Feng J, Atkinson M R, McCleary W, Stock J B, Wanner B L, Ninfa A J
Department of Biochemistry, Wayne State University School of Medicine, Detroit, Michigan 48201.
J Bacteriol. 1992 Oct;174(19):6061-70. doi: 10.1128/jb.174.19.6061-6070.1992.
Transcription of the Ntr regulon is controlled by the two-component system consisting of the response regulator NRI (NtrC) and the kinase/phosphatase NRII (NtrB), which both phosphorylates and dephosphorylates NRI. Even though in vitro transcription from nitrogen-regulated promoters requires phosphorylated NRI, NRII-independent activation of NRI also occurs in vivo. We show here that this activation likely involves acetyl phosphate; it is eliminated by mutations that reduce synthesis of acetyl phosphate and is elevated by a mutation expected to cause accumulation of acetyl phosphate. With purified components, we investigated the mechanism by which acetyl phosphate stimulates glutamine synthetase synthesis. Acetyl phosphate, carbamyl phosphate, and phosphoramidate but not ATP or phosphoenolpyruvate acted as substrates for the autophosphorylation of NRI in vitro. Phosphorylated NRI produced by this mechanism exhibited the properties associated with NRI phosphorylated by NRII, including the activated ATPase activity of the central domain of NRI and the ability to activate transcription from the nitrogen-regulated glutamine synthetase glnAp2 promoter.
Ntr 调控子的转录由双组分系统控制，该系统由应答调节因子 NRI（NtrC）和激酶/磷酸酶 NRII（NtrB）组成，二者均可使 NRI 磷酸化和去磷酸化。尽管体外氮调节启动子的转录需要磷酸化的 NRI，但 NRI 的 NRII 非依赖性激活在体内也会发生。我们在此表明，这种激活可能涉及乙酰磷酸；它可被减少乙酰磷酸合成的突变消除，并因预期会导致乙酰磷酸积累的突变而增强。利用纯化的组分，我们研究了乙酰磷酸刺激谷氨酰胺合成酶合成的机制。乙酰磷酸、氨基甲酰磷酸和氨基磷酸酯，但不是 ATP 或磷酸烯醇丙酮酸，在体外作为 NRI 自磷酸化的底物。通过这种机制产生的磷酸化 NRI 表现出与被 NRII 磷酸化的 NRI 相关的特性，包括 NRI 中央结构域的活化 ATP 酶活性以及激活氮调节的谷氨酰胺合成酶 glnAp2 启动子转录的能力。
