title: Purification and properties of the highly thermostable alkaline protease from an alkaliphilic and thermophilic Bacillus sp.
abstract: Thermostable alkaline protease from an alkaliphilic thermophile Bacillus sp. B18' was purified by using DEAE- and CM-Toyopearl 650M column chromatographies. Molecular weights of the enzyme determined by SDS-PAGE and gel filtration were 30,000 and 28,000, respectively. The optimum pH and temperature toward the hydrolysis of casein were pH 12-13 and 85 degrees C, both of which are higher than those of a mesophilic alkaline protease from an alkaliphile, Bacillus sp. B21-2. The enzyme was stable at pH 5.0-12.0 and about 60% of the initial enzymatic activity was retained after a 60 min incubation period at pH 10.0 and 70 degrees C. Thermostability of the enzyme was enhanced by Ca2+. The enzyme activity was inhibited by DFP, suggesting that the enzyme is a serine protease. The NH2-terminal amino acid is Gln, which is that of many subtilisin-type proteases. The 20 residues of the NH2-terminal amino acid sequence have a comparative high homology with those of other alkaline proteases from alkaliphiles (40-50%), especially thermostable alkaline protease from Bacillus sp. No. AH-101 (95%) and Thermoactinomyces sp. HS682 (95%).
authors.affiliation: Osaka Prefectural Industrial Technology Research Institute, Japan.
authors.forename: N
authors.initials: N
authors.lastname: Fujiwara
authors.forename: A
authors.initials: A
authors.lastname: Masui
authors.forename: T
authors.initials: T
authors.lastname: Imanaka
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000145:classification / Q000201:enzymology*; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D013696:Temperature
chemical_list: D012697:Serine Endopeptidases; C032923:microbial serine proteinases