Kinetic coefficients for simultaneous reduction of sulfate and uranium by Desulfovibrio desulfuricans

 Previously it was demonstrated that bacteria are capable of transforming soluble uranyl ion, U(VI), to insoluble uraninite, U(IV); however, the rate for this transformation has not been determined. We report the kinetic coefficients for Desulfovibrio desulfuricans DSM 1924 grown in a continuous-flow chemostat where pyruvate was the electron donor and sulfate was the electron acceptor. The medium was supplemented with 1 mM uranyl nitrate, and the chemostat flow rate ranged from 1.12 ml/h to 4.75 ml/h with incubation at 28°C. The maximum rate of pyruvate utilization (k) was determined to be 4.7 days^-1, while the half-velocity constant (K _s) was 127 mg/l. The yield coefficient (Y) of cells per mole of pyruvate oxidized was calculated to be 0.021 g, while the endogenous decay coefficient (k _d) was determined to be 0.072 days^-1. More than 90% of U(VI) was transformed to U(VI) in the chemostat under the conditions employed. Received: 7 September 1995/Received last revision: 10 January 1996/Accepted: 5 February 1996     

Effects of temperature and phosphorous concentration on microbial sulfate reduction by Desulfovibrio desulfuricans

The effects of temperature and phosphorous concentration on the rate and the extent of microbial sulfate reduction with lactate as carbon and energy source were investigated for Desulfovibrio desulfuricans. The continuous culture experiments (chemostat) were conducted at pH 7.0 from 12 to 48 degrees C. The maximum specific growth rate (mu(max)) was relatively constant in the range 25 degrees C-43 degrees C and dramatically decreased outside this temperature range. The half-saturation coefficient was minimum at 25 degrees C. Cell yield was highest in the optimum temperature range (35 degrees C-43 degrees C) for growth. Maintenance energy requirements for D. desulfuricans were not significant. Two moles of lactate is consumed for every mole of sulfate reduced, and this stoichiometric ratio is not temperature dependent. Steady state rate and stoichiometric coefficients accurately predicted transient behavior during temperature shifts. The extent of extracellular polymeric substance (EPS) is related to the concentration of phosphorous in the medium. EPS production rate increased with decreased phosphorous loading rate. Failure to discriminate between cell and EPS formation by D. desulfuricans leads to significant overestimates of the cell yield. The limiting C:P ratio for D. desulfuricans was in the range of 400:1 to 800:1.     

Comparative bioenergetics of sulfate reduction in Desulfovibrio and Desulfotomaculum spp.

Extracts of Desulfotomaculum nigrificans, Desulfotomaculum orientis, and Desulfotomaculum ruminis exhibit low levels of inorganic pyrophosphatase but were found to have high levels of pyrophosphate:acetate phosphotransferase. Conversely, extracts of Desulfovibrio gigas, Desulfovibrio vulgaris, and Desulfovibrio desulfuricans Norway 4 were shown to have high levels of inorganic pyrophosphatase but negligible amounts of pyrophosphate:acetate phosphotransferase. Both enzymes are reductant activated and appear to have an analogous function in removing pyrophosphate formed during the activation of sulfate. Conservation of the bond energy of pyrophosphate in Desulfotomaculum eliminates the necessity for invoking electron-transfer-coupled phosphorylation to account for the growth of these bacteria on lactate plus sulfate. Relative growth yields of Desulfovibrio vulgaris and Desulfotomaculum orientis on lactate plus sulfate indicate that the latter does not carry out significant electron-transfer-coupled phosphorylation in this mode of growth.     

Direct cathodic electron uptake coupled to sulfate reduction by Desulfovibrio ferrophilus IS5 biofilms

Direct electron uptake is emerging as a key process for electron transfer in anaerobic microbial communities, both between species and from extracellular sources, such as zero-valent iron (Fe⁰) or cathodic surfaces. In this study, we investigated cathodic electron uptake by Fe⁰-corroding Desulfovibrio ferrophilus IS5 and showed that electron uptake is dependent on direct cell contact via a biofilm on the cathode surface rather than through secreted intermediates. Induction of cathodic electron uptake by lactate-starved D. ferrophilus IS5 cells resulted in the expression of all components necessary for electron uptake; however, protein synthesis was required for full biofilm formation. Notably, proteinase K treatment uncoupled electron uptake from biofilm formation, likely through proteolytic degradation of proteinaceous components of the electron uptake machinery. We also showed that cathodic electron uptake is dependent on SO₄²⁻ reduction. The insensitivity of Fe⁰ corrosion to proteinase K treatment suggests that electron uptake from a cathode might involve different mechanism(s) than those involved in Fe⁰ corrosion.     

