title: Direct over-expression, characterization and H2O2 stability study of active Pleurotus eryngii versatile peroxidase in Escherichia coli.
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 (A(407)/A(280), a measure of hemin content of the peroxidases) of 1.2, oxidized ABTS veratryl alcohol, Mn(2+), and Reactive Black 5. Activity of the enzyme increased after removing the tag. It lost only 5 % of its activity in 6.4 mM H(2)O(2). This is the first report on direct over-expression of active VP in E. coli.
authors.affiliation: Key Laboratory of Biofuels, Qingdao Institute of BioEnergy and Bioprocess Technology, Chinese Academy of Sciences, No. 189 Songling Road, Qingdao, 266101, China. baoxue@qibebt.ac.cn
authors.forename: Xue
authors.initials: X
authors.lastname: Bao
authors.forename: Aiqiu
authors.initials: A
authors.lastname: Liu
authors.forename: Xuefeng
authors.initials: X
authors.lastname: Lu
authors.forename: Jian-Jun
authors.initials: JJ
authors.lastname: Li
mesh_terms: D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000737:chemistry*; D010544:Peroxidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020076:Pleurotus / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D005656:Fungal Proteins; D011994:Recombinant Proteins; D006861:Hydrogen Peroxide; D010544:Peroxidases