Study On The Mechanism Of Nitrogen Degradation By Rhodococcus Sp. LMD As An Efficient RDX Degradading

Cyclic trimethylene ammonium trinitrate(RDX),an energetic compound,has a great effect on environmental pollution and ring breaking,and is difficult to degrade.To screen environmental microorganisms for RDX degradation,the laboratory has previously isolated and domesticated a Rhodococcus sp.LMD(CGMCC NO.24210)from sludge containing RDX,which is resistant to high concentrations of RDX wastewater.The RDX tolerance and metabolic mechanism of the strain were preliminarily investigated by proteomics and metabolomics.Target: Supersaturated RDX tolerance test,RDX degradation test by key enzymes in vitro and dynamic transcriptomics were used to study the RDX tolerance and degradation mechanism of the strain.Method: The effect of high RDX concentration on the target strains was studied by acetone supersaturated RDX wastewater.The degradation mechanism of RDX was verified by recombinant expression of key P450 enzymes and accessory proteins.The differential expression of key genes in key tolerance,degradation,and nitrogen metabolism pathways of the strains were analyzed by dynamic transcriptome analysis(the differential expression of key genes at different growth stages between RDX as the only nitrogen source and the control group).(1)Acetone co-solvent was used to prepare supersaturated RDX solution.Rhodococcus sp.LMD could tolerate 54 mg/L supersaturated RDX solution and grow with it as the sole nitrogen source,and the optimal tolerance concentration of RDX was 36 mg/L.(2)Two P450 enzymes(gene722 and gene939)from Rhodococcus sp.LMD and the control P450 enzyme Xpl A were successfully expressed and degradated by RDX in vitro.The RDX degradation efficiency of the control enzyme was better than that of gene722 and gene939.(3)The dynamic transcriptome study found that several differentially expressed genes were involved in RDX tolerance,degradation and transformation metabolism,mainly including transporters of nitrogen-containing organic matter,nitrogen-containing organic matter degrading enzymes(such as P450 enzymes),and the key metabolic enzymes of glutamate,glutamine,and methionine(which were found to be increased in RDX experimental group by metabolomics).Conclusion and perspective: This study demonstrated that Rhodococcus sp.LMD could tolerate supersaturated RDX wastewater.The recombinant key enzymes and in vitro enzyme degradation experiments confirmed that the target strain could express P450 enzymes that degrade RDX.Dynamic transcriptomics combined with metabolomics and proteomics data confirmed that RDX as the only nitrogen source had a great impact on the nitrogen metabolism pathway of the target strain,indicating that the strain had the ability to transform RDX into other high-value nitrogen-containing metabolites.Further studies will focus on the metabolism and regulation mechanism of nitrogen-containing organic matter in the target monarch.