residue 148 interactions with residues of the substrate binding S1 subsite of the nsp5 active-site cavity. This study identifies novel residues in nsp5 that may be important for regulating substrate specificity and nsp5 proteinase activity.
