Characterization of a Rifampin-Inactivating Glycosyltransferase from a Screen of Environmental Actinomycetes
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ABSTRACT
Identifying and understanding the collection of all antibiotic resistance determinants presented in the global microbiota, the antibiotic resistome, provides insight into the evolution of antibiotic resistance and critical information for the development of future antimicrobials. The rifamycins are broad-spectrum antibiotics that target bacterial transcription by inhibition of RNA polymerase. Although mutational alteration of the drug target is the predominant mechanism of resistance to this family of antibiotics in the clinic, a number of diverse inactivation mechanisms have also been reported. In this report, we investigate a subset of environmental rifampin-resistant actinomycete isolates and identify a diverse collection of rifampin inactivation mechanisms. We describe a single isolate, WAC1438, capable of inactivating rifampin by glycosylation. A draft genome sequence of WAC1438 (most closely related to
Streptomyces speibonae
, according to a 16S rRNA gene comparison) was assembled, and the associated rifampin glycosyltransferase open reading frame,
rgt1438
, was identified. The role of
rgt1438
in rifampin resistance was confirmed by its disruption in the bacterial chromosome, resulting in a loss of antibiotic inactivation and a 4-fold decrease in MIC. Interestingly, examination of the RNA polymerase β-subunit sequence of WAC1438 suggests that it harbors a resistant target and thus possesses dual mechanisms of rifamycin resistance. Using an
in vitro
assay with purified enzyme, Rgt1438 could inactivate a variety of rifamycin antibiotics with comparable steady-state kinetics constants. Our results identify
rgt1438
as a rifampin resistance determinant from WAC1438 capable of inactivating an assortment of rifamycins, adding a new element to the rifampin resistome.
