Abstract
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.
Highlights
Nocardia spp. are Gram-positive bacteria that cause a lung infection, known as pulmonary nocordiosis, in immuno-compromised and immuno-competent individuals [1]
The mechanism of resistance can originate by mutation(s) of the target protein or enzyme, which leads to reduced antibiotic activity [10, 17, 21]
The most common mechanism of Rif resistance occurs via mutations on the RNA polymerase (RNAP) β-subunit; sequence analysis of resistant strains identified mutations in clusters that are known as the Rif resistant-determining region (RRDR) [16, 26, 27]
Summary
Nocardia spp. are Gram-positive bacteria that cause a lung infection, known as pulmonary nocordiosis, in immuno-compromised and immuno-competent individuals [1]. Rifampicin (Rif) is a member of the rifamycin family of antibiotics used in combinational therapy for the treatment of nocardiosis and numerous mycobacterial infections, including tuberculosis and leprosy [1, 2]. The emergence of Rif resistance has limited its prolonged use in the treatment for nocardiosis [4]. For this reason, different approaches have been considered, such as semi-synthetic Rif modifications, which leads to decreased toxicity and shortened treatment duration [5]. In Nocardia farcinica IFM 10152, one of PLOS ONE | DOI:10.1371/journal.pone.0162578. In Nocardia farcinica IFM 10152, one of PLOS ONE | DOI:10.1371/journal.pone.0162578 October 5, 2016
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