title: Secretory expression, functional characterization, and molecular genetic analysis of novel halo-solvent-tolerant protease from Bacillus gibsonii.
abstract: A novel protease gene from Bacillus gibsonii, aprBG, was cloned, expressed in B. subtilis, and characterized. High-level expression of aprBG was achieved in the recombinant strain when a junction was present between the promoter and the target gene. The purified recombinant enzyme exhibited similar N-terminal sequences and catalytic properties to the native enzyme, including high affinity and hydrolytic efficiency toward various substrates and a superior performance when exposed to various metal ions, surfactants, oxidants, and commercial detergents. AprBG was remarkably stable in 50% organic solvents and retained 100% activity and stability in 0-4 M NaCl, which is better than the characteristics of previously reported proteases. AprBG was most closely related to the high-alkaline proteases of the subtilisin family with a 57-68% identity. The secretion and maturation mechanism of AprBG was dependent on the enzyme activity, as analyzed by site-directed mutagenesis. Thus, when taken together, the results revealed that the halo-solvent-tolerant protease AprBG displays significant activity and stability under various extreme conditions, indicating its potential for use in many biotechnology applications.
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Aihua
authors.initials: A
authors.lastname: Deng
authors.forename: Guoqiang
authors.initials: G
authors.lastname: Zhang
authors.forename: Nana
authors.initials: N
authors.lastname: Shi
authors.forename: Jie
authors.initials: J
authors.lastname: Wu
authors.forename: Fuping
authors.initials: F
authors.lastname: Lu
authors.forename: Tingyi
authors.initials: T
authors.lastname: Wen
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D016000:Cluster Analysis; D004252:DNA Mutational Analysis; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D020536:Enzyme Activators / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D004795:Enzyme Stability; D015870:Gene Expression; D006868:Hydrolysis; D060847:Metabolic Engineering; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010447:Peptide Hydrolases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D010802:Phylogeny; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012965:Sodium Chloride / Q000378:metabolism
chemical_list: D004269:DNA, Bacterial; D020536:Enzyme Activators; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D012965:Sodium Chloride; D010447:Peptide Hydrolases