Construction and characterization a fixN mutant strain of Herbaspirillum seropedicae
Maria Augusta Schmidt, Emanuel Maltempi de Souza, Roseli Wassem, Liu Un Rigo, Geoffrey Yates, Fábio de Oliveira Pedrosa and Rose Adele Monteiro.
Universidade Federal do Paraná, Setor de Ciências Biológicas, Departamento de Bioquímica e Biologia Molecular, Núcleo de Fixação Biológica de Nitrogênio
Herbaspirillum seropedicae is an endophytic diazotrophic microrganism which associates with several gramineous crops of economic importance. The nitrogen fixed by this organism can probably be utilized by plants, therefore, it is important to understand the physiology and genetics of the nitrogen-fixing process. Cytochrome oxidase cbb3, coded by the fixNOQP gene, is necessary for microaerobic respiration in rhizobia and is essential for nitrogen fixation in these organisms. Mutation of the fixN or fixO genes of Bradyrhizobium japonicum produced strains capable of nodulating soybean but no nodules had nitrogenase activity, suggesting that the cytochrome oxidase cbb3 may be involved in the transport of electrons to nitrogenase. The operon fixNOQP of H. seropedicae was identified by the Genome Sequencing Project of Herbaspirillum seropedicae (GENOPAR).
The transposon EZ:TN <TET-1>, containing a gene for tetracycline resistance, was inserted into the fixN gene of H. seropedicae strain SmR1 by homologous recombination to produce the strain RAM21. The activity of nitrogenase of this mutant strain was 40-50% that of the wild type strain. These results suggest that cytochrome oxidase cbb3 is probably required for maximal nitrogenase activity in H. seropedicae.
Supported by CNPq/PIBIC, CAPES and Fundação Araucária.
|