title: Identification and Characterization of a New Alkaline Thermolysin-Like Protease, BtsTLP1, from Bacillus thuringiensis Serovar Sichuansis Strain MC28.
abstract: Thermolysin and its homologs are a group of metalloproteases that have been widely used in both therapeutic and biotechnological applications. We here report the identification and characterization of a novel thermolysin-like protease, BtsTLP1, from insect pathogen Bacillus thuringiensis serovar Sichuansis strain MC28. BtsTLP1 is extracellularly produced in Bacillus subtilis, and the active protein was purified via successive chromatographic steps. The mature form of BtsTLP1 has a molecule mass of 35.6 kDa as determined by mass spectrometry analyses. The biochemical characterization indicates that BtsTLP1 has an apparent Km value of 1.57 mg/ml for azocasein and is active between 20°C and 80°C. Unlike other reported neutral gram-positive thermolysin homologs with optimal pH around 7, BtsTLP1 exhibits an alkaline pH optimum around 10. The activity of BtsTLP1 is strongly inhibited by EDTA and a group of specific divalent ions, with Zn(2+) and Cu(2+) showing particular effects in promoting the enzyme autolysis. Furthermore, our data also indicate that BtsTLP1 has potential in cleaning applications.
authors.affiliation: China Research Center, DuPont Industrial Bioscience, Shanghai 200335, P.R. China.|State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, P.R. China.
authors.forename: Zhenghong
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: China Research Center, DuPont Industrial Bioscience, Shanghai 200335, P.R. China.
authors.forename: Helong
authors.initials: H
authors.lastname: Hao
authors.affiliation: China Research Center, DuPont Industrial Bioscience, Shanghai 200335, P.R. China.
authors.forename: Zhongmei
authors.initials: Z
authors.lastname: Tang
authors.affiliation: China Research Center, DuPont Industrial Bioscience, Shanghai 200335, P.R. China.
authors.forename: Zhengzheng
authors.initials: Z
authors.lastname: Zou
authors.affiliation: China Research Center, DuPont Industrial Bioscience, Shanghai 200335, P.R. China.
authors.forename: Keya
authors.initials: K
authors.lastname: Zhang
authors.affiliation: China Research Center, DuPont Industrial Bioscience, Shanghai 200335, P.R. China.
authors.forename: Zhiyong
authors.initials: Z
authors.lastname: Xie
authors.affiliation: Danisco US Inc., Genencor Division, DuPont Industrial Bioscience, Palo Alto, CA 94304, USA.|Genencor International B.V., DuPont Industrial Bioscience, 2333 CN Leiden, The Netherlands.
authors.forename: Lilia
authors.initials: L
authors.lastname: Babe
authors.forename: Frits
authors.initials: F
authors.lastname: Goedegebuur
authors.affiliation: China Research Center, DuPont Industrial Bioscience, Shanghai 200335, P.R. China.
authors.forename: Xiaogang
authors.initials: X
authors.lastname: Gu
mesh_terms: D000595:Amino Acid Sequence; D001413:Bacillus thuringiensis / Q000201:enzymology*; D002364:Caseins / Q000378:metabolism; D002413:Cations, Divalent / Q000378:metabolism; D002845:Chromatography; D016000:Cluster Analysis; D004492:Edetic Acid / Q000378:metabolism; D020536:Enzyme Activators / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D013058:Mass Spectrometry; D008666:Metalloendopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D008670:Metals / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid; D013696:Temperature
chemical_list: D002364:Caseins; D002413:Cations, Divalent; D020536:Enzyme Activators; D004791:Enzyme Inhibitors; D008670:Metals; C026018:azocasein; D004492:Edetic Acid; D008666:Metalloendopeptidases; C044194:microbial metalloproteinases
keywords: Alkaline metalloprotease; Bacillus thuringiensis; Thermolysin; Thermolysin-like protease