A Sodium-Translocating Module Linking Succinate Production to Formation of Membrane Potential in Prevotella bryantii.

Ruminants such as cattle and sheep depend on the breakdown of carbohydrates from plant-based feedstuff, which is accomplished by the microbial community in the rumen. Roughly 40% of the members of the rumen microbiota belong to the family , which ferments sugars to organic acids such as acetate, propionate, and succinate. These substrates are important nutrients for the ruminant. In a metaproteome analysis of the rumen of cattle, proteins that are homologous to the Na-translocating NADH:quinone oxidoreductase (NQR) and the quinone:fumarate reductase (QFR) were identified in different species. Here, we show that fumarate reduction to succinate in anaerobically growing Prevotella bryantii is coupled to chemiosmotic energy conservation by a supercomplex composed of NQR and QFR. This odium-translocating ADH:umarate oxidoeductase (SNFR) supercomplex was enriched by blue native PAGE (BN-PAGE) and characterized by in-gel enzyme activity staining and mass spectrometry. High NADH oxidation (850 nmol minmg), quinone reduction (490 nmol minmg), and fumarate reduction (1,200 nmol minmg) activities, together with high expression levels, demonstrate that SNFR represents a charge-separating unit in . Absorption spectroscopy of SNFR exposed to different substrates revealed intramolecular electron transfer from the flavin adenine dinucleotide (FAD) cofactor in NQR to heme cofactors in QFR. SNFR catalyzed the stoichiometric conversion of NADH and fumarate to NAD and succinate. We propose that the regeneration of NAD in is intimately linked to the buildup of an electrochemical gradient which powers ATP synthesis by electron transport phosphorylation. Feeding strategies for ruminants are designed to optimize nutrient efficiency for animals and to prevent energy losses like enhanced methane production. Key to this are the fermentative reactions of the rumen microbiota, dominated by spp. We show that succinate formation by is coupled to NADH oxidation and sodium gradient formation by a newly described supercomplex consisting of Na-translocating NADH:quinone oxidoreductase (NQR) and fumarate reductase (QFR), representing the odium-translocating ADH:umarate oxidoeductase (SNFR) supercomplex. SNFR is the major charge-separating module, generating an electrochemical sodium gradient in . Our findings offer clues to the observation that use of fumarate as feed additive does not significantly increase succinate production, or decrease methanogenesis, by the microbial community in the rumen.