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, 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.