Abstract:

The aims of this study were to investigate the fermentation conditions for the production of recombinant Mucor pusillus rennet (RMPR) and to enzymologically characterize the enzyme obtained. An optimum enzyme activity was obtained after 96 h of induction with 1% methanol as the inducer under the following conditions: initial OD600 nm, 0.5; and volume of medium contained in 500 mL flask, 100 mL．The incubation both obtained was centrifugated, and the supernatant was then salted out with ammonium sulfate to prepare crude enzyme solution, and enzymological characterization was carried out for the crude enzyme solution. The optimal reaction temperature of the enzyme was 60 ℃, and it kept stable in the temperature range from 30 to 45 ℃ and the residual enzyme activity was still 86.5% after 90 min of incubation at 45 ℃. As pH increased from 5.5 to 7.0, the activity of the enzyme presented a gradual decrease and the highest enzyme activity was observed at pH 5.5. At a Ca2+ concentration of 0.03 mol/L, the enzyme had the highest activity to clot milk, and Na+ had no notable effect on the enzyme, and Ni+ and Fe2+ were both an enzyme inhibitor, while Mn2+, K+ and Mg2+ all had promoting effect on the enzyme.

Key words: Mucor pusillus, rennet, enzymological characterization, fermentation, recombinant