The role of the Sphingomonas species UG30 pentachlorophenol-4-monooxygenase in p-nitrophenol degradation

Pentachlorophenol-4-monooxygenase is an aromatic flavoprotein monooxygenase which hydroxylates pentachlorophenol and a wide range of polyhalogenated p henols at their pare position. The PCP-degrading Sphingomonas species UG30 was recently shown to mineralize p-nitrophenol. In this study, the UG30 pcp B gene encoding the pentachlorophenol-4-monooxygenase gene was cloned for u se to study its potential role in p-nitrophenol degradation. The UG30 pcpB gene consists of 1614 bp with a predicted translational product of 538 amin o acids and a molecular mass of 59 933 Da. The primary sequence of pentachl orophenol-4-monooxygenase contained a highly conserved FAD binding site at its N-terminus associated with a beta alpha beta fold. UG30 has been shown previously to convert p-nitrophenol to 4-nitrocatechol. We observed that pe ntachlorophenol-4-monooxygenase catalyzed the hydroxylation of 4-nitrocatec hol to 1,2,4-benzenetriol. About 31.2% of the nitro substituent of 4-nitroc atechol (initial concentration of 200 mu M) was cleaved to yield nitrite ov er 2 h, indicating that the enzyme may be involved in the second step of p- nitrophenol degradation. The enzyme also hydroxylated p-nitrophenol at the para position, but only to a very slight extent. Our results confirm that p entachlorophenol-4-monooxygenase is not the primary enzyme in the initial s tep of p-nitrophenol metabolism by UG30. The UG30 pcpB sequence has been de posited at the GenBank under accession number AF059680. (C) 1999 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.