Mechanism of Rifampicin Inactivation in Nocardia farcinica

PLoS One. 2016 Oct 5;11(10):e0162578. doi: 10.1371/journal.pone.0162578. eCollection 2016.


A novel mechanism of rifampicin (Rif) resistance has recently been reported in Nocardia farcinica. This new mechanism involves the activity of rifampicin monooxygenase (RifMO), a flavin-dependent monooxygenase that catalyzes the hydroxylation of Rif, which is the first step in the degradation pathway. Recombinant RifMO was overexpressed and purified for biochemical analysis. Kinetic characterization revealed that Rif binding is necessary for effective FAD reduction. RifMO exhibits only a 3-fold coenzyme preference for NADPH over NADH. RifMO catalyzes the incorporation of a single oxygen atom forming an unstable intermediate that eventually is converted to 2'-N-hydroxy-4-oxo-Rif. Stable C4a-hydroperoxyflavin was not detected by rapid kinetics methods, which is consistent with only 30% of the activated oxygen leading to product formation. These findings represent the first reported detailed biochemical characterization of a flavin-monooxygenase involved in antibiotic resistance.

MeSH terms

  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Biocatalysis
  • Chromatography, High Pressure Liquid
  • Flavin-Adenine Dinucleotide / chemistry
  • Flavin-Adenine Dinucleotide / metabolism
  • Kinetics
  • NADP / chemistry
  • NADP / metabolism
  • Nocardia / drug effects
  • Nocardia / enzymology*
  • Oxidation-Reduction
  • Oxygen / chemistry
  • Oxygen / metabolism
  • Oxygenases / genetics
  • Oxygenases / metabolism
  • Protein Binding
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / isolation & purification
  • Rifampin / analysis
  • Rifampin / chemistry
  • Rifampin / metabolism*
  • Rifampin / pharmacology


  • Bacterial Proteins
  • Recombinant Proteins
  • Flavin-Adenine Dinucleotide
  • NADP
  • Oxygenases
  • dimethylaniline monooxygenase (N-oxide forming)
  • Oxygen
  • Rifampin

Grant support

This work was supported by National Science Foundation grants CHE-1506206 and MCB 1021384. This work was also supported by a Joint Supervision Fellowship to H. Abdelwahab funded by the Cultural Affairs and Mission sectors of the Egyptian Ministry of Higher Education. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.