Characterization Of Polycylic Aromatic Compounds Degradation By Pseudomonas Putida

Polychlorinated biphenyls(PCBs),polybrominated diphenyl ethers(PBDEs), polycyclic aromatic hydrocarbons(PAHs) and polychlorinated dibenzo-p-dioxin (PCDDs) are environmental persistent organic pollutants of environments.These compounds are usually carcinogenic,teratogenic and mutagenic.The release of these compounds into the environments will be serious threats to human health,so studies on microbial degradation of these compounds have very important theoretical meanings and application values for remediation of the contaminated environments.This work focused on a biphenyl-degrading strain B6-2 which was isolated in our laboratory previously and was assigned to Pseudomonas putida.Strain B6-2 was able to grow with biphenyl or benzoate acid as the sole carbon source,indicating that strain B6-2 could completely mineralized biphenyl.Although could not grow on DE,fluorene(FN) and DD,biphenyl-grown cells of P. putida B6-2 were shown to be capable of efficiently degrading these three models compounds of PBDEs,PAHs and PCDDs.Identification of metabolites suggested that strain B6-2 degraded DE to phenol through the lateral dioxygenation pathway, and the production of hydroquinone was also detected.During the degradation of FN,two metabolites,9-fluorenol and 9-fluorenone,were found to accumulate. Detection of another metabolite 1-indanone indicated that strain B6-2 could also degrade FN through the lateral dioxygenation pathway.DD was degraded to 4-(3-hydroxybenzol[1,4]dioxin-2-yl)-2-oxobut-3-enoic acid through the lateral dioxygenation pathway by strain B6-2,and the further degradation product catechol was also identified.The above results indicate that strain P.putida B6-2 degrades DE,FN and DD mainly through lateral dioxygenation pathway.The metabolites, phenol and catechol,were detected,and were shown to be capable of support B6-2 cell growth.Therefore,at least partile of DE,FN and DD could be minerilized throught a cometabolic way by strain B6-2.A genome library of P.putida B6-2 was constructed for cloning function gene cluster.Two clusters corresponding to the biphenyl-degrading and salicylic acid-degrading were cloned.In strain B6-2,the arrangement of the the "biphenyl-degrading" gene cluster was the same as those in strain Burkholderia sp. LB400 and Pseudomonas pseudoalcaligenes KF707.The "biphenyl-degrading" gene cluster contains eight genes named bphABCKHJID,bphABCD encoding enzymes catalyzing biphenyl to benzoic acid and 2-hydroxypenta-2,4-dienoate. bphKHJI is corresponding for the degradation of 2-hydroxypenta-2,4-dienoate. Salicylic acid degrading gene cluster of strain B6-2 was assigned nah.In strain KF707,the nah was 6-kilo base downstream from bphD.The amino acid sequence of bphABCKHJID in strain B6-2 is highly similar with the corresponding enzymes in strain KF707.However,there are difference between the sequences of several amino acid groups of biphenyl dioxygenase genes(bphA) in these two strains,which should be the reason for their differences in catabolic activities.Because of the extensive degradation ablity of Burkholderia sp.LB400 towards PCBs,the crucial biphenyl dioxygenase gene(bphA) for PCBs degradation of this strain was introduced into P.putida B6-2.However,the enzymatic activity was not detected,there was no difference between the abilities to degrade 2,2',3,3'-tetrachloro biphenyl by P.putida B6-2 containing bphA of strain LB400 and the wild strain of B6-2.