MOLECULAR-CLONING AND EXPRESSION ANALYSIS OF THE RHODOBACTER-CAPSULATUS SODB GENE, ENCODING AN IRON SUPEROXIDE-DISMUTASE

Genetic complementation of a sodA sodB Escherichia coli mutant strain was used to clone Rhodobacter capsulatus genes involved in detoxificat ion of superoxide radicals. After sequence analysis, 1 of the 16 ident ical clones obtained by this selection procedure was shown to contain an open reading frame with sequence similarity to that coding for Fe-c ontaining superoxide dismutases (SodB). The R. capsulatus sodB gene wa s expressed in E. call, and the nature of the metal ligand was confirm ed by inhibitor sensitivity assays with lysates from both bacterial sp ecies. Activity staining of cleared Rhodobacter lysates resolved by po lyacrylamide gel electrophoresis indicated that SodB was the only supe roxide dismutase present in this phototrophic organism. The sodB gene was expressed at low levels in R. capsulatus cells grown under anaerob ic or semiaerobic conditions, but expression was strongly induced upon exposure of the bacteria to air or to methyl viologen. Attempts to co nstruct a sodB mutant in this organism by allelic exchange of the chro mosomal copy of the gene with a suicide plasmid containing a mutated s odB gene were unsuccessful, strongly suggesting that the encoded super oxide dismutase is essential for viability of R. capsulatus in aerobic cultures.