Douglas R M, Ritchie G Y, Munro A W, McLaggan D, Booth I R
Department of Molecular and Cell Biology, University of Aberdeen, Marischal College, UK.
Mol Membr Biol. 1994 Jan-Mar;11(1):55-61. doi: 10.3109/09687689409161030.
KefC is a glutathione-gated K(+)-efflux system that is widespread in Gram-negative bacteria and which plays a role in the protection of cells from the toxic effects of electrophilic reagents, such as N-ethylmaleimide (NEM). The KefC gene from Escherichia coli has been cloned and the DNA sequenced. A number of kefC mutants that affect K+ retention by the KefC system have been isolated and all retain activation by NEM. Cloned kefC was found to suppress the phenotype of two such mutants kefC121 and kefC103. Analysis of this phenomenon has shown that suppression is specific to the KefC system, but that cloned kefC from Klebsiella and Erwinia can also mediate suppression of the mutant phenotype. Plasmid constructs of the E. coli gene in which expression of the cloned gene was diminished showed induced ability to suppress the mutant phenotype. KefC'-'LacZ hybrid proteins were inserted in the membrane but did not suppress the mutant phenotype. Cloned kefC did not suppress a mutant kefB allele that exhibited a similar phenotype to the kefC121 allele. These data suggest that suppression is unlikely to arise from exclusion of the mutant form of the protein from the membrane. Furthermore, NEM-activated K+ efflux from a strain carrying both the mutant and cloned wild-type alleles was faster than when either allele was present in cells alone, suggesting that both forms of the protein are inserted into the membrane. These data are discussed in terms of a model for the KefC protein in which the protein is composed of one or more identical subunits that interact in the membrane.
KefC是一种谷胱甘肽门控的钾离子外流系统，广泛存在于革兰氏阴性菌中，在保护细胞免受亲电试剂（如N - 乙基马来酰亚胺（NEM））的毒性作用方面发挥作用。大肠杆菌的KefC基因已被克隆并进行了DNA测序。已经分离出一些影响KefC系统钾离子保留的kefC突变体，并且所有这些突变体都保留了被NEM激活的特性。发现克隆的kefC可以抑制两个这样的突变体kefC121和kefC103的表型。对这一现象的分析表明，这种抑制作用对KefC系统具有特异性，但来自克雷伯氏菌和欧文氏菌的克隆kefC也可以介导对突变体表型的抑制。大肠杆菌基因的质粒构建体中，克隆基因的表达减弱，但显示出诱导抑制突变体表型的能力。KefC'-'LacZ杂合蛋白插入膜中，但不抑制突变体表型。克隆的kefC不抑制与kefC121等位基因表现出相似表型的突变kefB等位基因。这些数据表明，抑制作用不太可能源于蛋白质的突变形式被排除在膜外。此外，携带突变体和克隆的野生型等位基因的菌株中，NEM激活的钾离子外流比单独在细胞中存在任何一个等位基因时都要快，这表明两种形式的蛋白质都插入到了膜中。本文根据KefC蛋白的模型对这些数据进行了讨论，该模型认为该蛋白由一个或多个在膜中相互作用的相同亚基组成。
