The Rhizobium leguminosarum bv. viciae VF39 {gamma}-aminobutyrate (GABA) aminotransferase gene (gabT) is induced by GABA and highly expressed in bacteroids

Ökologie des Bodens, Botanisches Institut, RWTH-Aachen, Worringerweg 1, 52056 Aachen, Germany 1 Division of Microbiology, School of AMS, University of Reading, Whiteknights, Reading RG6 6AJ, UK 2 Author for correspondence: Jürgen Prell. Tel: +49 241 8026647. Fax: +49 241 8022637. e-mail: prell{at}bio1.rwth-aachen.de A Rhizobium leguminosarum bv. viciae VF39 gene ( gabT ) encoding a -aminobutyrate (GABA) aminotransferase was identified, cloned and characterized. This gene is thought to be involved in GABA metabolism via the GABA shunt pathway, a theoretical bypass of the 2-oxoglutarate dehydrogenase complex. Mutants in gabT are still able to grow on GABA as a sole carbon and nitrogen source. 2-Oxoglutarate-dependent GABA aminotransferase activity is absent in these mutants, while pyruvate-dependent activity remains unaffected. This indicates that at least two enzymes with different substrate specifities are involved in the GABA metabolism of R. leguminosarum bv. viciae VF39. The gabT promoter was cloned into a newly constructed, stable promoter-probe vector pJP2, suitable for the study of transcriptional GUS fusions in free-living bacteria and during symbiosis. Under free-living conditions the gabT promoter is induced by GABA and repressed by succinate. Transcriptional regulation is mediated by GabR in a repressor-like manner. During symbiosis with the pea host plant gabT is induced and highly expressed in the symbiotic zone. Nodules induced by gabT mutants, however, are still effective in nitrogen fixation. Keywords: carbon metabolism, GABA shunt, symbiotic expression, promoter-probe vector, pJP2 Abbreviations: GABA, -aminobutyrate; GUS, ß- D -glucuronidase; PEP, phosphoenolpyruvate; PNP, p -nitrophenol; SSDH, succinate semialdehyde dehydrogenase; TCA, tricarboxylic acid; X-Gal, 5-bromo-4-chloro-3-indolyl ß- D -galactopyranoside; X-Glc, 5-bromo-4-chloro-3-indolyl ß- D -glucuronic acid The GenBank accession number for the sequence determined in this work is AF335502 .