PURIFICATION AND PROTEOLYTIC ACTION OF MILK- CLOTTING ENZYME PRODUCED BY PENICILLIUM CITRINUM

Abstract

Electrophoresis of the partially purified milk-clotting enzyme produced by Penicillium citrinum with 0.02M acetate buffer (pH 3.42) showed four protein components. Fractionation of the enzyme preparation with acetone led to individual isolation of the four electrophoretic components. The course of proteolysis in the first phase of enzymic action of each of the partially purified fungal enzymes and the four pure fractions was conducted and compared with those of trypsin and calf rennin. The fungal enzyme contained two rennin-like fractions constituting most of the milk-clotting activity with limited proteolysis and another two fractions with less clotting activity and more proteolysis. The macroglycopeptide isolated by the action of the fungal rennin-like enzyme fraction comprised 12 amino acids while its carbohydrate moiety consisted of galactose, galactosamine, and N-acetylneuraminic acid. Further studies on the most active rennin-like enzyme fraction showed that calcium chloride enhanced only the primary stage of the enzymic phase but near the end of this phase the rate of NPN release was very slow.