Lux R, Jahreis K, Bettenbrock K, Parkinson J S, Lengeler J W
Fachbereich Biologie/Chemie, Universität Osnabrück, Germany.
Proc Natl Acad Sci U S A. 1995 Dec 5;92(25):11583-7. doi: 10.1073/pnas.92.25.11583.
Chemotactic responses in Escherichia coli are typically mediated by transmembrane receptors that monitor chemoeffector levels with periplasmic binding domains and communicate with the flagellar motors through two cytoplasmic proteins, CheA and CheY. CheA autophosphorylates and then donates its phosphate to CheY, which in turn controls flagellar rotation. E. coli also exhibits chemotactic responses to substrates that are transported by the phosphoenolpyruvate (PEP)-dependent carbohydrate phosphotransferase system (PTS). Unlike conventional chemoreception, PTS substrates are sensed during their uptake and concomitant phosphorylation by the cell. The phosphoryl groups are transferred from PEP to the carbohydrates through two common intermediates, enzyme I (EI) and phosphohistidine carrier protein (HPr), and then to sugar-specific enzymes II. We found that in mutant strains HPr-like proteins could substitute for HPr in transport but did not mediate chemotactic signaling. In in vitro assays, these proteins exhibited reduced phosphotransfer rates from EI, indicating that the phosphorylation state of EI might link the PTS phospho-relay to the flagellar signaling pathway. Tests with purified proteins revealed that unphosphorylated EI inhibited CheA autophosphorylation, whereas phosphorylated EI did not. These findings suggest the following model for signal transduction in PTS-dependent chemotaxis. During uptake of a PTS carbohydrate, EI is dephosphorylated more rapidly by HPr than it is phosphorylated at the expense of PEP. Consequently, unphosphorylated EI builds up and inhibits CheA autophosphorylation. This slows the flow of phosphates to CheY, eliciting an up-gradient swimming response by the cell.
大肠杆菌中的趋化反应通常由跨膜受体介导，这些受体通过周质结合结构域监测化学效应物水平，并通过两种细胞质蛋白CheA和CheY与鞭毛马达进行通信。CheA进行自身磷酸化，然后将其磷酸基团转移给CheY，CheY进而控制鞭毛的旋转。大肠杆菌对由磷酸烯醇丙酮酸（PEP）依赖性碳水化合物磷酸转移酶系统（PTS）转运的底物也表现出趋化反应。与传统的化学感受不同，PTS底物在细胞摄取和伴随磷酸化的过程中被感知。磷酸基团通过两种常见中间体，即酶I（EI）和磷酸组氨酸载体蛋白（HPr），从PEP转移到碳水化合物上，然后再转移到糖特异性的酶II上。我们发现，在突变菌株中，类HPr蛋白可以在转运过程中替代HPr，但不介导趋化信号传导。在体外试验中，这些蛋白显示出从EI的磷酸转移速率降低，这表明EI的磷酸化状态可能将PTS磷酸传递与鞭毛信号通路联系起来。对纯化蛋白的测试表明，未磷酸化的EI抑制CheA自身磷酸化，而磷酸化的EI则不会。这些发现提出了以下PTS依赖性趋化作用中的信号转导模型。在摄取PTS碳水化合物期间，HPr使EI去磷酸化的速度比PEP使其磷酸化的速度更快。因此，未磷酸化的EI积累并抑制CheA自身磷酸化。这减缓了磷酸基团向CheY的流动，引发细胞向上梯度游动的反应。
