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de Paula Soares C.,Federal University of Rio de Janeiro | Rodrigues E.P.,State University Londrina | de Paula Ferreira J.,Federal Rural University of Rio de Janeiro | Simoes Araujo J.L.,Genetics and Biochemistry Laboratory | And 3 more authors.
Archives of Microbiology | Year: 2014

TonB-dependent receptors in concert with the TonB–ExbB–ExbD protein complex are responsible for the uptake of iron and substances such as vitamin B12 in several bacterial species. In this study, Tn5 mutagenesis of the sugarcane endophytic bacterium Gluconacetobacter diazotrophicus led to the isolation of a mutant with a single Tn5-insertion in the promoter region of a tonB gene ortholog. This mutant, named Gdiaa31, displayed a reduced growth rate and a lack of response to iron availability when compared to the wild-type strain PAL5T. Several efforts to generate null-mutants for the tonB gene by insertional mutagenesis were without success. RT-qPCR analysis demonstrated reduced transcription of tonB in Gdiaa31 when compared to PAL5T. tonB transcription was inhibited in the presence of Fe3+ ions both in PAL5T and in Gdiaa31. In comparison with PAL5T, Gdiaa31 also demonstrated decreased nitrogenase activity and biofilm formation capability, two iron-requiring physiological characteristics of G. diazotrophicus. Additionally, Gdiaa31 accumulated higher siderophore levels in culture supernatant. The genetic complementation of the Gdiaa31 strain with a plasmid that carried the tonB gene including its putative promoter region (pPtonB) restored nitrogenase activity and siderophore accumulation phenotypes. These results indicate that the TonB complex has a role in iron/siderophore transport and may be essential in the physiology of G. diazotrophicus. © 2014, Springer-Verlag Berlin Heidelberg.

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