Catch me if you can: a biotinylated proteoliposome affinity assay for the investigation of assembly of the MexA-MexB-OprM efflux pump from Pseudomonas aeruginosa
Véronique Yvette Ntsogo Enguéné, Alice Verchère, Gilles Phan, Isabelle Broutin, Martin Picard
Abstract
Efflux pumps are membrane transporters that actively extrude various substrates, leading to multidrug resistance (MDR). In this study, we have designed a new test that allows investigating the assembly of the MexA-MexB-OprM efflux pump from the Gram negative bacteria Pseudomonas aeruginosa. The method relies on the streptavidin-mediated pull-down of OprM proteoliposomes upon interaction with MexAB proteoliposomes containing a biotin function carried by lipids. We give clear evidence for the importance of MexA in promoting and stabilizing the assembly of the MexAB-OprM complex. In addition, we have investigated the effect of the role of the lipid anchor of MexA as well as the role of the proton motive force on the assembly and disassembly of the efflux pump. The assay presented here allows for an accurate investigation of the assembly with only tens of microgram of protein and could be adapted to 96 wells plates. Hence, this work provides a basis for the medium-high screening of efflux pump inhibitors (EPIs).
Pharmacology, Toxicology and Pharmaceutical Science
2
4%
Engineering
2
4%
Other
5
11%
Unknown
13
29%
Attention Score in Context
Attention Score in Context
This research output has an Altmetric Attention Score of 2. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 18 June 2015.
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#14,810,408
of 22,803,211 outputs
Outputs from Frontiers in Microbiology
#13,783
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#148,074
of 267,789 outputs
Outputs of similar age from Frontiers in Microbiology
#199
of 390 outputs
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