Pushpker Rajnigandha, Bay Denice C, Turner Raymond J
Department of Biological Sciences, University of Calgary, Calgary, Alberta, Canada.
Department of Medical Microbiology and Infectious Diseases, University of Manitoba, Winnipeg, Manitoba, Canada.
Microbiology (Reading). 2022 Dec;168(12). doi: 10.1099/mic.0.001287.
The small multidrug resistance (SMR) protein EmrE resides in the inner membrane and provides resistance against a wide range of antiseptic quaternary cationic compounds (QCCs) for the Gram-negative bacterium . We have reported previously that overexpression of the gene results in the reduction of pH and osmotic tolerance, likely through EmrE-mediated biological QCC-based osmoprotectant efflux, indicating a potential physiological role for EmrE beyond providing drug resistance. EmrE is the most studied member of SMR transporter family; however, it is not known how the substrates translocated by EmrE move across the periplasm and through the outer membrane (OM). We have shown that the OM protein OmpW participates in the EmrE-mediated substrate efflux process and provided a hypothesis for the present study that additional OM and periplasmic proteins participate in the translocation process. To test the hypothesis, we conducted alkaline pH-based growth phenotype screens under overexpression conditions. This screen identified 10 additional genes that appear to contribute to the EmrE-coupled osmoprotectant efflux: , , , , , , , , and . Further screening of these genes using a hyperosmotic growth phenotype assay in the presence and the absence of the osmoprotectant glycine betaine identified and two periplasmic protein genes, and , are mechanistically linked to EmrE.
小多药耐药（SMR）蛋白EmrE位于内膜中，为革兰氏阴性菌提供对多种防腐季铵阳离子化合物（QCCs）的抗性。我们之前报道过，该基因的过表达会导致pH值降低和渗透压耐受性降低，可能是通过EmrE介导的基于生物QCC的渗透保护剂外排，这表明EmrE除了提供耐药性之外还有潜在的生理作用。EmrE是SMR转运蛋白家族中研究最多的成员；然而，尚不清楚EmrE转运的底物如何穿过周质和外膜（OM）。我们已经表明，外膜蛋白OmpW参与EmrE介导的底物外排过程，并为本研究提供了一个假设，即额外的外膜和周质蛋白参与转运过程。为了验证这一假设，我们在过表达条件下进行了基于碱性pH值的生长表型筛选。该筛选确定了另外10个似乎有助于EmrE偶联渗透保护剂外排的基因： 、 、 、 、 、 、 、 、 和 。在存在和不存在渗透保护剂甘氨酸甜菜碱的情况下，使用高渗生长表型分析对这些基因进行进一步筛选，确定 和两个周质蛋白基因 、 在机制上与EmrE相关。
