Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Yuseong-gu, Daejeon, Republic of Korea.
J Bacteriol. 2012 Apr;194(8):1979-88. doi: 10.1128/JB.06062-11. Epub 2012 Feb 10.
Glyoxal is toxic and mutagenic α-oxoaldehyde generated in vivo as an oxidation by-product of sugar metabolism. We selected glyoxal-resistant mutants from an Escherichia coli strain lacking major glyoxal-detoxifying genes, gloA and yqhD, by growing cells in medium containing a lethal concentration of glyoxal. The mutants carried diverse genomic rearrangements, such as multibase deletions and recombination, in the upstream region of the yafB gene, encoding an aldo-keto reductase. Since these genomic lesions create transcriptional fusions of the yafB gene to the upstream rrn regulon or eliminate a negative regulatory site, the mutants generally enhanced an expression of the yafB gene. Glyoxal resistances of the mutants are correlated with the levels of yafB transcripts as well as the activities of aldo-keto reductase. An overproduction of YafB in the glyoxal-resistant mutant lacking the putative NsrR-binding site provides evidence that the yafB gene is negatively regulated by this protein. We also observed that the expression of yafB is enhanced with an increased concentration of glyoxal as well as a mutation in the fnr gene, encoding a putative regulator. The bindings of NsrR and Fnr to the yafB promoter were also demonstrated by gel mobility shift assays.
乙二醛是一种有毒的、致突变的α-氧代醛，作为糖代谢的氧化副产物在体内生成。我们从缺乏主要乙二醛解毒基因 gloA 和 yqhD 的大肠杆菌菌株中选择了耐乙二醛的突变体，方法是在含有致死浓度乙二醛的培养基中培养细胞。这些突变体在编码醛酮还原酶的 yafB 基因的上游区域携带了多种基因组重排，如多碱基缺失和重组。由于这些基因组病变导致 yafB 基因与上游 rrn 调控子发生转录融合或消除了负调控位点，因此突变体通常会增强 yafB 基因的表达。突变体的乙二醛抗性与 yafB 转录本的水平以及醛酮还原酶的活性相关。在缺乏假定的 NsrR 结合位点的耐乙二醛突变体中过量表达 YafB 提供了证据，表明该 yafB 基因受到该蛋白的负调控。我们还观察到，yafB 的表达随着乙二醛浓度的增加以及编码假定调节剂的 fnr 基因的突变而增强。凝胶迁移率变动分析也证明了 NsrR 和 Fnr 对 yafB 启动子的结合。
