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Microbial milk-clotting enzymes are valued as calf rennet substitutes in the cheese industry. Mucor pusillus QM 436 was identified to produce the highest milkclotting activity during screening of 19 fungal strains. Salted whey resulting from Ras (Cephalotyre) cheese manufacture as a whole medium for growth of Mucor pusillus QM 436 and production of the enzyme. The milk-clotting enzyme from Mucor pusillus QM 436 was purified to 7.14-fold with 54.4% recovery by precipitation in ammonium sulfate, ethanol and fractionated by gel filtration on Sephadex G-100. The enzyme was active in the pH range 5.5-7.5 and was inactivated completely by heating 5 min at 70ºC and 30 min at 65ºC. The highest level of enzyme activity was obtained at 60ºC, pH 5.5. A positive and proportional relationship occurred in presence of CaCL2 in milk, with inhibition occurred in presence of NaCl. The high level of milk-clotting activity coupled with a low level of thermal stability suggested that the milk-clotting enzyme from Mucor pusillus QM 436 should be considered as a potential substitute for calf rennet.

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