Lawrence Berkeley National Laboratory

The anaerobic sulfate-reducing bacteria (SRB) of the genus Desulfovibrio are found in a remarkable variety of habitats, including soil, fresh water and salt water environments. SRB metabolism allows them to immobilize heavy toxic metals, such as uranium, through sulfide precipitation and/or through changing the redox state of the metal and thus its solubility. In studying the redox products of Desulfovibrio G20 during growth on various media, it was discovered a plasmid insertion mutant in the gene for the type-1 tetraheme cytochrome c3, cycA, was unable to grow on fumarate and the fumarate hydratase and fumarate reductase proteins were more than 10X decreased. Wildtype Desulfovibrio G20 anaerobic growth on fumarate appears to occur as a dismutation with the primary end products being succinate and acetate at approximately the theoretical ratio of 2:1, respectively. Yet wildtype growth with fumarate was inhibited with addition of as little as 5 mM formate. In wildtype G20, formate may inhibit growth directly by blocking an energy pathway or by down regulating the genes encoding the enzymes required for fumarate growth. Therefore the inability of the cycA mutant to grow on fumarate might result from an interruption in electron flow to the fumarate reductase or from an accumulation of inhibitory formate concentrations. To learn more about growth on fumarate, formate, or a combination of both, transposon mutants in formate dehydrogenases, formate C-acetyltransferases, malic enzymes, fumarate reductase, fumarate hydratase, and the pyruvate formate-lyase activating enzyme are being examined. Proteomic analysis of fumarate grown G20 cells, revealed that five of the eight proteins most increased in abundance compared to cells grown fermentatively on pyruvate, were specific to fumarate metabolism. Several of those highly expressed proteins are located in a single operon and include a fumarate reductase, fumarate hydratase and malic enzyme. To determine if the formate is having a regulatory effect on this operon, quantitative RT-PCR experiments are being performed to determine the expression of the genes in that operon