Purification and Characterization a Novel Alkaline Serine Protease from the Medicinal Mushroom Cordyceps sobolifera

摘要

A novel protease was purified from dried fruiting bodies of the mushroom Cordyceps sobolifera. The isolation procedure utilized ion exchange chromatography on DEAE-cellulose and SP-sepharose followed by gel filtration on Superdex 75. The protease did not adsorb on DEAE-cellulose but bound to SP-Sepharose. In sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE), the protease resolved as a single band with an apparent molecular mass of 31 kDa. It exhibited optimal activity at a temperature of 60 °C, a pH of 12.4 with a Km of 1. 98 mg/mL and a Vmax of 23. 56 μg/mL/min against its substrate casein. Protease activity was enhanced by the Fe2+ion at low concentrations in 1. 25-10 mM and was strongly inhibited by Hg2+up to 1. 25 mM. The protease was strongly inhibited by phenylmethylsulfonyl fluoride (PMSF), suggesting that it was a serine protease. It manifested significant inhibitory activity toward HIV-1 reverse transcriptase (RT) with an IC50 value of 8. 2×10-3 μM, which was the highest anti-HIV-1 RT activity of reported mushroom proteins.