Abstract

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The protease of malted Digitaria exilis (white acha) species was extracted, purified and characterized. The enzyme was extracted with 200 ml of 0.1M citrate phosphate buffer (pH 7) containing 0.4% (w/v) cysteine and 0.86% NaCl, purified by dialysis against 4M sucrose, Ion exchange chromatography on CM sepharose and gel filtration chromatography on Sepharose 4B gel. On ion exchange chromatography and gel filteration chromatography, the white acha protease was purified 2.21 folds with a specific activity of 362 Umg-1 protein. The relative molecular weight of the protease was estimated to be 88,000 daltons by Gel filtration. The white acha protease was optimally active at 50oC and pH 7, but retained about 40% of its activity at 70oC (30 mins) and pH 8. Appreciable stimulation (P<0.05) of the white acha protease was only achieved by Mn2+, while the other metal ions (Zn2 ,Ba2+ , Fe2+ ,Cu2+ ,Ca2+ ,Sr2+ & Hg2+) were inhibitory. Guanidine chloride, n-bromosuccinamide and EGTA were inhibitory (P<0.05) to the acha protease, while sodium sulphite and 2-mercaptoethanol (2-Me) were stimulatory with striking stimulation obtained with 2-ME. A significant effect (P<0.05) of inhibitors on acha protease was recorded. The enzyme exhibited broad specificity (70 – 100%) in the hydrolysis of various proteins (Bovine serum, albumin, casein, egg albumin and gelatin) and showed strongest affinity for casein when its km (0.188 mg/ml) and Vmax (0.208 U/mol) values were obtained, respectively. Therefore, Digitaria exilis protease can be useful in food industries if harnessed

Keywords: Digitaria exilis, enzymes, extraction, proteases, purification