CcpA-mediated regulation of cellular energy metabolism in the ruminal bacterium .

This study investigated the cellular energy metabolite profiles of S1. Glucose concentrations of 5 or 50 mM and the presence or absence of were applied during the cultivation of S1. Results revealed the identification of 51 metabolites categorized into seven types: amino acid derivatives, amino acids, coenzymes and vitamins, nucleotides and their metabolites, organic acids and their derivatives, phosphate sugars, and phosphoric acids. Each group exhibited distinct cellular energy metabolite profiles, as evidenced by principal component analysis showing unique metabolite compositions. Orthogonal projections to latent structures discriminant analysis (OPLS-DA) further distinguished between each pair of groups. KEGG prediction indicated that the abundance and enrichment of metabolites were primarily involved in carbon metabolism, amino acid metabolism, and nucleotide metabolomics. Based on the random forest algorithm, glutamine and UDP-GlcNac were identified as biomarkers under varying glucose conditions and in the presence or absence of . Further analysis reveals that low glucose restricts carbon flux to the Embden-Meyerhof-Parnas (EMP) pathway, while knockout further reduces flux through this pathway. Glucose and CcpA might regulate fructose-1,6-bisphosphate (FBP) concentration to modulate lactate dehydrogenase and pyruvate formate-lyase enzyme activity, thereby influencing the fermentation direction of pyruvate. In low-glucose environments, glutamine serves to alleviate glucose deficiency, and the interaction between glucose and CcpA may mediate the fate of amino acids differently. Low glucose limits guanine synthesis but not adenine, cytosine, or thymine synthesis, while knockout disrupts both synthesis pathways by inhibiting the pentose phosphate pathway (PPP). Overall, this study provides insights into the intricate interplay between glucose concentration, knockout, and cellular energy metabolism in S1.IMPORTANCE S1 plays a pivotal role in lactate production in the rumen, increasing the risk of rumen acidosis. Modulating the fermentation profile of S1 could reduce lactate accumulation, potentially improving rumen health. In this study, knockout decreased fermentation fluid lactate concentration but increased formate concentration. Liquid chromatography tandem mass spectrometry characterized the metabolic activity of S1 under varying glucose concentrations. We found that CcpA regulates central carbon metabolism, including the EMP pathway, gluconeogenesis, and the PPP in S1. Additionally, glucose and CcpA likely influence pyruvate fermentation, directing it toward lactate or formate production by modulating FBP concentrations. These findings underscore the regulatory roles of glucose concentration and CcpA in metabolic pathways, particularly in fermentation and energy metabolism in S1.