title: Significant improvement of thermal stability of glucose 1-dehydrogenase by introducing disulfide bonds at the tetramer interface.
abstract: Rational design was applied to glucose 1-dehydrogenase (LsGDH) from Lysinibacillus sphaericus G10 to improve its thermal stability by introduction of disulfide bridges between subunits. One out of the eleven mutants, designated as DS255, displayed significantly enhanced thermal stability with considerable soluble expression and high specific activity. It was extremely stable at pH ranging from 4.5 to 10.5, as it retained nearly 100% activity after incubating at different buffers for 1h. Mutant DS255 also exhibited high thermostability, having a half-life of 9900min at 50°C, which was 1868-fold as that of its wild type. Moreover, both of the increased free energy of denaturation and decreased entropy of denaturation of DS255 suggested that the enzyme structure was stabilized by the engineered disulfide bonds. On account of its robust stability, mutant DS255 would be a competitive candidate in practical applications of chiral chemicals synthesis, biofuel cells and glucose biosensors.
authors: Institute of Microbiology, College of Life Science, Zhejiang University, Hangzhou 310058, China. Electronic address: htding@outlook.com.
authors: Haitao
authors: H
authors: Ding
authors: Fen
authors: F
authors: Gao
authors: Danfeng
authors: D
authors: Liu
authors: Zeli
authors: Z
authors: Li
authors: Xiaohong
authors: X
authors: Xu
authors: Min
authors: M
authors: Wu
authors: Yuhua
authors: Y
authors: Zhao
mesh_terms: D001406:Bacillaceae / Q000201:enzymology / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry; D004795:Enzyme Stability; D042843:Glucose 1-Dehydrogenase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006207:Half-Life; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011489:Protein Denaturation; D015202:Protein Engineering; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017385:Sequence Homology; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D004220:Disulfides; D021122:Protein Subunits; D011994:Recombinant Proteins; D042843:Glucose 1-Dehydrogenase; D003545:Cysteine
keywords: BmGDH-IWG3; Disulfide bond; GDH; GDH from Bacillus megaterium IWG3; GDH from Lysinibacillus sphaericus G10; Glucose 1-dehydrogenase; LsGDH; NAD; NAD(P)-dependent glucose 1-dehydrogenase; NADP; Protein engineering; Protein stability; nicotinamide adenine dinucleotide; nicotinamide adenine dinucleotide phosphate