THE ESCHERICHIA-COLI PII SIGNAL-TRANSDUCTION PROTEIN IS ACTIVATED UPON BINDING 2-KETOGLUTARATE AND ATP
THE ESCHERICHIA-COLI PII SIGNAL-TRANSDUCTION PROTEIN IS ACTIVATED UPON BINDING 2-KETOGLUTARATE AND ATP
复制标题
DOI：
10.1074/jbc.270.30.17797
复制
发表时间：
1995-07-28
影响因子：
4.8
通讯作者：
NINFA, AJ
中科院分区：
文献类型：
--
作者：
KAMBEROV, ES;ATKINSON, MR;NINFA, AJ
在大肠杆菌中，氮对转录的调节需要感受细胞内的氮状态，并控制转录激活剂NRI的去磷酸化，这种去磷酸化是在存在可分离的PII蛋白的情况下由双功能激酶/磷酸酶NRII催化的。PII刺激NRII磷酸酶活性的能力是由一个信号转导的尿苷酰转移酶/尿苷酰去氢酶(Utase/UR)调节的，它在氮饥饿条件下将PII转化为PB-UMP；这种修饰阻止了PII刺激类似于P的NRI的去磷酸化。我们使用纯化的组分来检测小分子与PII的结合，小分子对PII刺激NRII磷酸酶活性的影响，PII在固定化NRII上的滞留，以及Utase/UR酶对PII尿苷基化的调节。我们的结果表明，PII在与ATP和2-酮戊二酸或谷氨酸结合时被激活，并且PII的配体形式与固定化的NRII结合得更好。我们还证明了抑制尿苷基转移酶活性所需的谷氨酰胺浓度与2-酮戊二酸的浓度无关。我们假设，在大肠杆菌中的氮感觉涉及到Utase/UR蛋白对谷氨酰胺的分离测定和PII蛋白对2-酮戊二酸的分离测定。
Nitrogen regulation of transcription in Escherichia coli requires sensation of the intracellular nitrogen status and control of the dephosphorylation of the transcriptional activator NRI similar to P. This dephosphorylation is catalyzed by the bifunctional kinase/phosphatase NRII in the presence of the dissociable PII protein. The ability of PII to stimulate the phosphatase activity of NRII is regulated by a signal transducing uridylyltransferase/uridylyl-removing enzyme (UTase/UR), which converts PII to PB-UMP under conditions of nitrogen starvation; this modification prevents PII from stimulating the dephosphorylation of NRI similar to P. We used purified components to examine the binding of small molecules to PII, the effect of small molecules on the stimulation of the NRII phosphatase activity by PII, the retention of PII on immobilized NRII, and the regulation of the uridylylation of PII by the UTase/UR enzyme. Our results indicate that PII is activated upon binding ATP and either 2-ketoglutarate or glutamate, and that the liganded form of PII binds much better to immobilized NRII. We also demonstrate that the concentration of glutamine required to inhibit the uridylyltransferase activity is independent of the concentration of 2-ketoglutarate present. We hypothesize that nitrogen sensation in E. coli involves the separate measurement of glutamine by the UTase/UR protein and 2-ketoglutarate by the PII protein.