Mutations altering the N-terminal receiver domain of NRI (NtrC) That prevent dephosphorylation by the NRII-PII complex in Escherichia coli.
The phosphorylated form of NRI is the transcriptional activator of nitrogen-regulated genes in Escherichia coli. NRI approximately P displays a slow autophosphatase activity and is rapidly dephosphorylated by the complex of the NRII and PII signal transduction proteins. Here we describe the isolation of two mutations, causing the alterations DeltaD10 and K104Q in the receiver domain of NRI, that were selected as conferring resistance to dephosphorylation by the NRII-PII complex. The mutations, which alter highly conserved residues near the D54 site of phosphorylation in the NRI receiver domain, resulted in elevated expression of nitrogen-regulated genes under nitrogen-rich conditions. The altered NRI receiver domains were phosphorylated by NRII in vitro but were defective in dephosphorylation. The DeltaD10 receiver domain retained normal autophosphatase activity but was resistant to dephosphorylation by the NRII-PII complex. The K104Q receiver domain lacked both the autophosphatase activity and the ability to be dephosphorylated by the NRII-PII complex. The properties of these altered proteins are consistent with the hypothesis that the NRII-PII complex is not a true phosphatase but rather collaborates with NRI approximately P to bring about its dephosphorylation.
Department of Biological Chemistry, University of Michigan Medical School, 1301 E. Catherine, Ann Arbor, MI 48109-0606, USA.
Augen A
AA
Pioszak
Alexander J
AJ
Ninfa
D000222:Adaptation, Physiological; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D003470:Culture Media / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D009154:Mutation; D009586:Nitrogen Fixation / Q000235:genetics; D051922:PII Nitrogen Regulatory Proteins; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D017384:Sequence Deletion; D015398:Signal Transduction; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism*
D001426:Bacterial Proteins; D003470:Culture Media; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D051922:PII Nitrogen Regulatory Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C497876:glnG protein, E coli; D011494:Protein Kinases; C055699:protein kinase-phosphatase NTRB; D010749:Phosphoprotein Phosphatases