Pigeornpea (Cajanus Cajun) Urease Immobilized on Alginate Beads Showing Improved Stability and Operational Pa Rameters

Abstract

Plant urease (urea amidohydrolase; EC 3.5.1.5) is a seed protein that is common in most legumes. It is also common in many bacteria and fungi and several species of yeast. Urease allows organisms to use exogenous and internally generated urea as a nitrogen source by catalyzing the hydrolysis of urea to ammonia and carbon dioxide. U~ease from pigeonpea (Cajanus cajan) was immobilized in alginate beads by entrapment method. Approximately 48% immobilization of urease in alginate was observed at 3.5% (w/v) alginate with 0.65 mg protein/5 ml alginate gel. From steady state kinetics, the optimum temperature for soluble urease was 65 °C and 80 °C for immobilized urease, the optimum pH of urease 7.3 (soluble) and 8.0 (alginate immobilized). It was observed that there was a increase in Km for alginate-immobilized urease (4.34 mM) when compared with soluble urease (2.22 mM).The alginate immobilized ureases showed a decrease in Vmax (0.769 ~mol NH3 min-1 mg-1 protein) as compare to soluble enzymes (0.80 ~mol NH3 min-1 mg-1protein).With time-dependent thermal inactivation studies. The immobilized enzyme showed improved stability at 75 °C and the t112 of decay in urease activity was 12 min and 50 min for soluble and immobilized urease, respectively. The reusability studies · revealed that the alginate immobilized urease after 8 uses retained more than 50% of the original activity. The immobilized urease was always kept in 50 mM Tris-acetate buffers, pH 7.3 at 4 °C. For storage stability studies, the loss of activity, for soluble urease after 50 days of storage at 4 °C was 80% and in comparison to alginate immobilized urease the loss was 51%. The plot of % residual activity versus the number of days gave a t112 of 30 days for soluble urease; 51 days for alginate-immobilized urease.