Enzymatic reduction of D-biotin-d-sulfoxide with cell-free extracts of Escherichia coli

D-biotin-d-sulfoxide can fulfill the growth requirement for $\text{E. coli}$ mutants deficient in biotin biosynthesis. In vitro reduction of D-biotin-d-sulfoxide to biotin requires a protein cell-free extract and NADPH. Preliminary experiments suggest that more than one protein participates in the reaction.     

Cysteine synthesis by Desulfovibrio vulgaris extracts.

Extracts of Desulfovibrio vulgaris were found to contain serine transacetylase and cysteine synthase activities. When extracts were incubated with bisulfite and o-acetylserine, or acetyl coenzyme A plus L-serine, under a hydrogen atmosphere, cysteine was formed. Pyruvate served as a reductant for bisulfite reduction to sulfide and concomitantly provided the acetyl moiety for acetyl coenzyme A formation. Consequently, when extracts were incubated with pyruvate, bisulfite, and L-serine, cysteine synthesis resulted.     

Periplasmic oxygen reduction by Desulfovibrio species

Washed cells of Desulfovibrio vulgaris strain Marburg (DSM 2119) reduced oxygen to water with H(2) as electron donor at a mean rate of 253 nmol O(2) min(-1) (mg protein)(-1). After separating the periplasm from the cells, more than 60% of the cytochrome c activity and 90% of the oxygen-reducing activity were found in the periplasmic fraction. Oxygen reduction and the reduction of cytochrome c with H(2) were inhibited by CuCl(2). After further separation of the periplasm by ultrafiltration (exclusion sizes 30, 50, and 100 kDa), oxygen reduction with H(2) occurred with the retentates only. Ascorbate plus tetramethyl-p-phenylenediamine (TMPD), however, were also oxidized by the filtrates. The stoichiometry of 1 mol O(2) reduced per 2 mol ascorbate oxidized indicated the formation of water. Our experiments present evidence that in D. vulgaris periplasmic hydrogenase and cytochrome c play a major role in oxygen reduction. Preliminary studies with other Desulfovibrio species indicated a similar function of periplasmic c-type cytochromes in D. desulfuricans CSN and D. termitidis KH1.     

Factors affecting microbial sulfate reduction by Desulfovibrio desulfuricans in continuous culture: limiting nutrients and sulfide concentration

The effects of sulfate and nitrogen concentrations of the rate and stoichiometry of microbial sulfate reduction were investigated for Desulfovibrio desulfuricans grown on lactate and sulfate in a chemostat at pH 7.0. Maximum specific growth rates (mu(max)), half-saturation coefficients (K(sul)), and cell yield (Y(c/Lac)) of 0.344 +/- 0.007 and 0.352 +/- 0.003 h (-1), 1.8 +/- 0.3 and 1.0 +/- 0.2 mg/L, and 0.020 +/- 0.003 and 0.017 +/- 0.003 g cell/g lactate, respectively, were obtained under sulfate-limiting conditions at 35 degrees C and 43 degrees C. Maintenance energy requirements for D. desulfuricans were significant under sulfate-limiting conditions. The extent of extracellular polymeric substance (EPS) produced was related to the carbon: nitrogen ratio in the medium. EPS production rate increased with decreased nitrogen loading rate. Nitrogen starvation also resulted in decreased cell size of D. desulfuricans. The limiting C : N ratio (w/w) for D. desulfuricans was in the range of 45 : 1 to 120 : 1. Effects of sulfide on microbial sulfate reduction, cell size, and biomass production were also ivestigated at pH 7.0. Fifty percent inhibition of lactate utilization occurred at a total sulfide concentration of approximately 500 mg/L. The cell size of D. desulfuricans decreased with increasing total sulfide concentration. Sulfide inhibition of D. desulfuricans was observed to be a reversible process. (c) 1992 John Wiley & Sons, Inc.