Lethality of glnD null mutations in Azotobacter vinelandii is suppressible by prevention of glutamine synthetase adenylylation

Rita Colnaghi, Paul Rudnick, Luhong He, Andrew Green, Dalai Yan, Ethan Larson, Christina Kennedy

Research output: Contribution to journalArticle

18 Scopus citations

Abstract

GlnD is a pivotal protein in sensing intracellular levels of fixed nitrogen and has been best studied in enteric bacteria, where it reversibly uridylylates two related proteins, PII and GlnK. The uridylylation state of these proteins determines the activities of glutamine synthase (GS) and NtrC. Results presented here demonstrate that glnD is an essential gene in Azotobacter vinelandii. Null glnD mutations were introduced into the A. vinelandii genome, but none could be stably maintained unless a second mutation was present that resulted in unregulated activity of GS. One mutation, gln-71, occurred spontaneously to give strain MV71, which failed to uridylylate the GlnK protein. The second, created by design, was glnAY407F (MV75), altering the adenylylation site of GS. The gln-71 mutation is probably located in glnE, encoding adenylyltransferase, because introducing the Escherichia coli glnE gene into MV72, a glnD+ derivative of MV71, restored the regulation of GS activity. GlnK-UMP is therefore apparently required for GS to be sufficiently deadenylylated in A. vinelandii for growth to occur. The ΔglnD GSc isolates were Nif-, which could be corrected by introducing a nifL mutation, confirming a role for GlnD in mediating nif gene regulation via some aspect of the NifL/NifA interaction. MV71 was unexpectedly NtrC+, suggesting that A. vinelandii NtrC activity might be regulated differently than in enteric organisms.

Original languageEnglish (US)
Pages (from-to)1267-1276
Number of pages10
JournalMicrobiology
Volume147
Issue number5
DOIs
StatePublished - May 2001

Keywords

  • GlnD
  • Nif gene regulation
  • Nitrogen assimilation
  • Nitrogen fixation
  • NtrC

ASJC Scopus subject areas

  • Microbiology

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