Tuberculosis (TB) is an infectious disease caused by Mtb. There are about 10 million new TB cases globally each year. Of note, the emergence and spread of multi-drug and extensive drug resistance Mtb strains challenge the control and treatment of TB.

In a study published in PNAS, a research group from Tianjin Institute of Industrial Biotechnology of the Chinese Academy of Sciences, along with collaborators, revealed the actual function of EfpA as a lipid transporter and uncovered the structural basis for its interaction either with both phosphatidic acid (PA) and the drug molecule, a BRD-8000.3, which will facilitate the development of anti-TB drugs.

It is known that Mtb develops drug resistance through various mechanism, with efflux pump-mediated drug extrusion being one of the majors. EfpA is the first MFS (facilitator superfamily) efflux pump identified in Mtb, and it is essential to the bacterium’s survival.

Previous studies have reported that the expression of EfpA contributes to resistance against both first- and second-line anti-TB drugs. More importantly, inhibitors targeting EfpA have been shown to inhibit the growth and kill Mtb, highlighting EfpA as a promising drug target. However, the exact function of EfpA and the mechanism of EfpA-dependent drug resistance remain largely unknown.

In this study, the researchers first solved the cryo-EM structure of EfpA in complex with multiple lipids at a resolution of 2.9 Å, and revealed that EfpA assembles into an antiparallel dimer, with each protomer consisting of 14 transmembrane helices. Then, a range of lipids bound to EfpA were detected by LC-MS/MS, among which PA matches best with the densities in the central cavity and the lateral fenestration. Using a lipid flipping assay, researchers demonstrated that EfpA functions as a lipid transporter, and its transport activity can be inhibited by BRD-8000.3.

Moreover, researchers solved the cryo-EM structure of EfpA in complex with BRD-8000.3, and revealed that two BRD-8000.3 molecules occupy the lipid binding site at the dimer interface. Through structure analysis, mutations at the position V319 of EfpA were constructed, and it turned out that the expression level of V319F mutant rather than V319M increased more than tenfold compared to wild type. The oligomeric state of V319F mutant was also altered, which may contribute to its resistance to BRD-8000.3.

The findings of this study suggest a potential role for EfpA in the synthesis of Mtb cell envelope, and that the lipid transport activity may be associated to the drug resistance caused by EfpA.