Yoneyama H, Ocaktan A, Gotoh N, Nishino T, Nakae T
Department of Molecular Life Science, School of Medicine, Tokai University, Isehara, Japan.
Biochem Biophys Res Commun. 1998 Mar 27;244(3):898-902. doi: 10.1006/bbrc.1998.8351.
Pseudomonas aeruginosa encodes three sets of antibiotic extrusion proteins designated as MexA,B.OprM, MexC,D-OprJ and MexE,F-OprN regulated by the nalB, nfxB and nfxC genes, respectively. MexB,D,F, OprM, J,N and MexA,C,E function as the inner membrane pumps, the outer membrane channels and the membrane fusion proteins, respectively. To investigate the possibility of subunit interchangeability, we constructed the following combinations of chimeric pumps: MexA,D-OprM/delta MexB, MexC,B-OprM/delta MexA, and MexA,B-OprJ/delta OprM. The strains producing MexA,D-OprM/delta MexB and MexC,B-OprM/delta MexA failed to restore the antibiotic resistance shown in the strains producing the natural combinations of the subunit proteins. These results suggested that the inner membrane components cannot be interchanged. In contrast, the stains producing MexA,B-OprJ/delta OprM exhibited higher resistance to several antibiotics than the mutant lacking OprM and lower resistance than the strain overexpressing OprM. This result suggests that OprJ may complement the OprM function partially. A spectrum of antibiotics, of which the minimum inhibitory concentrations were restored partially by the complementation, was the same as the spectrum to which the nalB type mutant shows resistance. We surmised from these results that the MexA/MexB unit sustains the substrate specificity of the MexA,B-OprM machinery.
铜绿假单胞菌编码三组抗生素外排蛋白，分别命名为MexA、B - OprM，MexC、D - OprJ和MexE、F - OprN，它们分别受nalB、nfxB和nfxC基因调控。MexB、D、F，OprM、J、N和MexA、C、E分别作为内膜泵、外膜通道和膜融合蛋白发挥作用。为了研究亚基互换的可能性，我们构建了以下嵌合泵组合：MexA、D - OprM/缺失MexB，MexC、B - OprM/缺失MexA，以及MexA、B - OprJ/缺失OprM。产生MexA、D - OprM/缺失MexB和MexC、B - OprM/缺失MexA的菌株未能恢复产生亚基蛋白天然组合的菌株所表现出的抗生素抗性。这些结果表明内膜成分不能互换。相比之下，产生MexA、B - OprJ/缺失OprM的菌株对几种抗生素的抗性高于缺失OprM的突变体，而低于过表达OprM的菌株。这一结果表明OprJ可能部分补充OprM的功能。通过互补部分恢复最低抑菌浓度的一系列抗生素，与nalB型突变体显示抗性的抗生素谱相同。我们从这些结果推测MexA/MexB单元维持了MexA、B - OprM机制的底物特异性。
