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Biochemical and genetic analysis of the nifHDKE region of Rhizobium ORS571

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Summary

Deletions and Tn5 insertions were obtained in a cloned 10 kb BamHI-BglII fragment carrying the nifHDKE region of Rhizobium ORS571 and were recombined into the host genome. Genetic analysis of the mutants, comparison of polypeptides synthesized under conditions of repression and depression of N2 fixation, and biochemical complementation of crude extracts were performed. All Nif- mutants were also Fix-. Three transcription units were identified, nifHDK, nifE and a new nif locus adjacent to nifE; no nif locus was found in the immediate vicinity upstream of nifH. Fifteen polypeptides synthesized under conditions of N2 fixation were characterized by two-dimensional gel electrophoresis. Ten of them are likely to be nif products and polypeptides encoded by nifH, nifD, nifK and tentatively nifE were identified. Physiological and biochemical evidence for the functioning of the second copy of nifH is reported. Nitrogenase component 2 synthesized by this copy could not be differentiated from component 2 synthesized in the wild-type strain. When the function of nifH copy 1 was abolished, the amount of component 2 synthesized was about 30% of that synthesized in the wild-type strain.

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Author notes

  1. Patrice Denèfle

    Present address: Génética, 160 Quai de Polangis, 94340, Joinville le Pont, France

  2. Anil Kush

    Present address: DST Unit of Genetic Engineering, JNU, 110067, New-Delhi, India

Authors and Affiliations

  1. Unité de Physiologie Cellulaire, Départment des Biotechnologies, Institut Pasteur, 25 rue du Dr. Roux, F-75724, Paris Cedex 15, France

    Patrice Denèfle, Anil Kush, Françoise Norel, Annick Paquelin & Claudine Elmerich

Authors
  1. Patrice Denèfle
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  2. Anil Kush
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  3. Françoise Norel
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  4. Annick Paquelin
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  5. Claudine Elmerich
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Communicated by J. Schell

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Denèfle, P., Kush, A., Norel, F. et al. Biochemical and genetic analysis of the nifHDKE region of Rhizobium ORS571. Mol Gen Genet 207, 280–287 (1987). https://doi.org/10.1007/BF00331590

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  • DOI: https://doi.org/10.1007/BF00331590

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