Expression and characterization of RNA-dependent RNA polymerase of Ectropis obliqua virus.
Replication of positive-strand RNA virus is mediated by a virus- encoded RNA-dependent RNA polymerase (RdRp). To study the replication of Ectropis obliqua virus (EoV), a newly identified insect virus belonging to the family Iflaviradae, we expressed the RNA polymerase domain in Escherichia coli and purified it on a Ni-chelating HisTrap affinity column. It is demonstrated that EoV RdRp initiated RNA synthesis in a primerand poly (A)-dependent manner in vitro. Furthermore, the effect of primer concentration, temperature, metal ions (Mg2+, Mn2+, and K+) on enzymatic activity were determined. Our study represented a first step towards understanding the mechanism of EoV replication.
Lin
Meijuan
M
State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei Province 430072, China.
Ye
Shan
S
Xiong
Yi
Y
Cai
Dawei
D
Zhang
Jiamin
J
Hu
Yuanyang
Y
D000595:Amino Acid Sequence; D017931:DNA Primers / Q000737:chemistry / Q000378:metabolism; D007304:Insect Viruses / Q000145:classification / Q000201:enzymology*; D008670:Metals / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D012313:RNA / Q000096:biosynthesis / Q000378:metabolism; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013696:Temperature; D014779:Virus Replication
D017931:DNA Primers; D008670:Metals; D011994:Recombinant Proteins; D012313:RNA; D012324:RNA-Dependent RNA Polymerase