The ATP-binding cassette transporter CstA as a novel therapeutic target for treating carbapenem-resistant Enterobacteriaceae bacteria.

BACKGROUND

Nosocomial infection rates by carbapenem-resistant Enterobacteriaceae (CRE) bacteria are increasing annually. Although carbapenemase is considered the main mechanism of carbapenem resistance, RND family multi-drug transporters are also associated with carbapenem resistance. However, it remains unclear whether CRE bacteria have other carbapenem transporters.

RESULTS

LMJ45_RS15935 was annotated as an ATP-binding cassette transporter-encoding gene, and phylogenetic affiliation analysis suggested that LMJ45_RS15935 evolved from the related Enterobacteriaceae protein YojI. MIC analysis revealed that the LMJ45_RS15935 transporter specifically mediates carbapenem resistance; therefore, LMJ45_RS15935 was renamed as CstA (carbapenem-specific transporter A). The natural compound α-thujone was found to significantly reduce the carbapenem MICs of the recombinant Escherichia coli strain expressing CstA, and molecular docking analysis revealed that CstA binds to meropenem and α-thujone. α-thujone effectively inhibited the efflux of meropenem and the ATPase activity of CstA. The key amino acid sites in CstA for binding to meropenem were R116, D183 and L108, and for binding to α-thujone were L460, V402 and V406. Combination of meropenem and α-thujone effectively eliminated the CstA-expressing E. coli strain; this drug combination also effectively treated the peritonitis in mice infected with the CstA-expressing E. coli strain, with enhanced survival of the mice and significantly reduced bacterial loads. These findings indicate that CstA mediates carbapenem-specific resistance and that α-thujone is an effective inhibitor of CstA.

CONCLUSIONS

This study indicates that the ATP-binding cassette transporter CstA mediates carbapenem resistance in Enterobacteriaceae bacteria and suggests CstA is a potential drug target for treating CRE bacterial infections.