This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Nam, J.-W.
Right arrow Articles by Omori, T.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Nam, J.-W.
Right arrow Articles by Omori, T.
Agricola
Right arrow Articles by Nam, J.-W.
Right arrow Articles by Omori, T.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, December 2002, p. 5882-5890, Vol. 68, No. 12
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.12.5882-5890.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Purification and Characterization of Carbazole 1,9a-Dioxygenase, a Three-Component Dioxygenase System of Pseudomonas resinovorans Strain CA10

Jeong-Won Nam, Hideaki Nojiri, Haruko Noguchi, Hiromasa Uchimura, Takako Yoshida, Hiroshi Habe, Hisakazu Yamane, and Toshio Omori*

Biotechnology Research Center, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan

Received 22 March 2002/ Accepted 13 September 2002

The carbazole 1,9a-dioxygenase (CARDO) system of Pseudomonas resinovorans strain CA10 consists of terminal oxygenase (CarAa), ferredoxin (CarAc), and ferredoxin reductase (CarAd). Each component of CARDO was expressed in Escherichia coli strain BL21(DE3) as a native form (CarAa) or a His-tagged form (CarAc and CarAd) and was purified to apparent homogeneity. CarAa was found to be trimeric and to have one Rieske type [2Fe-2S] cluster and one mononuclear iron center in each monomer. Both His-tagged proteins were found to be monomeric and to contain the prosthetic groups predicted from the deduced amino acid sequence (His-tagged CarAd, one FAD and one [2Fe-2S] cluster per monomer protein; His-tagged CarAc, one Rieske type [2Fe-2S] cluster per monomer protein). Both NADH and NADPH were effective as electron donors for His-tagged CarAd. However, since the kcat/Km for NADH is 22.3-fold higher than that for NADPH in the 2,6-dichlorophenolindophenol reductase assay, NADH was supposed to be the physiological electron donor of CarAd. In the presence of NADH, His-tagged CarAc was reduced by His-tagged CarAd. Similarly, CarAa was reduced by His-tagged CarAc, His-tagged CarAd, and NADH. The three purified proteins could reconstitute the CARDO activity in vitro. In the reconstituted CARDO system, His-tagged CarAc seemed to be indispensable for electron transport, while His-tagged CarAd could be replaced by some unrelated reductases.

* Corresponding author. Mailing address: Biotechnology Research Center, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Phone: 81 (3) 5841-3067. Fax: 81 (3) 5841-8030. E-mail: aseigyo{at}mail.ecc.u-tokyo.ac.jp.

Applied and Environmental Microbiology, December 2002, p. 5882-5890, Vol. 68, No. 12
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.12.5882-5890.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Urata, M., Uchimura, H., Noguchi, H., Sakaguchi, T., Takemura, T., Eto, K., Habe, H., Omori, T., Yamane, H., Nojiri, H. (2006). Plasmid pCAR3 Contains Multiple Gene Sets Involved in the Conversion of Carbazole to Anthranilate.. Appl. Environ. Microbiol. 72: 3198-3205 [Abstract] [Full Text]  
  • Inoue, K., Habe, H., Yamane, H., Nojiri, H. (2006). Characterization of Novel Carbazole Catabolism Genes from Gram-Positive Carbazole Degrader Nocardioides aromaticivorans IC177.. Appl. Environ. Microbiol. 72: 3321-3329 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»