School of Biological Sciences and Institute of Microbiology, Seoul National University, Seoul, 08826, Korea.
Research Institute of Basic Science, Seoul National University, Seoul, 08826, Korea.
Nucleic Acids Res. 2023 Jun 23;51(11):5432-5448. doi: 10.1093/nar/gkad220.
Phosphorylation state-dependent interactions of the phosphoenolpyruvate (PEP):carbohydrate phosphotransferase system (PTS) components with transcription factors play a key role in carbon catabolite repression (CCR) by glucose in bacteria. Glucose inhibits the PTS-dependent transport of fructose and is preferred over fructose in Vibrio cholerae, but the mechanism is unknown. We have recently shown that, contrary to Escherichia coli, the fructose-dependent transcriptional regulator FruR acts as an activator of the fru operon in V. cholerae and binding of the FruR-fructose 1-phosphate (F1P) complex to an operator facilitates RNA polymerase (RNAP) binding to the fru promoter. Here we show that, in the presence of glucose, dephosphorylated HPr, a general PTS component, binds to FruR. Whereas HPr does not affect DNA-binding affinity of FruR, regardless of the presence of F1P, it prevents the FruR-F1P complex from facilitating the binding of RNAP to the fru promoter. Structural and biochemical analyses of the FruR-HPr complex identify key residues responsible for the V. cholerae-specific FruR-HPr interaction not observed in E. coli. Finally, we reveal how the dephosphorylated HPr interacts with FruR in V. cholerae, whereas the phosphorylated HPr binds to CcpA, which is a global regulator of CCR in Bacillus subtilis and shows structural similarity to FruR.
磷酸烯醇丙酮酸（PEP）：碳水化合物磷酸转移酶系统（PTS）成分与转录因子的磷酸化状态依赖性相互作用在细菌的碳分解代谢物阻遏（CCR）中起着关键作用。葡萄糖抑制 PTS 依赖的果糖运输，并且在霍乱弧菌中优先于果糖，但机制尚不清楚。我们最近表明，与大肠杆菌相反，果糖依赖性转录调节剂 FruR 在霍乱弧菌中作为 fru 操纵子的激活剂起作用，并且 FruR-果糖 1-磷酸（F1P）复合物与操纵子的结合促进 RNA 聚合酶（RNAP）与 fru 启动子结合。在这里，我们表明，在葡萄糖存在下，一般 PTS 成分去磷酸化 HPr 与 FruR 结合。尽管 HPr 不影响 FruR 的 DNA 结合亲和力，无论是否存在 F1P，它都阻止 FruR-F1P 复合物促进 RNAP 与 fru 启动子的结合。对 FruR-HPr 复合物的结构和生化分析确定了霍乱弧菌中观察到的关键残基，这些残基负责 FruR-HPr 相互作用，而在大肠杆菌中未观察到这种相互作用。最后，我们揭示了去磷酸化的 HPr 如何与霍乱弧菌中的 FruR 相互作用，而磷酸化的 HPr 与 CcpA 结合，CcpA 是枯草芽孢杆菌中 CCR 的全局调节剂，并且与 FruR 具有结构相似性。
