Inactivation Of The Pta-Acka Pathway Impairs Fitness Of Bacillus Anthracis During Overflow Metabolism

Under conditions of glucose excess, aerobically growing bacteria predominantly direct carbon flux toward acetate fermentation, a phenomenon known as overflow metabolism or the bacterial "Crabtree effect." Numerous studies of the major acetate-generating pathway, the phosphotransacetylase (Pta)-acetate kinase (AckA) pathway, have revealed its important role in bacterial fitness through the control of central metabolism to sustain balanced growth and cellular homeostasis. In this work, we highlight the contribution of the Pta-AckA pathway to the fitness of the spore-forming bacterium Bacillus anthracis. We demonstrate that disruption of the Pta-AckA pathway causes drastic growth reduction in the mutants and alters the metabolic and energy status of the cells. Our results revealed that inactivation of the Pta-AckA pathway increases the glucose consumption rate, affects intracellular ATP, NAD(+), and NADH levels, and leads to a metabolic block at the pyruvate and acetyl coenzyme A (acetyl-CoA) nodes. Consequently, accumulation of intracellular acetylCoA and pyruvate forces bacteria to direct carbon into the tricarboxylic acid and/or glyoxylate cycles, as well as fatty acid and poly(3-hydroxybutyrate) biosynthesis pathways. Notably, the presence of phosphotransbutyrylase (Ptb) in B. anthracis partially compensates for the loss of Pta activity. Furthermore, overexpression of the ptb gene not only eliminates the negative impact of the pta mutation on B. anthracis fitness but also restores normal growth in the pta mutant of the non-butyrate-producing bacterium Staphylococcus aureus. Taken together, the results of this study demonstrate the importance of the Pta-AckA pathway for B. anthracis fitness by revealing its critical contribution to the maintenance of metabolic homeostasis during aerobic growth under conditions of carbon overflow.IMPORTANCE B. anthracis, the etiological agent of anthrax, is a highly pathogenic, spore-forming bacterium that causes acute, life-threatening disease in both humans and livestock. A greater understanding of the metabolic determinants governing the fitness of B. anthracis is essential for the development of successful therapeutic and vaccination strategies aimed at lessening the potential impact of this important biodefense pathogen. This study is the first to demonstrate the vital role of the PtaAckA pathway in preserving energy and metabolic homeostasis in B. anthracis under conditions of carbon overflow, thus highlighting this pathway as a potential therapeutic target for drug discovery. Overall, the results of this study provide important insights into the metabolic processes and requirements driving rapid B. anthracis proliferation during vegetative growth.