Efficient degradation of hydroquinone by a metabolically engineered Pseudarthrobacter sulfonivorans strain

Pseudarthrobacter sulfonivorans strain Ar51 can degrade crude oil and multi-substituted benzene compounds efficiently at low temperatures. However, it cannot degrade hydroquinone, which is a key intermediate in the degradation of several other compounds of environmental importance, such as 4-nitrophenol, g-hexachlorocyclohexane, 4-hydroxyacetophenone and 4-aminophenol. Here we co-expressed the two subunits of hydroquinone dioxygenase from Sphingomonas sp . strain TTNP3 with different promoters in the strain Ar51. The strain with 2 hdnO promoters exhibited the strongest hydroquinone catabolic activity. However, in the absence of antibiotic selection this ability to degrade hydroquinone was lost due to plasmid instability. Consequently, we constructed a hisD knockout strain, which was unable to synthesise histidine. By introducing the hisD gene onto the plasmid, the ability to degrade hydroquinone in the absence of antibiotic selection was stabilised. In addition, to make the strain more stable for industrial applications, we knocked out the recA gene and integrated the hydroquinone dioxygenase genes at this chromosomal locus. This strain exhibited the strongest activity in catabolizing hydroquinone, up to 470 mg/L in 16h without antibiotic selection. Additionally, this activity was shown to be stable when the strain has cultured in medium without antibiotic selection after 20 passages.