Calcium modulated activity enhancement and thermal stability study of a cationic peroxidase purified from wheat bran

A cationic form of peroxidase was purified from wheat bran, a major by-product of wheat milling industry to near homogeneity by ammonium sulphate precipitation, anion exchange, and cation exchange and gel filtration chromatography. It was a glycoprotein with a molecular weight of 44 kDa, pH optimum of 4.8 and carbohydrate content of 13.8%. The enzyme showed Ping-Pong Bi Bi type catalysis. Inclusion of calcium during purification increased the specific activity and yield of the enzyme. Activity of purified enzyme was enhanced by calcium more than 400% in a biphasic manner. Purified enzyme exhibited increased thermal stability when calcium was added. There was no change in tryptophan fluorescence by the addition of calcium, but the haem absorption at 403 nm showed a change indicating an alteration in the haem environment. Calcium is essential for maintaining the haem structure, enzymatic activity and thermal stability of wheat bran peroxidase.