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Microbiology 145 (1999), 2313-2322
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Microbiology (1999), 145, 2313-2322.
© 1999 Society for General Microbiology

Molecular Genetics of Streptomycetes

Nitrogen metabolism in Streptomyces coelicolor A3(2): modification of glutamine synthetase I by an adenylyltransferase

D. Finka,1, D. Falke1, W. Wohlleben1 and A. Engels1

Mikrobiologie/Biotechnologie, Eberhard-Karls-Universität Tübingen, Auf der Morgenstelle 28, D-72076 Tübingen, Germany1

Author for correspondence: A. Engels. Tel: +49 7071 29 76157. Fax: +49 7071 29 5979. e-mail: ae{at}biotech.uni-tuebingen.de

An internal adenylyltransferase gene (glnE) fragment from Streptomyces coelicolor was amplified using heterologous PCR primers derived from consensus motifs. The sequence had significant similarity to bacterial glnE genes, and included a motif typical of the C-terminal adenylyltransferase domain of GlnE. glnE from S. coelicolor lies on the AseI-C fragment of the chromosome and is localized near glnA (encoding glutamine synthetase I, GSI) and glnII (encoding GSII). To analyse the function of GlnE in S. coelicolor, glnE (S. coelicolor E4) and glnA (S. coelicolor HT107) gene replacement mutants were constructed. The GSI activity of the glnE mutant was not down-regulated after an ammonium shock. However, the GSI activity of the wild-type cells decreased to 60% of the original activity. The glnA mutant is not glutamine auxotrophic, but in the {gamma}-glutamyltransferase assay no GSI activity was detected in unshifted and shifted HT107 cells. By snake venom phosphodiesterase treatment the GSI activity in the wild-type can be reconstituted, whereas no alteration is observed in the E4 mutant. Additionally, the loss of short-term GSI regulation in the E4 mutant was accompanied by an increased glutamine:glutamate ratio.

Keywords: Streptomyces coelicolor, nitrogen metabolism, adenylyltransferase, glutamine synthetase

Abbreviations: GS, glutamine synthetase; SVPDE, snake venom phosphodiesterase

The EMBL accession number of the internal Streptomyces coelicolor glnE fragment is Y17736.

a Present address: Genetics Department, John Innes Centre, Colney Lane, Norwich NR4 7UH, UK.




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