Production and characterization of milk-clotting enzymes by Bacillus circulans NRC-3

ABSTRACT

The production of Milk-clotting enzyme was investigated using seven bacterial isolates. Bacillus circulans NRC-3 proved to be the most potent and produced a highly active milk-clotting enzyme with negligible proteolytic activity on wheat bran medium as a food byproduct. The milk- clotting enzyme activity was optimal at 50°C and CaC12 of final concentration of 0.3 mM. The enzyme was highly stable at elevated temperatures [70°C] and retained more than 60% of the original activity by heating for 5min.at 70°C. Partial purification of the enzyme was achieved by fractional precipitation with acetone or ethanol. The 40% acetone fraction was the most promising and possessed rennin properties with no proteolytic activity. It reached about 4.3-fold that of the culture filtrate. The partial purified B.circulans NRC-3 milk-coltting enzyme was applied on three buffalo fresh milk specimens, i.e. full- cream, half- cream and skimmed fresh. The enzyme exhibited the maximum relative activity [100%] with both half- cream and skimmed fresh milk specimens and dropped to 67.1% with the full- cream specimen