Transposon mutations in the 5' end of glnD, the gene for a nitrogen regulatory sensor, that suppress the osmosensitive phenotype caused by otsBA lesions in Escherichia coli.
GlnD of Escherichia coli is a bifunctional signal-transducing enzyme (102.4 kDa) which uridylylates the allosteric regulatory protein PII and deuridylylates PII-UMP in response to growth with nitrogen excess or limitation, respectively. GlnD catalyzes these reactions in response to high or low levels of cytoplasmic glutamine, respectively, and indirectly directs the expression of nitrogen-regulated genes, e.g., the glnK- amtB operon. We report that chromosomal mini-Tn10 insertions situated after nucleotide number 997 or 1075 of glnD partially suppressed the osmosensitive phenotype of DeltaotsBA or otsA::Tn10 mutations (defective osmoregulatory trehalose synthesis). Strains carrying these glnD::mini-Tn10 mutations either completely repressed the expression of trp::(glnKp-lacZ) or induced this reporter system to nearly 60% of the wild-type glnD level in response to nitrogen availability, an essentially normal response. This was in contrast to the much-studied glnD99::Tn10 mutation, which carries its insertion in the 3' end of the gene, causes a complete repression of glnKp-lacZ expression under all growth conditions, and also confers leaky glutamine auxotrophy. When expressed from the Pm promoter in plasmid constructs, the present glnD mutations produced proteins with an apparent mass of 39 or 42 kDa. These proteins were deduced to comprise 344 or 370 N-terminal residues, respectively, harboring the known nucleotidyltransferase domain of GlnD, plus a common C-terminal addition of 12 residues encoded by IS10. They lacked three other domains of GlnD. Apparently, the transferase domain by itself enabled the cells to catalyze the uridylylation reaction and direct nitrogen-regulated gene expression. Our data indicate that there exists a link between osmotic stress and the nitrogen response.[1]References
- Transposon mutations in the 5' end of glnD, the gene for a nitrogen regulatory sensor, that suppress the osmosensitive phenotype caused by otsBA lesions in Escherichia coli. Tøndervik, A., Torgersen, H.R., Botnmark, H.K., Strøm, A.R. J. Bacteriol. (2006) [Pubmed]
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