Jones H M, Gunsalus R P
J Bacteriol. 1987 Jul;169(7):3340-9. doi: 10.1128/jb.169.7.3340-3349.1987.
The fumarate reductase enzyme complex, encoded by the frdABCD operon, allows Escherichia coli to utilize fumarate as a terminal electron acceptor for anaerobic oxidative phosphorylation. To analyze the expression of fumarate reductase, protein and operon fusions were constructed between the frdA and the lacZ genes and introduced onto the E. coli chromosome at the lambda attachment site. Expression of beta-galactosidase from either fusion was increased 10-fold during anaerobic versus aerobic cell growth, increased an additional 1.5-fold by the presence of fumarate, the substrate, and decreased 23-fold by nitrate, a preferred electron acceptor. The addition of trimethylamine-N-oxide as an electron acceptor did not significantly alter frdA'-'lacZ expression. Control of frd operon expression is therefore exerted at the transcriptional level in response to the availability of the electron acceptors oxygen, fumarate, and nitrate. Anaerobic induction of frdA'-'lacZ expression was impaired in an fnr mutant and was restored when the fnr+ gene was provided in trans, thus establishing that the fnr gene product, Fnr, is responsible for the anaerobic activation of frd operon expression. Nitrate repression of frdA'-'lacZ expression was observed under either aerobic or anaerobic cell growth conditions in both wild-type and fnr mutant strains, demonstrating that the mechanism for nitrate repression is independent of nitrate respiration and oxygen control imparted by Fnr. Studies performed with a fnr'-'lacZ protein fusion confirmed that the fnr gene is expressed both aerobically and anaerobically. A model is proposed for the regulation of frdABCD operon expression in response to the availability of the alternate terminal electron acceptors oxygen, nitrate, and fumarate.
由frdABCD操纵子编码的延胡索酸还原酶复合体,使大肠杆菌能够利用延胡索酸作为厌氧氧化磷酸化的末端电子受体。为了分析延胡索酸还原酶的表达,构建了frdA与lacZ基因之间的蛋白质和操纵子融合体,并将其导入大肠杆菌染色体的λ附着位点。在厌氧与需氧细胞生长过程中,来自任一融合体的β-半乳糖苷酶表达增加了10倍,由于底物延胡索酸的存在又额外增加了1.5倍,而由于优先电子受体硝酸盐的存在则降低了23倍。添加三甲胺-N-氧化物作为电子受体并没有显著改变frdA'-'lacZ的表达。因此,frd操纵子的表达调控是在转录水平上响应电子受体氧气、延胡索酸和硝酸盐的可用性而进行的。在fnr突变体中,frdA'-'lacZ表达的厌氧诱导受损,当反式提供fnr+基因时恢复,从而确定fnr基因产物Fnr负责frd操纵子表达的厌氧激活。在野生型和fnr突变体菌株的需氧或厌氧细胞生长条件下,均观察到frdA'-'lacZ表达的硝酸盐抑制作用,这表明硝酸盐抑制机制独立于Fnr赋予的硝酸盐呼吸和氧气控制。用fnr'-'lacZ蛋白质融合体进行的研究证实,fnr基因在需氧和厌氧条件下均有表达。针对frdABCD操纵子表达响应交替末端电子受体氧气、硝酸盐和延胡索酸的可用性的调控提出了一个模型。