Identification of the amino acid residue involved in rabbit hemorrhagic disease virus VPg uridylylation
Identification of the amino acid residue involved in rabbit hemorrhagic disease virus VPg uridylylation
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DOI：
10.1074/jbc.m100707200
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发表时间：
2001-07-27
影响因子：
4.8
通讯作者：
Parra, F
中科院分区：
文献类型：
--
作者：
Machín, A;Alonso, JMM;Parra, F
利用谷胱甘肽S-转移酶载体在大肠杆菌中表达兔出血症病毒（RHDV）AST/89株VPg基因。重组多肽可以以良好的产率纯化，并在体外由来自RHDV的重组RNA依赖性RNA聚合酶催化的反应中由[α-P-32]UTP在不存在添加的模板RNA的情况下进行尿苷酰化。使用缺失突变体和点突变体可以鉴定Tyr-21为参与尿苷酰化的残基，因此参与VPg和病毒基因组之间的连接。这些数据构成的第一份报告的身份参与VPg尿苷酰化的杯状病毒科的成员中的氨基酸残基。
The virus genome-linked protein (VPg) coding region from rabbit hemorrhagic disease virus (RHDV) (isolate AST/89) was expressed in Escherichia coli by using a glutathione S-transferase-based vector. The recombinant polypeptide could be purified in good yields and was uridylylated in vitro from [alpha-P-32]UTP in a reaction catalyzed by the recombinant RNA-dependent RNA polymerase from RHDV in the absence of added template RNA. The use of deletion and point mutants allowed the identification of Tyr-21 as the residue involved in uridylylation and consequently in the linkage between VPg and the viral genome. These data constitute the first report on the identity of the amino acid residue involved in VPg uridylylation in a member of the Caliciviridae family.