Production and isolation of Glutaminase and to study its role as anti-cancer drug
Shipra Kalra, Kanav Midha, Sandeep Singh
Glutamine is the respiratory fuel of tumor cells. When Glutamine concentrationrnis depleting in the tumor cells, it undergoes cell death. Glutaminase can actrnas anticancer drug. This research focuses on production and isolation ofrnGlutaminase and to study its role as an anticancer drug. For isolation ofrnGlutaminase produced by E-coli acetone precipitation was carried outrnwhich causes complete precipitation. Glutaminase activity was determined spectrophotometricallyrnby Nessler’s reagent at 630 nm. Purification of the enzyme was done by HPLC andrnion exchange chromatography. Anti-tumor properties of L-Glutaminase can bernchecked with the help of Rhodamine Assay and NBT Assay. The optimumrnconcentration was observed when 12.5% of glutamine was added in the productionrnmedium. E-coli showed max Glutaminase activity at 12.5% Glutamine, 100mMrnInorganic Phosphate and 250mM Ammonium Sulphate. Glutaminase activity afterrnacetone precipitation is 1665 U/ml and Specific activity of 244. 9 IU/mg. AfterrnAmmonium Sulphate extraction Glutaminase activity was found to be 1790.2 U/ml.rnThere is almost no change in the specific activity of the enzyme. Results ofrnIon exchange and HPLC were compared and were found that HPLC showed betterrnenzyme activity. From both the cytotoxicity assay, it was concluded that thernGlutaminase inhibits the growth of cancer cells by depleting the glutamine fromrnthe media.
Shipra Kalra, Kanav Midha, Sandeep Singh. Production and isolation of Glutaminase and to study its role as anti-cancer drug. European Journal of Biotechnology and Bioscience, Volume 4, Issue 8, 2016, Pages 31-37