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Analysis of Two Gene Clusters Involved in the Degradation of 4-Fluorophenol by Arthrobacter sp. Strain IF1

Ferreira, Maria Isabel M., Iida, Toshiya, Hasan, Syed A., Nakamura, Kaoru, Fraaije, Marco W., Janssen, Dick B., Kudo, Toshiaki
Applied and environmental microbiology 2009 v.75 no.24 pp. 7767-7773
Arthrobacter, DNA libraries, DNA primers, Escherichia coli, NAD (coenzyme), Northern blotting, adenine, carbon, clones, dehalogenation, energy, gene expression, hydroquinone, hydroxylation, multigene family, p-nitrophenol, reverse transcriptase polymerase chain reaction, screening
Arthrobacter sp. strain IF1 is able to grow on 4-fluorophenol (4-FP) as a sole source of carbon and energy. To clone the 4-FP degradation genes, DNA libraries were constructed and screened with a probe obtained by PCR using primers designed on the basis of conserved regions of aromatic two-component monooxygenases. Sequencing of positive clones yielded two gene clusters, each harboring a gene encoding a monooxygenase with high sequence similarity to the oxygenase component of 4-nitrophenol and 4-chlorophenol monooxygenase systems. Both these monooxygenase genes were differentially expressed during growth on 4-FP, as revealed by Northern blotting and reverse transcription-PCR. One cluster also contained a gene for a flavin reductase. The monooxygenase and reductase were purified from Escherichia coli cells expressing the corresponding genes, and together they catalyzed NADH-dependent hydroxylation and dehalogenation of 4-halophenols. The results indicate that strain IF1 transforms 4-FP to hydroquinone by a two-component monooxygenase system of which one component provides reduced flavin adenine dinucleotide at the expense of NADH and the other catalyzes para-hydroxylation of 4-FP and other 4-substituted phenols.