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Journal of Bacteriology, October 1998, p. 5413-5420, Vol. 180, No. 20
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Molecular Cloning and Expression Analysis of the Rhodobacter capsulatus sodB Gene, Encoding an Iron Superoxide Dismutase

Néstor Cortez,1,* Néstor Carrillo,1 Cécile Pasternak,2 Angelika Balzer,2 and Gabriele Klug2

Programa Multidisciplinario de Biología Experimental (PROMUBIE), National University of Rosario and CONICET, RA2000 Rosario, Argentina,1 and Institut für Mikro und Molekularbiologie, Justus-Liebig Universität, D-35392 Giessen, Germany2

Received 24 March 1998/Accepted 12 August 1998

Genetic complementation of a sodA sodB Escherichia coli mutant strain was used to clone Rhodobacter capsulatus genes involved in detoxification of superoxide radicals. After sequence analysis, 1 of the 16 identical clones obtained by this selection procedure was shown to contain an open reading frame with sequence similarity to that coding for Fe-containing superoxide dismutases (SodB). The R. capsulatus sodB gene was expressed in E. coli, and the nature of the metal ligand was confirmed by inhibitor sensitivity assays with lysates from both bacterial species. Activity staining of cleared Rhodobacter lysates resolved by polyacrylamide gel electrophoresis indicated that SodB was the only superoxide dismutase present in this phototrophic organism. The sodB gene was expressed at low levels in R. capsulatus cells grown under anaerobic or semiaerobic conditions, but expression was strongly induced upon exposure of the bacteria to air or to methyl viologen. Attempts to construct a sodB mutant in this organism by allelic exchange of the chromosomal copy of the gene with a suicide plasmid containing a mutated sodB gene were unsuccessful, strongly suggesting that the encoded superoxide dismutase is essential for viability of R. capsulatus in aerobic cultures.

* Corresponding author. Mailing address: PROMUBIE, Area Biología Molecular, Facultad de Cs. Bioquímicas y Farmacéuticas, U.N.R., Suipacha 531, RA2000 Rosario, Argentina. Phone: (54) (41) 350661/350596. Fax: (54) (41) 390465. E-mail: cortez{at}arnet.com.ar.

Journal of Bacteriology, October 1998, p. 5413-5420, Vol. 180, No. 20
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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