Two W-containing formate dehydrogenases (CO2-reductases) involved in syntrophic propionate oxidation by Syntrophobacter fumaroxidans

Two formate dehydrogenases (CO2-reductases) (FDH-1 and FDH-2) were isolated from the syntrophic propionate-oxidizing bacterium Syntrophobacter fumaroxidans. Both enzymes were produced in axenic fumarate-grown cells as well as in cells which were grown syntrophically on propionate with Methanospirillum hungatei as the H-2 and formate scavenger. The purified enzymes exhibited extremely high formate-oxidation and CO2-reduction rates, and low K-m values for formate. For the enzyme designated FDH-1, a specific formate oxidation rate of 700 U.mg(-1) and a K-m for formate of 0.04 mm were measured when benzyl viologen was used as an artificial electron acceptor. The enzyme designated FDH-2 oxidized formate with a specific activity of 2700 U.mg(-1) and a K-m of 0.01 mm for formate with benzyl viologen as electron acceptor. The specific CO2 -reduction (to formate) rates measured for FDH-1 and FDH-2, using dithionite-reduced methyl viologen as the electron donor, were 900 U.mg(-1) and 89 U.mg(-1) , respectively. From gel filtration and polyacrylamide gel electrophoresis it was concluded that FDH-1 is composed of three subunits (89+/-3, 56+/-2 and 19+/-1 kDa) and has a native molecular mass of approximately 350 kDa. FDH-2 appeared to be a heterodimer composed of a 92+/-3 kDa and a 33+/-2 kDa subunit. Both enzymes contained tungsten and selenium, while molybdenum was not detected. EPR spectroscopy suggested that FDH-1 contains at least four [2Fe-2S] clusters per molecule and additionally paramagnetically coupled [4Fe-4S] clusters. FDH-2 contains at least two [4Fe-4S] clusters per molecule. As both enzymes are produced under all growth conditions tested, but with differences in levels, expression may depend on unknown parameters.