Interchangeability of periplasmic adaptor proteins AcrA and AcrE in forming functional efflux pumps with AcrD in Salmonella enterica serovar Typhimurium.
Interchangeability of periplasmic adaptor proteins AcrA and AcrE in forming functional efflux pumps with AcrD in Salmonella enterica serovar Typhimurium.
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DOI：
10.1093/jac/dkab237
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发表时间：
2021-09-15
期刊：
影响因子：
--
通讯作者：
Blair JMA
中科院分区：
文献类型：
--
作者：
Alav I;Bavro VN;Blair JMA
耐药-增殖-分裂（RND）外排泵是抗生素耐药性的重要介质。RND泵，包括沙门氏菌中主要的多药外排泵AcrAB-TolC，是具有内膜RND转运蛋白、周质衔接蛋白（PAP）和外膜因子（OMF）的三重系统。我们先前确定了PAP AcrA和RND转运蛋白AcrB之间结合所需的残基，并证明了PAP可以与非同源转运蛋白一起发挥作用。AcrE和AcrD/AcrF分别是AcrA和AcrB的同源物。在这里，我们表明，AcrE可以与AcrD，它不拥有自己的PAP相互作用，并建立以前确定的AcrB结合的残基也参与AcrD结合。 acrD和acrE基因在缺乏acrABDEF（Δ3RND）的菌株中表达。基于先前定义的PAP-RND相互作用和acrA和acrE中产生的相应突变，预测参与混杂相互作用的PAP残基。测定突变株的抗菌药物敏感性。acrD和acrE的共表达显著降低了Δ3RND菌株对AcrD底物的敏感性，表明AcrE可以与AcrD形成功能复合物。沙门氏菌AcrD的底物分布不同于大肠杆菌AcrD。针对AcrA/AcrE中先前定义的PAP-RND相互作用位点的突变损害了AcrD依赖性底物的外排。这些数据表明，AcrE与AcrD形成了一个有流出能力的泵，因此为该泵提供了一种替代PAP。保守的RND结合位点的突变验证了AcrA和AcrE的可重复性，突出了它们作为外排抑制的潜在药物靶标。
Resistance-nodulation-division (RND) efflux pumps are important mediators of antibiotic resistance. RND pumps, including the principal multidrug efflux pump AcrAB-TolC in Salmonella, are tripartite systems with an inner membrane RND transporter, a periplasmic adaptor protein (PAP) and an outer membrane factor (OMF). We previously identified the residues required for binding between the PAP AcrA and the RND transporter AcrB and have demonstrated that PAPs can function with non-cognate transporters. AcrE and AcrD/AcrF are homologues of AcrA and AcrB, respectively. Here, we show that AcrE can interact with AcrD, which does not possess its own PAP, and establish that the residues previously identified in AcrB binding are also involved in AcrD binding. The acrD and acrE genes were expressed in a strain lacking acrABDEF (Δ3RND). PAP residues involved in promiscuous interactions were predicted based on previously defined PAP-RND interactions and corresponding mutations generated in acrA and acrE. Antimicrobial susceptibility of the mutant strains was determined. Co-expression of acrD and acrE significantly decreased susceptibility of the Δ3RND strain to AcrD substrates, showing that AcrE can form a functional complex with AcrD. The substrate profile of Salmonella AcrD differed from that of Escherichia coli AcrD. Mutations targeting the previously defined PAP-RND interaction sites in AcrA/AcrE impaired efflux of AcrD-dependent substrates. These data indicate that AcrE forms an efflux-competent pump with AcrD and thus presents an alternative PAP for this pump. Mutagenesis of the conserved RND binding sites validates the interchangeability of AcrA and AcrE, highlighting them as potential drug targets for efflux inhibition.
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影响因子：
3.2
作者：
Elkins, CA;Nikaido, H
通讯作者：
Nikaido, H
影响因子：
3.6
作者：
Nishino, K;Latifi, T;Groisman, EA
通讯作者：
Groisman, EA
影响因子：
3.4
作者：
Buckley, AM;Webber, MA;Piddock, LJV
通讯作者：
Piddock, LJV
DOI：
10.1016/j.bbapap.2008.10.004
发表时间：
2009-05
期刊：
Biochimica et biophysica acta
影响因子：
--
作者：
Nikaido H;Takatsuka Y
通讯作者：
Takatsuka Y
DOI：
10.1073/pnas.120163297
发表时间：
2000-06-06
影响因子：
11.1
作者：
Datsenko, KA;Wanner, BL
通讯作者：
Wanner, BL