Abstract
The vpl2 gene, encoding versatile peroxidase (VP) from Pleurotus eryngii, was synthesized with codon optimization and cloned into vector-pET-32a(+) and over-expressed in Escherichia coli BL21(DE3). An active peroxidase fused to the thioredoxin–hexahistidine tag was directly obtained by IPTG induction in the presence of hemin. Most of over-expressed protein was in the soluble form, and was purified on a nickel column with >85 % purity at a yield of 12.5 mg/l. The purified fusion protein, having an Rz value (A407/A280, a measure of hemin content of the peroxidases) of 1.2, oxidized ABTS veratryl alcohol, Mn2+, and Reactive Black 5. Activity of the enzyme increased after removing the tag. It lost only 5 % of its activity in 6.4 mM H2O2. This is the first report on direct over-expression of active VP in E. coli.
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Acknowledgments
We are grateful to Knowledge Innovation Program of the Chinese Academy of Sciences (Grant KSCX1-YW-11A3) and Natural Science Foundation of Shandong Province (China) (Grant ZR2010CM058) for the financial support.
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Bao, X., Liu, A., Lu, X. et al. Direct over-expression, characterization and H2O2 stability study of active Pleurotus eryngii versatile peroxidase in Escherichia coli . Biotechnol Lett 34, 1537–1543 (2012). https://doi.org/10.1007/s10529-012-0940-5
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DOI: https://doi.org/10.1007/s10529-012-0940-5