Physiological function of regulatory protein RHOGL007659 involved in metabolism biphenyl/polychlorinated biphenyl in Rhodococcus sp.R04

Abstract:

[Objective] The physiological function of regulatory protein RHOGL007659 in Rhodococcus sp. R04 and the metabolic properties of the mutant strain were investigated to explore its regulation mechanism of biphenyl degradation. [Methods] The RHOGL007659 gene was knocked out by homologous recombination, and the growth of the wild-type strain and deficient mutant strain in different carbon sources were compared. The transformation of biphenyl by the wild-type strain and the deficient mutant strain was detected by HPLC. Total RNAs were extracted from the wild-type strain and deficient mutant strain, and quantitative RT-PCR was carried out to analyze the transcriptional variation of bphA, bphB, bphC and bphD between the wild-type strain and the deficient mutant strain. BphB and BphD were expressed in Escherichia coli BL 21(DE3) and polyclonal antibodies were prepared after purification. BphB and BphD expression levels in the wild strain and deficient mutant strains were detected by Western blot. [Results] The deficient mutant strain R04Δ7659 of RHOGL007659 gene was obtained. The biomass of the deficient mutant strain significantly reduced in biphenyl culture, and almost did not grow. HPLC analysis showed that the deletion of RHOGL007659 gene resulted in inability to transform biphenyl by Rhodococcus sp. R04. Q-RT PCR experiment disclosed that the key genes of biphenyl degradation were down regulated in different degrees under biphenyl culture condition in the RHOGL007659 deficient mutant strain, the same results were obtained by Western blot. [Conclusion] RHOGL007659 is a regulatory protein of the upper biphenyl degradation gene in Rhodococcus sp. R04, and functions as a positive regulatory factor of the metabolism of biphenyl by Rhodococcus sp. R04.