Isolamento e caracterização de bactérias fixadoras de nitrogênio associadas com a cultura do morango na região Serrana do Rio de Janeiro.

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Manual de soluções e reagentes da Embrapa Agrobiologia.

Solução de ácido acético a 10%.Solução de ácido acético a15%. Solução de ácido bórico a 1%. Soluções: Àcido sulfúrico a 5%. Água oxigenada a 3%. de azul de brotimol a 0,1%. de azul de brotimol a 0,5%. de azul de brotimol a 0,5% alcoolica p/v. Cloreto de sódio 2,5 %. Cloreto de sódio 10%. Cloreto de potássio. Dicromato de potássio. Edta de ferro. Fosfato de potássio Dibásico. Fosfato de potássio dibásico. Fosfato de potássio monobásico. fosfato de potássio monobásico. Hidroxido de potássio. Hidroxido de potássio. Hidroxido de de sódio. Molibdato de sódio dihidratado. Sulfato de magnésio. Sulfato de magnésio heptahidratado. Sulfato de potássio. Sulfato de manganês. Sulfato de ferro.bitstream/CNPAB-2010/27283/1/doc086.pd

Antioxidant pathways are up-regulated during biological nitrogen fixation to prevent ROS-induced nitrogenase inhibition in Gluconacetobacter diazotrophicus

Gluconacetobacter diazotrophicus, an endophyte isolated from sugarcane, is a strict aerobe that fixates N2. This process is catalyzed by nitrogenase and requires copious amounts of ATP. Nitrogenase activity is extremely sensitive to inhibition by oxygen and reactive oxygen species (ROS). However, the elevated oxidative metabolic rates required to sustain biological nitrogen fixation (BNF) may favor an increased production of ROS. Here, we explored this paradox and observed that ROS levels are, in fact, decreased in nitrogen-fixing cells due to the up-regulation of transcript levels of six ROS-detoxifying genes. A cluster analyses based on common expression patterns revealed the existence of a stable cluster with 99.8% similarity made up of the genes encoding the α-subunit of nitrogenase Mo–Fe protein (nifD), superoxide dismutase (sodA) and catalase type E (katE). Finally, nitrogenase activity was inhibited in a dose-dependent manner by paraquat, a redox cycler that increases cellular ROS levels. Our data revealed that ROS can strongly inhibit nitrogenase activity, and G. diazotrophicus alters its redox metabolism during BNF by increasing antioxidant transcript levels resulting in a lower ROS generation. We suggest that careful controlled ROS production during this critical phase is an adaptive mechanism to allow nitrogen fixation

Discovery and characterization of a sulfoquinovose mutarotase using kinetic analysis at equilibrium by exchange spectroscopy

Bacterial sulfoglycolytic pathways catabolize sulfoquinovose (SQ), or glycosides thereof, to generate a three-carbon metabolite for primary cellular metabolism and a three-carbon sulfonate that is expelled from the cell. Sulfoglycolytic operons encoding an Embden–Meyerhof–Parnas-like or Entner–Doudoroff (ED)-like pathway harbor an uncharacterized gene (yihR in Escherichia coli; PpSQ1_00415 in Pseudomonas putida) that is up-regulated in the presence of SQ, has been annotated as an aldose-1-epimerase and which may encode an SQ mutarotase. Our sequence analyses and structural modeling confirmed that these proteins possess mutarotase-like active sites with conserved catalytic residues. We overexpressed the homolog from the sulfo-ED operon of Herbaspirillum seropedicaea (HsSQM) and used it to demonstrate SQ mutarotase activity for the first time. This was accomplished using nuclear magnetic resonance exchange spectroscopy, a method that allows the chemical exchange of magnetization between the two SQ anomers at equilibrium. HsSQM also catalyzed the mutarotation of various aldohexoses with an equatorial 2-hydroxy group, including D-galactose, D-glucose, D-glucose-6-phosphate (Glc-6-P), and D-glucuronic acid, but not D-mannose. HsSQM displayed only 5-fold selectivity in terms of efficiency (kcat/KM) for SQ versus the glycolysis intermediate Glc-6-P; however, its proficiency [kuncat/(kcat/KM)] for SQ was 17 000-fold better than for Glc-6-P, revealing that HsSQM preferentially stabilizes the SQ transition state