Promoting effect of the recombinant resuscitation promoting factors-2 of Rhodococcus erythropolis on petroleum degradation and cultivable bacterial diversities of the oil-contaminated soils.

Abstract

Resuscitation-promoting factors (Rpfs) belong to peptidoglycan hydrolases, which participate in recovery of dormant cells and promoting bacteria growth. In this study, the resuscitation promoting factor rpf2 gene of Rhodococcus erythropolis KB1 was expressed in Escherichia coli and purified by Ni2+ affinity chromatography. The purified recombinant fusion protein Rpf2 showed a closely 50 kDa band on sodium dodecyl sulphate polyacrylamide gel electrophoresis. The protein showed muralytic activity, with a specific activity of 1503 ± 123 U mg-1 when determined with 4-methylumbelliferyl-β-d-N, N',N″-triacetotri-ylchitoside as substrate. It also showed protease activity when measured with azocasein as substrate, with a specific activity of 1528 ± 411 U mg-1 . The addition of the recombinant Rpf2 protein significantly increased petroleum degradation efficiency of the indigenous micro-organisms and the petroleum degradation rates increased from 30·86 to 43·45%, 45·20 and 49·23% in the treatment groups. The recombinant protein also increased the petroleum-degrading bacterial diversities enriched from the contaminated soils. The cultivable bacterial flora of the treatment groups supplemented with different concentrations of Rpf2 increased from 82 genera in 9 phyla to 116 genera in 16 phyla and 138 genera in 16 phyla respectively. Thirteen extra petroleum-degrading bacteria strains were isolated from the petroleum-contaminated soils in the groups containing the recombinant Rpf2.