Characterization of Phosphofructokinase Activity in Mycobacterium tuberculosis Reveals That a Functional Glycolytic Carbon Flow Is Necessary to Limit the Accumulation of Toxic Metabolic Intermediates under Hypoxia

Mycobacterium tuberculosis metabolic versatility has been increasingly recognized as a major virulence mechanism enabling this pathogen to persist in many microenvironments encountered in its host. Glucose is one of the most abundant source of carbon that is exploited by many pathogenic bacteria in the human host. M. tuberculosis has an intact glycolytic pathway that is highly conserved in all clinical isolates sequenced to date suggesting that, in addition to lipids, glucose may represent a non-negligible source of carbon and energy for this pathogen in vivo. Fructose-6-phosphate phosphorylation represents the key-committing step in glycolysis and is catalyzed by phosphofructokinase (PFK). Two genes, pfkA and pfkB, have been annotated to encode putative PFK in M. tuberculosis. Here, we show that PFKA is the sole PFK enzyme in M. tuberculosis with no functional redundancy with PFKB. PFKA is required for growth on glucose as sole carbon source. Furthermore, in co-metabolism experiments, a disrupted glycolytic pathway resulted in decreased survival due to the accumulation of glucose-derived toxic intermediate metabolites. Coincidentally we found that glucose metabolism is highly toxic for the long term survival of hypoxic non-replicating M. tuberculosis. Indeed, M. tuberculosis survived several order of magnitudes better in a glucose-depleted culture medium, compared to what is traditionally achieved in the original glucose-supplemented medium. This novel finding improves the potential and relevance of the Wayne model for the study of the mechanisms of persistence in M. tuberculosis. In conclusion, although a functional glycolytic pathway is not required for infection and persistence in the mouse model, we propose that glycolysis is required for regulating the pool of sugar phosphate that may be otherwise toxic for hypoxic mycobacteria