Overexpression of the mexC–mexD–oprJ efflux operon in nfxB‐type multidrug‐resistant strains of Pseudomonas aeruginosa

作者：Keith G. Poole, Naomasa Gotoh, Hideto Tsujimoto, Qixun Zhao, 和田明久, Tetsuo Yamasaki, Shádi Neshat, Jun‐ichi Yamagishi, Xian-Zhi Li, Takeshi Nishino · 发表于：Molecular Microbiology · 年份：1996 · DOI：10.1046/j.1365-2958.1996.281397.x · 被引用次数：447 · 研究领域：Antibiotic Resistance in Bacteria、Bacterial Genetics and Biotechnology、Bacterial biofilms and quorum sensing

OprJ, overproduced in nfxB multidrug-resistant strains of Pseudomonas aeruginosa, and OprK, overproduced in the multidrug-resistant strain K385, were demonstrated to be immunologically cross-reactive using an OprJ-specific monoclonal antibody. Treatment of the purified proteins with trypsin or chymotrypsin yielded virtually indistinguishable digestion patterns, and the N-terminal sequence of two trypsin fragments was identical for both proteins, indicating that OprJ and OprK share identity. The N-terminal amino acid sequences were used to facilitate cloning of the oprJ gene on a 5kbp Kpnl fragment and a 10 kbp BamHl fragment. Nucleotide sequencing of portions of these fragments revealed that oprJ was the terminal gene in a putative three-gene operon, mexC-mexD-oprJ. The predicted mexC-mexD-oprJ gene products exhibit homology to the MexA-MexB-OprM components of the multidrug-resistance efflux pump of P. aeruginosa (43-46% identity). Consistent with an implied role for mexC-mexD-oprJ in drug efflux, the mexC-mexD-oprJ-hyperexpressing strain K385 showed reduced accumulation of a variety of antibiotics as compared with its parent strain, and this drug 'exclusion' was abrogated by energy inhibitors. The mexC and oprJ products are putative lipoproteins of a molecular mass of 40,707 and 51,742 Da, respectively, while mexD was predicted to encode a protein of 111 936 Da. Sequencing upstream of mexC revealed the presence of the nfxB gene transcribed divergently from the efflux genes. ...