Immobilization of Rhodococcus opacus 1CP azoreductase to obtain azo dye degrading biocatalysts operative at acidic pH

Abstract

A novel FMN-containing oxygen-insensitive azoreductase designated as AzoRo from strain Rhodococcus opacus 1CP was recently reported to enable higher dye degradation rate between pH 3.8 and 5, but its stability was poor. Attempts to improve it by making cross-linked enzyme aggregates and crystals were not successful. Thus covalent immobilization of AzoRo on modified mesoporous silicas was carried out using two types of mesoporous silica carriers with different pore structure: SBA-15 (ordered mesoporous silica) and meso-cellular foams (MCF). The supports were functionalized with amino and epoxy groups. Immobilized AzoRo (7.35 U mg−1) showed almost the same preference for pH and temperature as free enzyme (16.89 U mg−1) but stability was significantly improved. The immobilized enzyme was active after incubation at pH of 4.0 for 60 h, its storability was also improved and maximum activity was 5.2-fold higher than at standard conditions and even over 2.2 times higher than of free enzyme at pH 6.0. Compared to SBA-15, MCF catalysts showed higher activity probably due to larger pores and more open structure. Overall, AzoRo immobilization on mesoporous silicas appeared to be a successful strategy to obtain biocatalysts degrading azo dyes at lower pH up to 38.24 U mg−1 and over a longer period.