PMID:22012340

Citation	Porter, AW, Campbell, BR, Kolvenbach, BA, Corvini, PF, Benndorf, D, Rivera-Cancel, G and Hay, AG (2012) Identification of the flavin monooxygenase responsible for ipso substitution of alkyl and alkoxyphenols in Sphingomonas sp. TTNP3 and Sphingobium xenophagum Bayram. Appl. Microbiol. Biotechnol. 94:261-72
Abstract	We previously showed that opdA from Sphingomonas sp. PWE1 encodes a putative flavin monooxygenase capable of transforming octylphenol (OP) via type II ipso substitution. Here, we demonstrate that an opdA homolog is responsible for OP and related alkyl/alkoxyphenol degradation in the nonylphenol degrader Sphingomonas sp. TTNP3. PCR and Southern blot analyses revealed that TTNP3 contained an opdA homolog, while a TTNP3 derivative unable to grow on nonylphenol (TTNP3d) did not. OpdA expression was confirmed in wild-type TTNP3 via two dimensional gel electrophoresis. Activity was restored to TTNP3d following complementation with opdA. Sequence analysis of an opdA homolog from another nonylphenol degrader, Sphingobium xenophagum Bayram, revealed that the predicted protein sequences from PWE1 and Bayram were identical, but differed from TTNP3 by four amino acids. In order to assess differences, we heterologously expressed the two unique opdA homologs and compared their effect on the disappearance of five alkyl/alkoxyphenol substrates and subsequent appearance of hydroquinone. For all substrates, except OP, the levels of substrate disappearance and hydroquinone appearance were significantly lower in cultures expressing odpA (TTNP3) than those expressing opdA (PWE1/Bayram). These differences in substrate specificity were consistent with an in silico model which predicted that two of the amino acid differences between odpA (TTNP3) and opdA (PWE1/Bayram) lay in a putative substrate binding pocket. While these strains are known to use the same type II ipso substitution mechanism for alkylphenol degradation, this work provides the first preliminary evidence that opdA homologs also encode the type I ipso substitution activity responsible for the degradation of alkoxyphenols.
Links	PubMed Online version:10.1007/s00253-011-3621-8
Keywords	Bacterial Proteins/genetics; Bacterial Proteins/metabolism; Biodegradation, Environmental; Mixed Function Oxygenases/genetics; Mixed Function Oxygenases/metabolism; Molecular Sequence Data; Molecular Structure; Phenols/chemistry; Phenols/metabolism; Sphingomonadaceae/enzymology; Sphingomonadaceae/genetics; Sphingomonadaceae/metabolism; Sphingomonas/enzymology; Sphingomonas/genetics; Sphingomonas/metabolism
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
9SPHN:A7YJX0		GO:0019336: phenol-containing compound catabolic process	ECO:0000315:		P		Fig. 5 showing the reduction in phenol between wt and mutant strains	complete CACAO 8556
9SPHN:A7YJX0	involved_in	GO:0019336: phenol-containing compound catabolic process	ECO:0000315: mutant phenotype evidence used in manual assertion		P		Seeded From UniProt	complete

References

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