Abstrato

Partial Purification and Characterization of Urease from Germinating Chickpea (Cicer arietinum L.) Seed

Niranjan Kumar Sana, Sahela Pervin, Md.Golam Sarowar Jahan, Md. Masudul Hasan Khan, Md. Rezaul Karim, Bidhan Chandra Sarkar, Ranajit Kumar Shaha

Urease, the urea-hydrolyzing enzyme, was identified abundantly in germinating chickpea (Cicer arietinumL.) seed. The enzyme was purified to homogeneity by the sequential steps of 20% acetone extraction, followed by 50% acetone fractionation, gel filtration on Sephadex G-200, and DEAEcellulose chromatography. The purification fold was 44.99 with a final specific activity of 489.57mMmin-1mg-1. The purified urease was a hexamer of identical subunits. The native enzyme had amolecularmass of 510 kDa (Gel filtration, Sephadex G-200) whereas subunit values of 85 kDa were determined on PAGE with sodium dodecyl sulfate. The optimum pH and temperature of the purified urease were 7.2 and 480C, respectively, using urea as substrate. The half-life of urease was 30 days in 50 mM phosphate buffer (pH7.2) at 40C.Kmvalue of the purified urease for ureawas 3.1mM. Urease activity was decreased by 50% within 5 minutes at 700C. The optimum substrate (urea) concentration for urease was 25 mM. The enzyme showed the highest activity when incubated for 30 minutes at 480C. Ca2+ enhanced urease activity by 120.47%, while Pb2+, Cu2+, Zn2+ and Hg2+ almost completely inhibited the urease activity.