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Purification and characterization of a novel serine protease from the mushroom Pholiota nameko
- Guan, Gui-Ping, Zhang, Guo-Qing, Wu, Ying-Ying, Wang, He-Xiang, Ng, Tzi-Bun
- Journal of bioscience and bioengineering 2011 v.111 no.6 pp. 641-645
- Pholiota microspora, aluminum, calcium, casein, cations, cobalt, copper, enzyme activity, fruiting bodies, gel chromatography, ion exchange chromatography, iron, lead, lithium, magnesium, manganese, mercury, metal ions, molecular weight, mushroom bodies, mushrooms, pH, potassium, serine proteinases, temperature, zinc
- A novel serine protease, with a molecular mass of 19kDa and the N-terminal sequence of ARTPEAPAEV, was isolated from dried fruiting bodies of the mushroom Pholiota nameko. The purification protocol comprised ion exchange chromatography on DEAE-cellulose, Q-Sepharose and SP-Sepharose, and gel filtration on Superdex 75. It was unadsorbed on DEAE-cellulose and Q-Sepharose but adsorbed on SP-Sepharose. It exhibited an optimum temperature at 50°C, an optimum pH at pH 8.8, a Km of 5.64mg/mL and a Vmax of 0.98μmol/min/mL against substrate casein. A number of metal ions inhibited the enzyme including Pb²⁺, Mn²⁺, Ca²⁺, Hg²⁺, Zn²⁺, Cu²⁺, Co²⁺, Fe³⁺ and Al³⁺, with the inhibition of the last two cations being the most potent. K⁺ and Mg²⁺ slightly enhanced, while Li⁺ moderately potentiated the activity of the protease. The protease was strongly inhibited by phenylmethylsulfonyl fluoride (PMSF), suggesting that it is a serine protease.