Energetics of methanogenic benzoate degradation by Syntrophus gentianae in syntrophic coculture

Abstract

Summary: Growing cocultures of Syntrophus gentianae with Methanospirillum hungatei degraded benzoate to CH4 and acetate. During growth, the change of free energy available for Syntrophus gentianae ranged between -50 and -55 kJ mol−1. At the end-point of benzoate degradation, a residual concentration of benzoate of 0.2 mM was found, correlating with a free energy change of -45 kJ mol−1 available to the fermenting bacterium. Benzoate thresholds were also observed in dense cell suspensions. They corresponded 1 a final energy situation in the range -31.8 to -45.8 kJ mol−1 for the fermentin bacterium. Addition of a H2-oxidizing sulfate reducer to the methanogenic coculture inhibited by bromoethanesulfonate (BES) resulted in benzoate degradation to below the limit of benzoate detection (10 μM). Accumulated acetate proved to be thermodynamically inhibitory; removal of acetate by Methanosaeta concilii in methanogenic or molybdate-inhibited sulfate-reducing cocultures led to degradation of residual benzoate with a final δG’ -45.8 kJ mol−1. In methanogenic cocultures, the residual Gibbs free energy (δG’) available for the fermenting bacterium at the end of benzoate degradation correlated with the concentration of acetate built up during the course of benzoate degradation; higher concentrations led to more positive values for δG’. Addition of different concentrations of propionate resulted in different values for δG when benzoate degradation had ceased; higher concentrations led to more positive values for δG’. Addition of acetate or propionate to benzoate-degrading cocultures also lowered the rate of benzoate degradation. The protonophore carbonylcyanide chlorophenylhydrazone (CCCP) facilitated further benzoate degradation in methanogenic BES-inhibited cocultures until a δG’ of -31 kJ mol−1 was reache We conclude that the minimum energy required for growth and energy conservation of the benzoate-fermenting bacterium S. gentianae is approximately -45 kJ (mol benzoate)−1, equivalent to two-thirds of an ATP unit. Both hydrogen and acetate inhibit benzoate degradation thermodynamically, and acetate also partly uncouples substrate degradation from energy conservation.

Energetics and biochemistry of fermentative benzoate degradation by Syntrophus gentianae

“…With this, the overall energy balance of benzoate degradation by S. gentianae comes to a net energy gain of one-third of an ATP unit per reaction run provided that the dearomatizing benzoyl-CoA reduction does not require any energy input, as hypothesized above. This energy balance agrees with the overall energy balance calculated on the basis of substrate and product concentrations in metabolizing cell suspensions of S. aciditrophicus (Warikoo et al 1996) and S. gentianae (Schöcke and Schink 1997), which in both cases came up with a minimum free energy change of -40 kJ per mol benzoate degraded. Thus, these bacteria could synthesize no more than a net amount of one-third to two-thirds of an ATP unit per reaction, and our analysis of reaction steps coupled to ATP expenditure and ATP synthesis balance out to nearly the same overall ATP gain.…”

“…Benzoate activation consumes two ATP equivalents in the hydrolysis of one ATP to AMP + 2 P i . Further energy has to be invested in reversed electron transport steps to allow release of electrons from oxidation reactions of comparably high redox potential [e.g., the pimelyl-CoA dehydrogenase reaction (compound 6 to compound 7) and the glutaryl-CoA dehydrogenase reaction (compound 10 to compound 11)], to be released as molecular hydrogen at hydrogen pressures in the range of 10 Pa, as observed in such cultures (Schöcke and Schink 1997;Schink 1997). In analogy to the butyryl-CoA dehydrogenase reaction in syntrophic butyrate oxidation, these reversed electron transport reactions can be assumed to consume two-thirds of an ATP unit each (Wallrabenstein and Schink 1994).…”

“…Benzoate is one of the products of primary fermentation of organic matter and is formed as an intermediate, e.g., during degradation of aromatic amino acids. Assessments of the energetics of benzoate degradation by syntrophically fermenting bacteria such as Syntrophus buswellii or Syntrophus gentianae on the basis of substrate and product measurements in suspensions of intact cells with and without partner bacteria have shown that these bacteria have an energy span of approximately -40 to -45 kJ available per mol benzoate degraded, equivalent to one-third to two-thirds of an ATP unit (Warikoo et al 1996;Schöcke and Schink 1997).…”

Metabolism of Benzoate, Cyclohex-1-ene Carboxylate, and Cyclohexane Carboxylate by “ Syntrophus aciditrophicus ” Strain SB in Syntrophic Association with H 2 -Using Microorganisms

“…First, members of the genus Syntrophus, which belong to the ␦ subgroup of the class Proteobacteria, T. aromatica, which belongs to the ␤ subgroup of the Proteobacteria, and R. palustris, which belongs to the ␣ subgroup of the Proteobacteria, are phylogenetically distinct. Second, the energy constraints encountered in syntrophic aromatic metabolism (45)(46)(47) are not encountered by organisms that can obtain energy from respiration and photosynthesis. It is obvious that syntrophic microorganisms can obtain energy for growth on benzoate, yet it is not clear how net ATP production occurs if benzoate activation and ring reduction during syntrophic benzoate metabolism occur, as found in benzoatedegrading phototrophs and denitrifiers (18,46,47).…”

2-Hydroxycyclohexanecarboxyl Coenzyme A Dehydrogenase, an Enzyme Characteristic of the Anaerobic Benzoate Degradation Pathway Used by Rhodopseudomonas palustris

“…Bacteria of the genus Syntrophus can grow with benzoate as a sole carbon and energy source when they are in coculture with hydrogen-consuming bacteria, such as methanogens or sulfate reducers, but energy calculations show that they cannot possibly generate sufficient ATP to support growth by using either of the benzoyl-CoA degradation pathways shown in Fig. 1 (31,37). Thermodynamic calculations suggest that a four-electron rather than a two-electron reduction of benzoyl-CoA would not necessarily require the hydrolysis of ATP.…”