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Isolation and characterization of Tn5-induced NADPH-glutamate synthase (GOGAT-) mutants of Azorhizobium sesbaniae ORS571 and cloning of the corresponding glt locus

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Summary

Random Tn5 mutagenesis was used to isolate two independent Azorhizobium sesbaniae ORS571 mutants disturbed in ammonium assimilation (Asm-). Both Asm- mutant strains were shown to lack NADPH-glutamate synthase (NADPH-GOGAT) activity and to carry Tn5 insertions ca. 1.5 kb apart in the ORS571 chromosome. The Tn5-containing region of one of the GOGAT- mutant strains was cloned in pACYC184 and used to identify the wild-type glt (GOGAT) locus in a phage clone bank of ORS571. The cloned region was shown to have DNA homology with the Escherichia coli glt locus and to complement the Asm- phenotype of E. coli and ORS571 GOGAT- strains. The ORS571 GOGAT- mutations were found to interfere with free-living as well as symbiotic nitrogen fixation. Expression of ORS571 NADPH-GOGAT activity was shown to be independent of the nitrogen regulation (ntr) system.

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Authors and Affiliations

  1. Max-Planck-Institut für Züchtungsforschung, D-5000, Köln 30, Germany

    U. Hilgert, J. Schell & F. J. de Bruijn

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  1. U. Hilgert
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  2. J. Schell
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  3. F. J. de Bruijn
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Communicated by H. Saedler

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Hilgert, U., Schell, J. & de Bruijn, F.J. Isolation and characterization of Tn5-induced NADPH-glutamate synthase (GOGAT-) mutants of Azorhizobium sesbaniae ORS571 and cloning of the corresponding glt locus. Molec Gen Genet 210, 195–202 (1987). https://doi.org/10.1007/BF00325684

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  • DOI: https://doi.org/10.1007/BF00325684

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