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Abstract

Major facilitator superfamily (MFS) transporters aid bacteria in expelling toxic compounds such as antibiotics, bile acids, and other substances. Previous bioinformatic studies identified the emrB motif as a potential regulatory RNA element. In Lactobacillus, this RNA motif is generally found in the intergenic region upstream of an MFS transporter gene with homology to the emrB gene from Escherichia coli. Secondary structure predictions of the intergenic region were used to define the structural features of this RNA motif. The emrB motif comprises two hairpins, though inclusion of neighboring regions is predicted to stabilize alternative secondary structures. A potential rho independent terminator was identified adjacent to the emrB motif. Our working hypothesis is that in the presence of antibiotics or bile salts, the emrB motif acts as an anti-terminator to suppress transcriptional termination and allow for expression of emrB gene. The operon containing the emrB gene includes: an antibiotic resistance gene, vanZ, and a gene for glucose-6-phosphate-isomerase. The emrB motif may bind to antibiotics or bile salts that stabilize the antiterminator structure to promote transcription of the emrB gene. Initial characterization of the emrB motif used DNA templates derived by PCR from Lactobacillus acidophilus genomic DNA to synthesize various RNA constructs. The results obtained from thermal melt assays of the RNA transcripts indicated that the motif forms a very stable structure, which is consistent with 3D structure predictions showing a 3-helix junction that may form a binding site for antibiotics associated with the regulatory function of the emrB motif. Analyzing the structure and function of the emrB motif is important for understanding possible mechanisms of antibiotic resistance and bile tolerance in the gastrointestinal microbiota of humans.

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