PMID:19251893

Citation	Shin, KA and Spain, JC (2009) Pathway and evolutionary implications of diphenylamine biodegradation by Burkholderia sp. strain JS667. Appl. Environ. Microbiol. 75:2694-704
Abstract	Diphenylamine (DPA) is a common contaminant at munitions-contaminated sites as well as at aniline manufacturing sites. Little is known about the biodegradation of the compound, and bacteria able to use DPA as the growth substrate have not been reported. Burkholderia sp. strain JS667 and Ralstonia sp. strain JS668 were isolated by selective enrichment from DPA-contaminated sediment. The isolates grew aerobically with DPA as the sole carbon, nitrogen, and energy source. During induction of DPA degradation, stoichiometric amounts of aniline accumulated and then disappeared, which suggested that aniline is on the DPA degradation pathway. Genes encoding the enzymes that catalyze the initial steps in DPA degradation were cloned from the genomic DNA of strain JS667. The Escherichia coli clone catalyzed stoichiometric transformation of DPA to aniline and catechol. Transposon mutagenesis, the sequence similarity of putative open reading frames to those of well-characterized dioxygenases, and (18)O(2) experiments support the conclusion that the initial reaction in DPA degradation is catalyzed by a multicomponent ring-hydroxylating dioxygenase. DPA is converted to aniline and catechol via dioxygenation at the 1,2 position of the aromatic ring and spontaneous rearomatization. Aniline and catechol are further biodegraded by the well-established aniline degradation pathway. Genes that encode the complete aniline degradation pathway were found 12 kb downstream of the genes that encode the initial dioxygenase. Expression of the relevant dioxygenases was confirmed by reverse transcription-PCR analysis. Both the sequence similarity and the gene organization suggest that the DPA degradation pathway evolved recently by the recruitment of two gene clusters that encode the DPA dioxygenase and aniline degradation pathway.
Links	PubMed PMC2681709 Online version:10.1128/AEM.02198-08
Keywords	Aerobiosis; Aniline Compounds/metabolism; Burkholderia/genetics; Burkholderia/growth & development; Burkholderia/isolation & purification; Burkholderia/metabolism; Catechols/metabolism; Cloning, Molecular; DNA, Bacterial/chemistry; DNA, Bacterial/genetics; Dioxygenases/metabolism; Diphenylamine/metabolism; Escherichia coli/genetics; Evolution, Molecular; Gene Expression Profiling; Gene Order; Metabolic Networks and Pathways/genetics; Molecular Sequence Data; Mutagenesis, Insertional; Oxygen Isotopes/metabolism; Ralstonia/isolation & purification; Sequence Analysis, DNA; Sequence Homology, Amino Acid
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
9BURK:C3UVB8		GO:0051213: dioxygenase activity	ECO:0000314:		F		Fig 4. shows how DPA in enzyme forms stoichiometric amounts of the expected products. Fig 5. shows that the enzyme inserts both atoms of molecular oxygen into DPA, proving that it has dioxygenase activity	complete CACAO 4746

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