Molecular characterization of azoreductase and its potential for the decolorization of Remazol Red R and Acid Blue 29

Abstract

Azoreductase is a reductive enzyme that efficiently biotransformed textile azo dyes. This study demonstrated the heterologous overexpression of the azoreductase gene in Escherichia coli for the effective degradation of Remazol Red-R and Acid-Blue 29 dyes. The AzK gene of Klebsiella pneumoniae encoding a ≈22 kDa azoreductase enzyme was cloned into the pET21⁺C expression vector. The inoculum size of 1.5%, IPTG concentration of 0.5 mM, and incubation time of 6 h were optimized by response surface methodology a statistical tool. The crude extract showed 76% and 74%, while the purified enzyme achieved 94% and 93% decolorization of RRR and AB-29, respectively in 0.3 h. The reaction kinetics showed that RRR had a K_m and V_max value of 0.058 mM and 1416 U mg^-1, respectively at an NADH concentration of 10 mM. HPLC and GC-MS analyses showed that RRR was effectively bio-transformed by azoreductase to 2-[3-(hydroxy-amino) benzene-1-sulfonyl and AB-29 to aniline and 3-nitrosoaniline. This study explored the potential of recombinant azoreductase isolated from K. pneumoniae in the degradation of toxic textile azo dyes into less toxic metabolites.

Keywords: Azoreductase; Biodegradation; Bioremediation; Decolorization; Remazol red.