Biochemical and kinetic analysis of the influenza virus RNA polymerase purified from insect cells

Biochemical and Biophysical Research Communications
Shijian ZhangTetsuya Toyoda

Abstract

The influenza virus RNA polymerase (RdRp) was purified from insect cells (around 0.2mg/l). The RdRp catalyzed all the biochemical reactions of influenza virus transcription and replication in vitro; dinucleotide ApG and globin mRNA-primed transcription, de novo initiation (replication), and polyadenylation. The optimal Mg concentration, pH and temperature were 8mM, 8.0 and 25 degrees C, respectively, which were slightly different from those measured for RdRp of virions. This system is a single-round transcription system. K(m) (microM) were 10.74+/-0.26 (GTP), 33.22+/-3.37 (ATP), 28.93+/-0.48 (CTP) and 22.01+/-1.48 (UTP), and V(max) (fmol nucleotide/pmol RdRp/min) were 2.40+/-0.032 (GTP), 1.95+/-0.17 (ATP), 2.07+/-0.17 (CTP), and 1.52+/-0.38 (UTP), which agreed with high mutation of influenza viruses.

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Citations

Sep 23, 2010·The Journal of Biological Chemistry·Shijian ZhangTetsuya Toyoda
Aug 13, 2010·Journal of Virology·Patricia Resa-InfanteJuan Ortín
Jan 31, 2012·Biochimica Et Biophysica Acta·Qiang WangTetsuya Toyoda
Jan 12, 2011·Biochimica Et Biophysica Acta·Hongbing JiangTetsuya Toyoda
Oct 30, 2012·Analytical Biochemistry·Qiang WangTetsuya Toyoda
Mar 14, 2013·Nature Communications·Lauren TurrellFrank T Vreede

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