title: A novel alkaline protease from wild edible mushroom Termitomyces albuminosus.
abstract: A protease with a molecular mass of 30 kDa and the N-terminal sequence of GLQTNAPWGLARSS, was isolated from fresh fruiting bodies of the wild edible mushroom Termitomyces albuminosus. The purification protocol included ion exchange chromatography on DEAE-cellulose, Q-Sepharose, SP-Sepharose and FPLC-gel filtration on Superdex 75. The protein was unadsorbed on DEAE-cellulose and Q-Sepharose, but adsorbed on SP-Sepharose. The optimal pH and temperature of the purified enzyme were 10.6 and 60 °C, respectively. The enzyme was stable in the presence of 2 % (v/v) Tween 80 and 4 M urea. More than 80 % of the enzyme activity was retained in 2 % (v/v) Triton X 100, 54 % in 10 mM EDTA and 31 % in 2 % (w/v) SDS. The enzyme was strongly inhibited by phenylmethylsulfonyl fluoride (PMSF), but not inhibited by dithiothreitol (DTT), pepstatin or lima bean trypsin inhibitor suggesting that it was a serine protease but not a trypsin-like one. The protease was inhibited by Hg(2+), Cu(2+), and Fe(3+) ions. The K(m) and V(max) values of the purified enzyme for casein were 8.26 mg ∙ ml(-1) and 0.668 mg ∙ ml(-1) ∙ min(-1), respectively.
authors.affiliation: Hebei Engineering University, Handan, China.
authors.forename: Suyue
authors.initials: S
authors.lastname: Zheng
authors.forename: Hexiang
authors.initials: H
authors.lastname: Wang
authors.forename: Guoqing
authors.initials: G
authors.lastname: Zhang
mesh_terms: D000595:Amino Acid Sequence; D002412:Cations; D002853:Chromatography, Liquid; D057075:Enzyme Assays; D004791:Enzyme Inhibitors / Q000737:chemistry; D004795:Enzyme Stability; D048690:Fruiting Bodies, Fungal / Q000201:enzymology*; D005656:Fungal Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D020539:Sequence Analysis, Protein; D057057:Serine Proteases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D055434:Termitomyces / Q000201:enzymology*
chemical_list: D002412:Cations; D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D057057:Serine Proteases