The stringent response enables metabolic adaptation of bacteria under stress conditions and is governed by RelA/SpoT Homolog (RSH)-type enzymes. Long RSH-type enzymes encompass an N-terminal domain (NTD) harboring the second messenger nucleotide (p)ppGpp hydrolase and synthetase activity and a stress-perceiving and regulatory C-terminal domain (CTD). CTD-mediated binding of Rel to stalled ribosomes boosts (p)ppGpp synthesis. However, how the opposing activities of the NTD are controlled in the absence of stress was poorly understood. Here, we demonstrate on the RSH-type protein Rel that the critical regulative elements reside within the TGS (ThrRS, GTPase, and SpoT) subdomain of the CTD, which associates to and represses the synthetase to concomitantly allow for activation of the hydrolase. Furthermore, we show that Rel forms homodimers, which appear to control the interaction with deacylated-tRNA, but not the enzymatic activity of Rel. Collectively, our study provides a detailed molecular view into the mechanism of stringent response repression in the absence of stress.

严格的响应使细菌能够在压力条件下进行代谢适应，并受RelA / SpoT同源（RSH）型酶控制。长RSH型酶包含一个N末端域（NTD），该域带有第二个信使核苷酸（p）ppGpp水解酶和合成酶活性，以及​​一个压力感知和调节性C末端域（CTD）。CTD介导的Rel与停滞的核糖体结合可增强（p）ppGpp合成。但是，人们对在没有压力的情况下如何控制NTD的相反活动知之甚少。在这里，我们在RSH型蛋白Rel上证明了关键的调控元件位于CTD的TGS（ThrRS，GTPase和SpoT）子域内，该子域与该合成酶相关联并抑制该合成酶，从而可以激活水解酶。此外，我们证明Rel会形成同二聚体，似乎控制与脱酰tRNA的相互作用，但不控制Rel的酶促活性。总的来说，我们的研究提供了在没有压力的情况下严格反应抑制机制的详细分子观点。