Biochemical and genetic evidence of benzylsuccinate synthase in toluene-degrading, ferric iron-reducing Geobacter metallireducens

In vitro assays demonstrated that toluene-grown cells of Geobacter metallireducens catalyzed the addition of toluene to fumarate to form benzylsuccinate under anaerobic conditions. The specific in vitro rate of benzylsuccinate formation was ca. 45% of the specific in vivo rate of toluene consumption. In addition, bssA and bssB, which code for the alpha and beta subunits of benzylsuccinate synthase (BSS), respectively, were found to have sequences in G. metallireducens similar to the only sequences heretofore available (for three denitrifying strains). This is the first report of the presence of BSS in a ferric iron-reducing bacterium; BSS activity has previously been reported in denitrifying, sulfate-reducing, and anoxygenic phototrophic toluene degraders, as well as in a highly enriched methanogenic, toluene-degrading culture.