الفهرس 
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Abstract
L-glutaminase (L-glutamine amidohydrolase, EC 3.5.1.2) is an important enzyme that catalyzes the deamination of L-glutamine to L-glutamic acid and ammonium ions. Recently, it has received significant attention owing to its potential applications in cancer therapy, food industry and preparation of high-value chemicals like threonine. All fungi isolated from agricultural soil are able to produce L-glutaminase. In the present study, a newly locally isolated fungal strain was found to produce extracellular L-glutaminase and it was identified genetically as Aspergillus fumigatus MT071344 using the 16S rRNA gene sequence analysis.
The production of L-glutaminase from Aspergillus fumigatus MT071344 was evaluated and optimized under different fermentation modes. The optimum L-glutaminase production was obtained at pH 6, after 5.5 incubation time and at 28.5°C temperature. The growth medium contained glucose as carbon source (2%w/v) and soybean (5g/L) as nitrogen source, glycine (0.1g/L) as amino acids source and CaCl2 (0.05 g/L) as minerals. These optimized conditions when applied to a solid state fermentation (SSF) have produced 94.05 U/ml of L-glutaminase activity.
Ammonium sulfate precipitation (75% saturation) was used to precipitate proteins from fermented substrate culture. The crude enzyme was purified about 7.55 times more and the yield rate was enhanced up to 87.5% with a specific activity of 6.5 U/mg, then purified by dialysis against 0.05M Tris-HCl buffer (pH 8.0) before being applied to Sephadex G-100 column and DEAE-cellulose column chromatography.
However, the yield and the final specific activity after gel filtration chromatography were 43.75% and 40 U/mg, respectively and purification fold increased to 46.5. The molecular mass of the purified L-glutaminase enzyme was found to be 55KDa. Moreover, the purified L-glutaminase exerted antitumor activity against different cell lines such as Human hepatocellular carcinoma cell line (HepG2), Human colon carcinoma (HCT-116), Human breast carcinoma (MCF-7), and Human promyelocytic leukemia carcinoma (HL-60) cell lines with the highest cytotoxic activity against HCT-116 cell line with an IC50 of 60.4±1.7 µg/ml. In conclusion, L-glutaminase was purified from Aspergillus fumigatus MT071344 using simple methods and its unique properties with its relatively low molecular weight can facilitate its usage in medical applications as antitumor drug.