Helvellisin, a novel alkaline protease from the wild ascomycete mushroom Helvella lacunosa(MICROBIAL PHYSIOLOGY AND BIOTECHNOLOGY)

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A 33.5-kDa serine protease designated as helvellisin was isolated from dried fruiting bodies of the wild ascomycete mushroom Helvella lacunosa. It was purified by using a procedure which entailed ion exchange chromatography on DEAE-cellulose, CM-Sepharose, Q-Sepharose, and FPLC-gel filtration on Superdex 75. The protease was characterized by unique N-terminal amino acid sequence, thermostability and pH stability. The protease exhibited a pH optimum of 11.0 and a temperature optimum of 65℃, with about 40% activity remaining at 87℃ and pH5 and 13. Helvellisin demonstrated a protease activity of 14600U/mg toward casein. The K_m of the purified protease for casein was 3.81mg/ml at pH 11.0 and 37℃. The V_<max> was 5.35×10^<-2>mg ml^<-1> min^<-1>. It was adversely affected by phenylmethylsulfonyl fluoride, suggesting that it is serine protease. The activity of the protease was enhanced by Mg^<2+>, Fe^<2+> and Mn^<2+>, but was curtailed by Cu^<2+>, Hg^<2+> and Fe^<3+>. It was devoid of antifungal and ribonuclease activities.