Enzymatic transformation of nitro-aromatic compounds by a flavin-free NADH azoreductase from Lysinibacillus sphaericus.

Azo dyes and nitro-aromatic compounds are the largest group of pollutants released in the environment as industrial wastes. They create serious health and environmental problems. Azoreductases catalyze the reduction of azo dyes and nitro compounds to their respective amines. AN azoreductase was purified up to 12-fold from Lysinibacillus sphaericus using ion-exchange and size exclusion chromatography. It was optimally active at pH 7.4 and 75 °C. It was stable at 70 °C for 30 min. The purified enzyme utilized NADH rather than NADPH as an electron donor to reduce substrates. The molecular weight of the purified enzyme was ~29 kDa. The enzyme also acted as nitroreductase and could selectively reduce the nitro group of 2-nitrophenol, 4-nitrobenzoic acid, 2-nitro-benzaldehyde and 3-nitrophenol. Reduction products of these compounds were identified by IR and NMR.

Properties of NAD (P) H azoreductase from alkaliphilic red bacteria Aquiflexum sp. DL6.

Azoreductase plays a key role in bioremediation and biotransformation of azo dyes. It initializes the reduction of azo bond in azo dye metabolism under aerobic or anaerobic conditions. In the present study, we isolated an alkaliphilic red-colored Aquiflexum sp. DL6 bacterial strain and identified by 16S rRNA method. We report nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide phosphate-dependent azoreductase purified from Aquiflexum sp. DL6 by a combination of ammonium sulfate precipitation and chromatography methods. The azoreductase was purified up to 30-fold with 37 % recovery. The molecular weight was found to be 80 kDa. The optimum activity was observed at pH 7.4 and temperature 60 °C with amaranth azo dye as a substrate. The thermal stability of azoreductase was up to 80 °C. The azoreductase has shown a wide range of substrate specificity, including azo dyes and nitro aromatic compounds. Metal ions have no significant inhibitory action on azoreductase activity. The apparent K m and V max values for amaranth azo dye were 1.11 mM and 30.77 U/mg protein respectively. This NAD (P) H azoreductase represents the first azoreductase to be characterized from alkaliphilic bacteria.

Proteomic profiling and interactome analysis of ER-positive/HER2/neu negative invasive ductal carcinoma of the breast: towards proteomics biomarkers.

Breast cancer, especially ER positive/HER2/neu negative IDC, is the predominant subtype of invasive ductal carcinoma. Although proteomic approaches have been used towards biomarker discovery in clinical breast cancer, ER positive/HER2/neu negative IDC is the least studied subtype. To discover biomarkers, as well as to understand the molecular events associated with disease progression of estrogen receptor positive/HER2/neu negative subtype of invasive ductal carcinoma, differential protein expression profiling was performed by using LC-MS(E) (MS at elevated energy). A total of 118 proteins were identified, of which 26 were differentially expressed. These identified proteins were functionally classified and their interactions and coexpression were analyzed by using bioinformatic tools PANTHER (Protein Analysis THrough Evolutionary Relationships) and STRING (Search Tool for the Retrieval of Interacting Genes). These proteins were found to be upregulated and were involved in cytoskeletal organization, calcium binding, and stress response. Interactions of annexin A5, actin, S100 A10, glyceraldehyde 3 phosphate dehydrogenase, superoxide dismutase 1, apolipoprotein, fibrinogen, and heat shock proteins were prominent. Differential expression of these proteins was validated by two-dimensional gel electrophoresis and Western blot analysis. The cluster of these proteins may serve as a signature profile for estrogen receptor positive/ HER2/neu negative subtype.

Analysis of AGE modified proteins and RAGE expression in HER2/neu negative invasive ductal carcinoma.

Cancer is associated with increased glycolysis and carbonyl stress. In view of this, AGE modified proteins were identified from clinical breast cancer tissue using 2DE-immunoblot and mass-spectrometry. These proteins were identified to be serotransferrin, fibrinogen gamma chain, glycerol-3-phosphate dehydrogenase, lactate dehydrogenase, annexin II, prohibitin and peroxiredoxin 6, which have established role in cancer. Further, RAGE expression and its downstream signaling proteins NADPH oxidase and NF-kB were studied. Role of these AGE modified proteins and RAGE signaling in breast cancer is discussed.

Mechanistic investigation of decolorization and degradation of reactive red 120 by Bacillus lentus BI377.

Bacillus lentus BI377, isolated from textile effluent-contaminated soil, was able to degrade 97% and 92% of Reactive Red 120 dye when 1200 and 1500 mg/l, respectively, of dye was added to nutrient broth (NB) at 35 °C within 12 h. UV-vis spectroscopy, GC-MS, FTIR and 1H NMR revealed the formation of catechol which may be further utilized by the bacterium via the TCA cycle, leading to complete mineralization. Structural analysis of metabolites in conjunction with enzyme activity studies confirmed the involvement of azoreductase, cytochrome P450 monooxygenase and other antioxidant enzymes. Decreases in total organic carbon and in biological and chemical oxygen demand suggest formation of low molecular weight metabolites that could be completely mineralized. These results suggest the potential use of B. lentus BI377 towards online treatment of textile dye effluents by using an appropriate bioreactor over a wide range of pH. This study opens-up a dependable and proficient way to use industrially viable non-pathogenic strains for biotransformation of carcinogenic dyes to ecofriendly compounds.

Decolorization and degradation of Disperse Blue 79 and Acid Orange 10, by Bacillus fusiformis KMK5 isolated from the textile dye contaminated soil.

The release of azo dyes into the environment is a concern due to coloration of natural waters and due to the toxicity, mutagenicity and carcinogenicity of the dyes and their biotransformation products. The dye degrading bacterial strain KMK 5 was isolated from the textile dyes contaminated soil of Ichalkaranji, Maharashtra, India. It was identified as Bacillus fusiformis based on the biochemical and morphological characterization as well as 16S rDNA sequencing. KMK 5 could tolerate and degrade azo dyes, Disperse Blue 79 (DB79) and Acid Orange 10 (AO10) under anoxic conditions. Complete mineralization of DB79 and AO10 at the concentration of 1.5g/l was observed within 48h. This degradation potential increased the applicability of this microorganism for the dye removal.