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Characterization of an Azospirillum brasilense Sp7 gene homologous to Alcaligenes eutropbus pbbB and to Rbizobium meliloti nodG

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Summary

A 4 kb SalI fragment from Azospirillum brasilense Sp7 that shares homology with a 6.8 kb EcoRI fragment carrying nodGEFH and part of nodP of Rhizobium meliloti 41 was cloned in pUC18 to yield pAB503. The nucleotide sequence of a 2 kb SalI-SmaI fragment of the pAB503 insert revealed an open reading frame, named ORF3, encoding a polypeptide sharing 40% identity with R. mehloti NodG. The deduced polypeptide also shared 60% identity with the Alcaligenes eutrophus NADPH-dependent acetoacetyl-CoA (AA-CoA) reductase, encoded by the pbbB gene and involved in poly-β-hydroxybutyrate (PHB) synthesis. Northern blot analysis and promoter extension map** indicated that ORF3 is expressed as a monocistronic operon from a promoter that resembles the Escherichia coli σ70 consensus promoter. An ORF3-lacZ translational fusion was constructed and was very poorly expressed in E. coli, but was functional and constitutively expressed in Azospirillum. Tn5-Mob insertions in ORF3 did not affect growth, nitrogen fixation, PHB synthesis or NAD(P)H-linked AA-CoA reductase activity. An ORF3 DNA sequence was used to probe total DNA of several Azospirillum strains. No ORF3 homologues were found in A. irakense, A. amazonense, A. halopraeferens or in several A. lipoferum strains.

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  1. Unité de Physiologic Cellulaire, URA 1300 CNRS, Département des Biotechnologies, Institut Pasteur, 25 rue du Dr. Roux, F-75724, Paris Cedex 15, France

    Claire Vieille & Claudine Elmerich

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  1. Claire Vieille
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  2. Claudine Elmerich
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Vieille, C., Elmerich, C. Characterization of an Azospirillum brasilense Sp7 gene homologous to Alcaligenes eutropbus pbbB and to Rbizobium meliloti nodG. Molec. Gen. Genet. 231, 375–384 (1992). https://doi.org/10.1007/BF00292706

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