Metabolism Mechanism Of Phenol Degradtion In Acinetobacter Radioresistens APH1

As a crucial organic raw chemical,phenol is widely used in industrial manufacturing,pharmaceutical synthesis,and agricultural production.Thereby,phenol occupies a large proportion of the industrial wastewater,which is discharged from chemical plants.While flowing into the environment,the untreated wastewater poses a serious threat to the environment and human health.The accumulated phenol in the human body does harm to organs and tissues,interferes the nervous system,and even causes poisoning or death.Compared with physical and chemical detoxification methods,which have low efficiency and bring secondary pollution,using microorganisms screened from the actual polluted environment to repair phenol pollution is of great significance for environmental protection.In this study,an Acinetobacter radioresistens strain APH1 was isolated and identified for its powerful capability of utilizing phenol as the sole carbon source to grow.The optimal conditions for cell culture and phenol removal were determined to be 30°C,p H 6.0,0.2% Na Cl,and a phenol concentration of 500 mg/L.Under the optimal growth degradation conditions,500 mg/L phenol can be completely removed within 18 h.A draft genome sequence containing 3,290,330 bases with 45 contigs was obtained,and 22 genes were found to be involved in phenol metabolism and 51 putative drug resistance genes were annotated by genomic analysis.Based on GC-MS analysis of the key metabolites,and combined with the annotation of functional enzymes,the phenol metabolism pathway in the strain was deduced as follows: First,phenol was oxidized to catechol,then the catechol was further metabolized to form cis,cis-muconic acid by ringopening cleavage.Finally,through a series of reactions of β-ketoadipate pathway,succinic acid was produced and eventually flowed into the TCA cycle,achieving complete mineralization of phenol.Three catechol 1,2-dioxygenases,Cat Ap1,Cat Ap2,and Cat Ap3,were heterologously expressed and characterized.Cat Ap2,and Cat Ap3 can catalyze catechol to cis,cis-muconic acid,whereas Cat Ap1 has no activity.During bioremediation experiment using 450 mg/kg(dry weight)of phenol-contaminated soil,the strain APH1 removed 99% of the phenol within 3 days.According to microbial diversity analysis,the microbial abundance of Chungangia,Bacillus,Nitrospira,Lysinibacillus and Planomicrobium increased after the addition of phenol.Furthermore,at day23,the abundance of strain APH1 was substantially reduced,and the microbial diversity and structure of the whole microbial community were gradually recovered,indicating that strain APH1 would not affect the microbial ecosystem after the bioremediation.These findings provide a theoretical basis on metabolic mechanism of APH1 and microcosmic insights into the bioremediation of soil contaminated with phenol.