The Rhizobium leguminosarum bv. viciae glnD gene, encoding a uridylyltransferase/uridylyl-removing enzyme, is expressed in the root nodule but is notessential for nitrogen fixation

A Rhizobium leguminosarum by. viciae VF39 gene (glnD) encoding the uridylyl transferase/uridylyl-removing enzyme, which constitutes the sensory compone nt of the nitrogen regulation (ntr) system, was identified, cloned and char acterized. The deduced amino acid sequence contains the conserved active si te motif of the nucleotidyltransferase superfamily and is highly homologous to the glnD gene products of other bacterial species. Downstream of the VF 39 glnD resides an open reading frame with similarity to the Salmonella typ himurium virulence factor gene mviN. Mutation of the glnD gene abolished th e ability to use nitrate as a sole nitrogen source but not glutamine. In ad dition, neither uridylylation of P-II nor induction of the ntr-regulated gI nII gene (encoding glutamine synthetase II) under ammonium deficiency could be observed in mutant strains. This strongly suggests that glnD mutants ha rbour a permanently deuridylylated P-II protein and as a consequence are un able to activate transcription from NtrC-dependent promoters. The glnD gene itself is expressed constitutively, irrespective of the nitrogen content o f the medium. A functional GlnD protein is not essential for nitrogen fixat ion in R. leguminosarum by. viciae, but in situ detection of glnD expressio n in the symbiotic and infection zone of the root nodule and quantitative m easurements suggest that at least part of the ntr system functions in symbi osis. The results also indicate that the N-terminal part of GlnD is essenti al for the cell, as deletions in the 5'-region of the gene appear to be let hal and mutations possibly affecting the expression of the first half of th e protein have a significant effect on the vitality of the mutant strain.