Screening, nutritional optimization and purification for phytase produced by Enterobacter aerogenes and its role in enhancement of hydrocarbons degradation and biofilm inhibition.

In this study, a novel isolate of Enterobacter aerogenes isolated from contaminated soils with hydrocarbons had extracellular phytate-degrading activity. Enterobacter aerogenes isolates were identified by biochemical tests and confirmed by16S rRNA gene products (amplified size 211bp) for genotypic detection. The phytase activity was reached to maximum activity when this isolate was cultivated under the optimal conditions which consisted of using minimal salt medium containing 1%(w/v) rice bran as a sole source for carbon and 2% (w/v) yeast extract at pH 5.5 and temperature of 50°C for 48 h. The phytase had purified to homogeneity by 50% ammonium sulphate precipitation, ion exchange and gel filtration chromatography with 75.7 fold of purification and a yield of 30.35%. The purified phytase is a single peptide with approximate molecular mass of 42 kDa as assessed by SDS-PAGE. The highest degradative ability by Enterobacter aerogenes of black oil, white oil and used engine oil had observed after 72 h of incubation. Rapid degradation of black oil and used engine oil had also observed while slow degradation of white oilat all time of incubation. The purified phytase inhibited biofilm formation ability in a dose-dependent manner for all Gram-negative and Gram-positive biofilm-forming bacteria and a significant difference in cell surface hydrophobicity was observed after exposure of planktonic cells to phytase for hour. The hydrolyzing effect of phytase released by Enterobacter aerogenes for complex salts of phosphorus that are insoluble in the soil led to increase of phosphorus concentrations and enhanced the ability of Enterobacter aerogenes to degrade a specific hydrocarbon in contaminated soil so that the phytase has a promising application in bioremediation of contaminated soils with hydrocarbons.

Two NAD-linked redox shuttles maintain the peroxisomal redox balance in Saccharomyces cerevisiae.

In Saccharomyces cerevisiae, peroxisomes are the sole site of fatty acid ß-oxidation. During this process, NAD+ is reduced to NADH. When cells are grown on oleate medium, peroxisomal NADH is reoxidised to NAD+ by malate dehydrogenase (Mdh3p) and reduction equivalents are transferred to the cytosol by the malate/oxaloacetate shuttle. The ultimate step in lysine biosynthesis, the NAD+-dependent dehydrogenation of saccharopine to lysine, is another NAD+-dependent reaction performed inside peroxisomes. We have found that in glucose grown cells, both the malate/oxaloacetate shuttle and a glycerol-3-phosphate dehydrogenase 1(Gpd1p)-dependent shuttle are able to maintain the intraperoxisomal redox balance. Single mutants in MDH3 or GPD1 grow on lysine-deficient medium, but an mdh3/gpd1Δ double mutant accumulates saccharopine and displays lysine bradytrophy. Lysine biosynthesis is restored when saccharopine dehydrogenase is mislocalised to the cytosol in mdh3/gpd1Δ cells. We conclude that the availability of intraperoxisomal NAD+ required for saccharopine dehydrogenase activity can be sustained by both shuttles. The extent to which each of these shuttles contributes to the intraperoxisomal redox balance may depend on the growth medium. We propose that the presence of multiple peroxisomal redox shuttles allows eukaryotic cells to maintain the peroxisomal redox status under different metabolic conditions.