Role of glutamine synthetase in phenazine antibiotic production by Pantoea agglomerans Eh1087

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Pantoea agglomerans strain Eh1087 produces the phenazine antibiotic D-alanylgriseoluteic acid. A glutamine auxotroph harboring an insertion in a putative glnA gene was obtained by transposon-mutagenesis of Eh1087 that produced less D-alanylgriseoluteic acid than the parental strain (strain Eh7.1). Cosmids encoding the Eh1087 glnA were isolated by their ability to complement the mutant for prototrophy. The role of the Eh1087 glnA locus was functionally confirmed by complementation of an Escherichia coli glnA mutant. Analysis of the nucleotide and deduced amino acid sequences of the Eh1087 glnA gene indicated a high degree of similarity to the glnA genes and glutamine synthetase enzymes of other Enterobacteriaceae. Isotopic labelling experiments with 15N-labelled ammonium sulfate demonstrated that wild-type Eh1087 incorporated 15N into griseoluteic acid more readily than the glnA mutant Eh7.1. We conclude that the 2 nitrogens in the phenazine nucleus originate from glutamine and the intracellular glutamine synthesized by Eh1087 is a source of the phenazine nucleus nitrogens even in glutamine-rich environments.Key words: phenazine, Pantoea, Erwinia, glutamine synthetase, biosynthesis.

La souche Eh1087 de Pantoea agglomerans produit l'antibiotique de type phénazine acide D-alanylgriseolutéique (AGA). Un auxotrophe de la glutamine renfermant une insertion dans le gène putatif glnA a été obtenu par mutagénèse par transposon de Eh1087, et celui-ci produisait moins de AGA que la souche parentale (souche Eh7.1). Des cosmides codant le gène glnA de Eh1087 ont été isolés grâce à leur capacité à complémenter le mutant pour la phototrophie. Le rôle du locus Eh1087 glnA a été confirmés fonctionnellement par complémentation d'un mutant glnA de E. coli. Une analyse des séquences de nucléotides et d'acides aminés déduits du gène Eh1087 glnA ont signalé un haut degré de similitude aux gènes glnA et aux enzymes glutamine synthétase d'autres Enterobacteriaceae. Des expériences de marquage isotopique avec du sulfate d'ammonium 15N ont démontré que la Eh1087 sauvage incorporait du 15N plus facilement que le mutant glnA, Eh7.1. Nous en déduisons que les deux azotes dans le noyau phénazine proviennent de la glutamine, et que la glutamine intracellulaire synthétisée par Eh1087 est une source d'atomes d'azote pour le noyau phénazine même dans des environnements riches en glutamine.Mots clés : phénazine, Pantoea, Erwinia, glutamine synthétase, biosynthèse.[Traduit par la Rédaction]

Document Type: Research Article

Publication date: October 1, 2004

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  • Published since 1954, this monthly journal contains new research in the field of microbiology including applied microbiology and biotechnology; microbial structure and function; fungi and other eucaryotic protists; infection and immunity; microbial ecology; physiology, metabolism and enzymology; and virology, genetics, and molecular biology. It also publishes review articles and notes on an occasional basis, contributed by recognized scientists worldwide.
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