摘要

外膜蛋白（OMPs）在抗生素抗性中发挥重要作用，特别是在革兰氏阴性菌中;然而，他们仍然有很多关于他们对抗生素应激反应的行为的不明功能。在目前的工作中，使用基于定量串联质量标签标记的质谱法来比较土霉素嗜水气单胞菌中抗土霉素（OXY-R）和其原始对照染色（OXY-O）之间的外膜相关蛋白。因此，在两个生物重复中共鉴定出261种常见的蛋白质，其中29种蛋白质增加，28种蛋白质减少。基因本体分析显示转运蛋白的表达显着降低，并且翻译相关蛋白在OXY-R株中下调。在使用蛋白质印迹验证选择的改变的蛋白质后，敲除了8个OMP相关基因，并进一步评估了它们在抗生素抗性中的作用。存活测定显示一些突变体如ΔAHA_4281，ΔAHA_2766，ΔAHA_2282，ΔAHA_1181和ΔAHA_1280影响嗜水气单胞菌对抗微生物剂的敏感性。此外，最小抑制浓度测定显示这些候选突变体对其他类型的抗生素也有不同的反应。我们的研究结果揭示了嗜水气单胞菌的几种新型外膜相关蛋白，它们在抗生素抗性中起重要作用，因此，可能有助于筛选研究以鉴定新的药物靶点。

Outer membrane proteins (OMPs) play essential roles in antibiotic resistance, particularly in Gram-negative bacteria; however, they still have many unidentified functions regarding their behavior in response to antibiotic stress. In the current work, quantitative tandem mass tag labeling-based mass spectrometry was used to compare the outer membrane related proteins between an oxytetracycline-resistant (OXY-R) and its original control stain (OXY-O) in Aeromonas hydrophila. Consequently, a total of 261 commonly altered proteins in two biological repeats were identified including 29 proteins that increased and 28 that decreased. Gene ontology analysis showed that the expression of transport proteins was significantly reduced, and translation-related proteins were downregulated in the OXY-R strain. After using western blotting to validate selected altered proteins, eight OMP-related genes were knocked out and their roles in antibiotic resistance were further evaluated. The survival assays showed that some mutants such as ΔAHA_4281, ΔAHA_2766, ΔAHA_2282, ΔAHA_1181, and ΔAHA_1280 affected the susceptibility of A. hydrophila to antimicrobials. Moreover, the minimum inhibitory concentration assay showed that these candidate mutants also respond differently to other types of antibiotics. Our results reveal several novel outer membrane related proteins of A. hydrophila that play important roles in antibiotic resistance, and as such, may be helpful for screening studies to identify novel drug targets.

https://www.ncbi.nlm.nih.gov/pubmed/30460208