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

PLoS One. 2013;8(2):e56037. doi: 10.1371/journal.pone.0056037. Epub 2013 Feb 7.


Metabolic versatility has been increasingly recognized as a major virulence mechanism that enables Mycobacterium tuberculosis to persist in many microenvironments encountered in its host. Glucose is one of the most abundant carbon sources 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 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 a phosphofructokinase (PFK) activity. 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. In co-metabolism experiments, we report that disruption of the glycolytic pathway at the PFK step results in intracellular accumulation of sugar-phosphates that correlated with significant impairment of the cell viability. Concomitantly, we found that the presence of glucose is highly toxic for the long-term survival of hypoxic non-replicating mycobacteria, suggesting that accumulation of glucose-derived toxic metabolites does occur in the absence of sustained aerobic respiration. The culture medium traditionally used to study the physiology of hypoxic mycobacteria is supplemented with glucose. In this medium, M. tuberculosis can survive for only 7-10 days in a true non-replicating state before death is observed. By omitting glucose in the medium this period could be extended for up to at least 40 days without significant viability loss. Therefore, our study suggests that glycolysis leads to accumulation of glucose-derived toxic metabolites that limits long-term survival of hypoxic mycobacteria. Such toxic effect is exacerbated when the glycolytic pathway is disrupted at the PKF step.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Carbon / metabolism*
  • Disease Models, Animal
  • Enzyme Activation
  • Female
  • Fructosephosphates / metabolism
  • Gene Knockout Techniques
  • Gene Order
  • Glucose / metabolism
  • Glycolysis
  • Humans
  • Hypoxia*
  • Kinetics
  • Mice
  • Mutation
  • Mycobacterium tuberculosis / genetics
  • Mycobacterium tuberculosis / growth & development
  • Mycobacterium tuberculosis / metabolism*
  • Phenotype
  • Phosphofructokinases / genetics
  • Phosphofructokinases / metabolism*
  • Tuberculosis / microbiology
  • Tuberculosis / mortality
  • Virulence / genetics


  • Fructosephosphates
  • fructose-6-phosphate
  • Carbon
  • Phosphofructokinases
  • Glucose

Grant support

This work was funded by the Novartis Institute of Tropical Diseases and the Singapore National Research Foundation under the Singapore-MIT Alliance for Research and Technology (SMART). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.