Li Hui, Zhang Dan-feng, Lin Xiang-min, Peng Xuan-xian
Center for Proteomics and Metabolomics, State Key Laboratory of Biocontrol, MOE Key Lab Aquatic Food Safety, School of Life Sciences, Sun Yat-sen University, University City, Guangzhou 510006, People's Republic of China.
School of Biological Science and Biotechnology, Minnan Normal University, Zhangzhou 363000, People's Republic of China.
FEMS Microbiol Lett. 2015 Jun;362(11). doi: 10.1093/femsle/fnv074. Epub 2015 May 3.
Antibiotic-resistant bacteria are a great threat to human health and food safety and there is an urgent need to understand the mechanisms of resistance for combating these bacteria. In the current study, comparative proteomic methodologies were applied to identify Escherichia coli K-12 outer membrane (OM) proteins related to kanamycin resistance. Mass spectrometry and western blotting results revealed that OM proteins TolC, Tsx and OstA were up-regulated, whereas MipA, OmpA, FadL and OmpW were down-regulated in kanamycin-resistant E. coli K-12 strain. Genetic deletion of tolC (ΔtolC-Km) led to a 2-fold decrease in the minimum inhibitory concentration (MIC) of kanamycin and deletion of mipA (ΔmipA-Km) resulted in a 4-fold increase in the MIC of kanamycin. Changes in the MICs for genetically modified strains could be completely recovered by gene complementation. Compared with the wild-type strain, the survival capability of ΔompA-Km was significantly increased and that of Δtsx-Km was significantly decreased. We further evaluated the role and expression of MipA in response to four other antibiotics including nalidixic acid, streptomycin, chloramphenicol and aureomycin, which suggested that MipA was a novel OM protein related to antibiotic resistance.
抗生素耐药菌对人类健康和食品安全构成巨大威胁，迫切需要了解耐药机制以对抗这些细菌。在本研究中，应用比较蛋白质组学方法鉴定与卡那霉素耐药相关的大肠杆菌K-12外膜（OM）蛋白。质谱和蛋白质印迹结果显示，在卡那霉素耐药的大肠杆菌K-12菌株中，OM蛋白TolC、Tsx和OstA上调，而MipA、OmpA、FadL和OmpW下调。tolC基因缺失（ΔtolC-Km）导致卡那霉素的最低抑菌浓度（MIC）降低2倍，mipA基因缺失（ΔmipA-Km）导致卡那霉素的MIC增加4倍。基因改造菌株的MIC变化可通过基因互补完全恢复。与野生型菌株相比，ΔompA-Km的生存能力显著增加，而Δtsx-Km的生存能力显著降低。我们进一步评估了MipA在应对其他四种抗生素（包括萘啶酸、链霉素、氯霉素和金霉素）时的作用和表达，这表明MipA是一种与抗生素耐药相关的新型OM蛋白。
