Production of highly active fungal milk-clotting enzyme by solid-state fermentation

Cheese production is projected to reach 20 million metric tons by 2020, of which 33% is being produced using calf rennet (EC 3.4.23.4). There is shortage of calf rennet, and use of plant and microbial rennets, hydrolyze milk proteins non-specifically resulting in low curd yields. This study reports fungal enzymes obtained from cost effective medium, with minimal down streaming, whose activity is comparable with calf and Mucor rennet. Of the fifteen fungi that were screened, Mucor thermohyalospora (MTCC 1384) and Rhizopus azygosporus (MTCC 10195) exhibited the highest milk-clotting activity (MCA) of 18,383 ± 486 U/ml and 16,373 ±  558 U/ml, respectively. Optimization exhibited a 33% increase in enzyme production (30 g wheat bran containing 6% defatted soy meal at 30 °C, pH 7) for M. thermohyalospora. The enzyme was active from pH 5-10 and temperature 45-55 °C. Rhizopus azygosporus exhibited 31% increase in enzyme production (30 g wheat bran containing 4% defatted soy meal at 30 °C, pH 6) and the enzyme was active from pH 6-9 at 50 °C. Curd yields prepared from fungal enzyme extract decreased (5-9%), when compared with calf rennet and Mucor rennet. This study describes the potential of fungal enzymes, hitherto unreported, as a viable alternative to calf rennet.