Dye-decolorizing peroxidase (DyP) with heme as a prosthetic group is regarded as a novel type of peroxidase subfamily. DyP has become a hot research topic of green economy because of its ability to degrade anthraquinone dyes. Since the effective degradation of dyes is closely related to enzyme activity, it is important to seek the ways of improving enzyme activity for the subsequent production and application of DyP. DypB gene from Rhodococcus jostii (RhDypB) was cloned to realize high-efficiency expression in Escherichia coli BL21(DE3), and to study methods for improving the enzyme activity. Recombinant strain BL21(DE3)/pET24b-DypB was constructed by one-step cloning technology. The recombinant protein was expressed by IPTG induction and purified by affinity chromatography using a nickel column. The effect on the activity of RhDypB by exogenous addition of heme synthesis precursor 5-aminolevulinic acid (5-ALA) and co-expression of the glutamyl-tRNA reductase gene (hemA) in the heme synthesis pathway in E. coli was compared. SDS-PAGE showed that there was an obvious target band at 40 kDa (purity-90%), and the concentration of protein was 100 mg/L. The addition of 5-ALA and co-expression of hemA increased the concentration of heme to 8.9 and 2.4 μmol/L, and the corresponding activity of enzyme was increased by 102% and 93%, respectively. The soluble expression of RhDypB and the increase in content of heme provide a basis for related research on peroxidase with heme as a cofactor.
PI Qian
,
XIA Rong
,
TANG Lei
. Heterologous expression and activity analysis of dye-decolorizing peroxidase from Rhodococcus jostii[J]. Food and Fermentation Industries, 2021
, 47(18)
: 86
-91
.
DOI: 10.13995/j.cnki.11-1802/ts.026822
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