This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Muraki, T. Articles by Lau, P. C. K. Search for Related Content PubMed PubMed Citation Articles by Muraki, T. Articles by Lau, P. C. K. Agricola Articles by Muraki, T. Articles by Lau, P. C. K.

Prokaryotic Homologs of the Eukaryotic 3-Hydroxyanthranilate 3,4-Dioxygenase and 2-Amino-3-Carboxymuconate-6-Semialdehyde Decarboxylase in the 2-Nitrobenzoate Degradation Pathway of Pseudomonas fluorescens Strain KU-7

Department of Biotechnology, Faculty of Engineering and High Technology Research Center, Kansai University, Suita, Osaka 564-8680, Japan,¹ Biotechnology Research Institute, National Research Council of Canada, Montreal, Quebec H4P 2R2, Canada²

Received 12 August 2002/ Accepted 10 December 2002

The 2-nitrobenzoic acid degradation pathway of Pseudomonas fluorescens strain KU-7 proceeds via a novel 3-hydroxyanthranilate intermediate. In this study, we cloned and sequenced a 19-kb DNA locus of strain KU-7 that encompasses the 3-hydroxyanthranilate meta-cleavage pathway genes. The gene cluster, designated nbaEXHJIGFCDR, is organized tightly and in the same direction. The nbaC and nbaD gene products were found to be novel homologs of the eukaryotic 3-hydroxyanthranilate 3,4-dioxygenase and 2-amino-3-carboxymuconate-6-semialdehyde decarboxylase, respectively. The NbaC enzyme carries out the oxidation of 3-hydroxyanthranilate to 2-amino-3-carboxymuconate-6-semialdehyde, while the NbaD enzyme catalyzes the decarboxylation of the latter compound to 2-aminomuconate-6-semialdehyde. The NbaC and NbaD proteins were overexpressed in Escherichia coli and characterized. The substrate specificity of the 23.8-kDa NbaC protein was found to be restricted to 3-hydroxyanthranilate. In E. coli, this enzyme oxidizes 3-hydroxyanthranilate with a specific activity of 8 U/mg of protein. Site-directed mutagenesis experiments revealed the essential role of two conserved histidine residues (His52 and His96) in the NbaC sequence. The NbaC activity is also dependent on the presence of Fe²⁺ but is inhibited by other metal ions, such as Zn²⁺, Cu²⁺, and Cd²⁺. The NbaD protein was overproduced as a 38.7-kDa protein, and its specific activity towards 2-amino-3-carboxymuconate-6-semialdehyde was 195 U/mg of protein. Further processing of 2-aminomuconate-6-semialdehyde to pyruvic acid and acetyl coenzyme A was predicted to proceed via the activities of NbaE, NbaF, NbaG, NbaH, NbaI, and NbaJ. The predicted amino acid sequences of these proteins are highly homologous to those of the corresponding proteins involved in the metabolism of 2-aminophenol (e.g., AmnCDEFGH in Pseudomonas sp. strain AP-3). The NbaR-encoding gene is predicted to have a regulatory function of the LysR family type. The function of the product of the small open reading frame, NbaX, like the homologous sequences in the nitrobenzene or 2-aminophenol metabolic pathway, remains elusive.

This paper is issued as NRCC number 45897.

This article has been cited by other articles:

• Iwaki, H., Muraki, T., Ishihara, S., Hasegawa, Y., Rankin, K. N., Sulea, T., Boyd, J., Lau, P. C. K. (2007). Characterization of a Pseudomonad 2-Nitrobenzoate Nitroreductase and Its Catabolic Pathway-Associated 2-Hydroxylaminobenzoate Mutase and a Chemoreceptor Involved in 2-Nitrobenzoate Chemotaxis. J. Bacteriol. 189: 3502-3514 [Abstract] [Full Text]  
• Liu, L., Wu, J.-F., Ma, Y.-F., Wang, S.-Y., Zhao, G.-P., Liu, S.-J. (2007). A Novel Deaminase Involved in Chloronitrobenzene and Nitrobenzene Degradation with Comamonas sp. Strain CNB-1. J. Bacteriol. 189: 2677-2682 [Abstract] [Full Text]  
• Iwaki, H., Wang, S., Grosse, S., Bergeron, H., Nagahashi, A., Lertvorachon, J., Yang, J., Konishi, Y., Hasegawa, Y., Lau, P. C. K. (2006). Pseudomonad Cyclopentadecanone Monooxygenase Displaying an Uncommon Spectrum of Baeyer-Villiger Oxidations of Cyclic Ketones. Appl. Environ. Microbiol. 72: 2707-2720 [Abstract] [Full Text]  
• Li, X., Guo, M., Fan, J., Tang, W., Wang, D., Ge, H., Rong, H., Teng, M., Niu, L., Liu, Q., Hao, Q. (2006). Crystal structure of 3-hydroxyanthranilic acid 3,4-dioxygenase from Saccharomyces cerevisiae: A special subgroup of the type III extradiol dioxygenases. Protein Sci. 15: 761-773 [Abstract] [Full Text]  
• Wu, J.-f., Jiang, C.-y., Wang, B.-j., Ma, Y.-f., Liu, Z.-p., Liu, S.-j. (2006). Novel Partial Reductive Pathway for 4-Chloronitrobenzene and Nitrobenzene Degradation in Comamonas sp. Strain CNB-1.. Appl. Environ. Microbiol. 72: 1759-1765 [Abstract] [Full Text]  
• Gaillard, M., Vallaeys, T., Vorholter, F. J., Minoia, M., Werlen, C., Sentchilo, V., Puhler, A., van der Meer, J. R. (2006). The clc Element of Pseudomonas sp. Strain B13, a Genomic Island with Various Catabolic Properties.. J. Bacteriol. 188: 1999-2013 [Abstract] [Full Text]  
• Colabroy, K. L., Begley, T. P. (2005). Tryptophan Catabolism: Identification and Characterization of a New Degradative Pathway. J. Bacteriol. 187: 7866-7869 [Abstract] [Full Text]