Structural basis for enzyme I inhibition by α-ketoglutarate

ACS Chem Biol. 2013;8(6):1232-40. doi: 10.1021/cb400027q. Epub 2013 Mar 29.


Creating new bacterial strains in which carbon and nitrogen metabolism are uncoupled is potentially very useful for optimizing yields of microbial produced chemicals from renewable carbon sources. However, the mechanisms that balance carbon and nitrogen consumption in bacteria are poorly understood. Recently, α-ketoglutarate (αKG), the carbon substrate for ammonia assimilation, has been observed to inhibit Escherichia coli enzyme I (EI), the first component of the bacterial phosphotransferase system (PTS), thereby providing a direct biochemical link between central carbon and nitrogen metabolism. Here we investigate the EI-αKG interaction by NMR and enzymatic assays. We show that αKG binds with a KD of ∼2.2 mM at the active site of EI, acting as a competitive inhibitor. In addition, we use molecular docking simulations to derive a structural model of the enzyme-inhibitor complex that is fully consistent with NMR and analytical ultracentrifugation data. We expect that the EI-αKG structure presented here will provide a starting point for structure-based design of EI mutants resistant to αKG.

Publication types

  • Research Support, N.I.H., Intramural

MeSH terms

  • Amino Acid Sequence
  • Escherichia coli / chemistry
  • Escherichia coli / enzymology*
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / chemistry*
  • Escherichia coli Proteins / metabolism*
  • Ketoglutaric Acids / chemistry
  • Ketoglutaric Acids / metabolism*
  • Molecular Docking Simulation
  • Molecular Sequence Data
  • Monosaccharide Transport Proteins / chemistry*
  • Monosaccharide Transport Proteins / metabolism*
  • Phosphoenolpyruvate Sugar Phosphotransferase System / chemistry*
  • Phosphoenolpyruvate Sugar Phosphotransferase System / metabolism*
  • Protein Structure, Tertiary
  • Sequence Alignment


  • Escherichia coli Proteins
  • Ketoglutaric Acids
  • Monosaccharide Transport Proteins
  • Phosphoenolpyruvate Sugar Phosphotransferase System
  • mannitol PTS permease, E coli