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Mycobacterium tuberculosis efpA encodes an efflux protein of the QacA transporter family.

ABSTRACT: The Mycobacterium tuberculosis H37Rv efpA gene encodes a putative efflux protein, EfpA, of 55,670 Da. The deduced EfpA protein was similar in secondary structure to Pur8, MmrA, TcmA, LfrA, EmrB, and other members of the QacA transporter family (QacA TF) which mediate antibiotic and chemical resistance in bacteria and yeast. The predicted EfpA sequence possessed all transporter motifs characteristic of the QacA TF, including those associated with proton-antiport function and the motif considered to be specific to exporters. The 1,590-bp efpA open reading frame was G+C rich (65%), whereas the 40-bp region immediately upstream had an A+T bias (35% G+C). Reverse transcriptase-PCR assays indicated that efpA was expressed in vitro and in situ. Putative promoter sequences were partially overlapped by the A+T-rich region and by a region capable of forming alternative secondary structures indicative of transcriptional regulation in analogous systems. PCR single-stranded conformational polymorphism analysis demonstrated that these upstream flanking sequences and the 231-bp, 5' coding region are highly conserved among both drug-sensitive and multiply-drug-resistant isolates of M. tuberculosis. The efpA gene was present in the slow-growing human pathogens M. tuberculosis, Mycobacterium leprae, and Mycobacterium bovis and in the opportunistic human pathogens Mycobacterium avium and Mycobacterium intracellular. However, efpA was not present in 17 other opportunistically pathogenic or nonpathogenic mycobacterial species.

SUBMITTER: Doran JL

PROVIDER: S-EPMC170471 | biostudies-other | 1997 Jan

REPOSITORIES: biostudies-other

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Mycobacterium tuberculosis efpA encodes an efflux protein of the QacA transporter family.

Doran J L JL Pang Y Y Mdluli K E KE Moran A J AJ Victor T C TC Stokes R W RW Mahenthiralingam E E Kreiswirth B N BN Butt J L JL Baron G S GS Treit J D JD Kerr V J VJ Van Helden P D PD Roberts M C MC Nano F E FE

Clinical and diagnostic laboratory immunology 19970101 1

The Mycobacterium tuberculosis H37Rv efpA gene encodes a putative efflux protein, EfpA, of 55,670 Da. The deduced EfpA protein was similar in secondary structure to Pur8, MmrA, TcmA, LfrA, EmrB, and other members of the QacA transporter family (QacA TF) which mediate antibiotic and chemical resistance in bacteria and yeast. The predicted EfpA sequence possessed all transporter motifs characteristic of the QacA TF, including those associated with proton-antiport function and the motif considered ...[more]

PMID: 9008277

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Project description:The increasing occurrence of multidrug-resistant Mycobacterium tuberculosis (Mtb) is a serious threat to global public health. Among the many mechanisms of drug resistance, only resistance-nodulation-division (RND)-type multidrug efflux systems can simultaneously render bacteria tolerant to numerous toxic compounds, including antibiotics. The elevated expression of RND-type xenobiotic efflux transporter complexes, which consist of an inner membrane transporter, membrane fusion protein, and outer membrane channel, plays a major role in multidrug resistance. Among the 14 mycobacterial membrane protein large (MmpL) proteins identified as inner membrane transporters of Mtb, MmpL5 is known to participate in the acquisition of resistance to bedaquiline and clofazimine. MmpL5 exports these drugs by forming a complex with the membrane fusion protein mycobacterial membrane protein small 5 (MmpS5). However, the role of MmpS5 in the efflux of antituberculous drugs by MmpL5 remains unclear. In this study, we focused on the in vivo dynamics of MmpL5 using green fluorescent protein (GFP). Single-molecule observations of MmpL5 showed substantial lateral displacements of MmpL5-GFP without the expression of MmpS5. Nondiffusing MmpL5-GFP foci typically showed three-step photobleaching, suggesting that MmpL5 formed a homotrimeric functional complex on the inner membrane in the presence of MmpS5. These results suggest that the expression of MmpS5 facilitates the assembly of monomeric MmpL5 into a homotrimer that is anchored to the inner membrane to transport various antimycobacterial drugs.IMPORTANCE It has been reported that mycobacterial membrane protein large 5 (MmpL5), a resistance-nodulation-division (RND)-type inner membrane transporter in Mycobacterium tuberculosis (Mtb), is involved in the transport of antimycobacterial drugs. However, the functional roles of the membrane fusion protein mycobacterial membrane protein small 5 (MmpS5), organized as an operon with MmpL5, are unclear. Via the single-molecule imaging of MmpL5, we uncovered the maintenance of the functional trimeric complex structure of MmpL5 in the presence of MmpS5. These findings demonstrate that the assembly mechanisms of mycobacterial RND efflux systems are the dynamically regulated process through interactions among the components. This represents the first report of the single-molecule observation of Mtb efflux transporters, which may enhance our understanding of innate antibiotic resistance.

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Dataset Information

Mycobacterium tuberculosis efpA encodes an efflux protein of the QacA transporter family.

Publications

Mycobacterium tuberculosis efpA encodes an efflux protein of the QacA transporter family.

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