Project description:In order to genetically determine the essential components of electron flow during respiration of sulfate by the anaerobe Desulfovibrio desulfuricans G20, a growth mode independent of sulfate is needed. However, a mutant of the G20 strain, I2, lacking the abundant type-1 tetraheme cytochrome c3 (TpI-c3), was found to be altered in growth with sulfate. Here we report that I2 does not reduce sulfate with electrons from pyruvate. In contrast, in the absence of sulfate, fermentative growth of I2 on pyruvate actually yielded slightly greater cellular protein than G20. When provided lactate with sulfate or when fermenting pyruvate, I2 produced more hydrogen and accumulated little if any succinate. G20 grows by fumarate dismutation and accumulates the theoretical 2:1 ratio of the end products, succinate and acetate, respectively. In contrast, I2 did not grow on fumarate alone. We inferred that TpI-c3 might be required for transfer of electrons to the membrane-bound fumarate reductase or for establishing a redox environment needed to trigger synthesis of the enzymes for succinate production. Microarray and proteomic analyses of gene and protein expression in I2 compared with G20 were consistent with increases in those enzymes predicted to be necessary for the generation of end products measured. These results support a role for the periplasmic TpI-c3 in electron flow from pyruvate to sulfate and in establishing conditions for enzyme production for fumarate dismutation. In addition, evidence for the simultaneous operation of respiratory electron flow and substrate-level phosphorylation was obtained from end product analysis. Strains I2 and G20 of Desulfovibrio desulfuricans were grown in different media. Four biological replicates of G20 on lactate media, and two biological replicates of G20 on Pyruvate media were compared with three replicates of I2 on lactate media and two replicates of I2 on pyruvate media.
