title: Expression, purification and thermostability of MBP-chondroitinase ABC I from Proteus vulgaris.
abstract: Chondroitinase ABC I (ChSase ABC I) which can degrade chondroitin sulfate (CS) and other glycosaminoglycan to oligosaccharide or unsaturated disaccharide, was fusionally expressed with maltose-binding protein (MBP) in Escherichia coli BL21(DE3) (E. coli BL21(DE3)) and purified for the first time in this study. The result showed that the productivity of recombinant MBP-ChSase ABC I was 3180 IU/(L fermentation liquor) with CS A as substrate, and the productivity might be the highest level when compared to the reported ones. The specific activity of recombinant MBP-ChSase ABC I was 76 IU/(mg protein) after purification. The Vmax, Km and kcat were 18.7 ± 0.3 μmol/Ls, 73.1 ± 4.1 μmol/L and 586.7 ± 10.8 s(-1), respectively. Enzyme activity of the purified enzyme remained about 78% after 210 min when the enzyme incubated at 30 °C. This study introduces a rapid method for highly expressing ChSase ABC I, and the method could be adopted in the process of industrial production. Furthermore the investigation of thermostability might lead to an important guide in clinical treatment.
authors: State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, No. 15 East Road of North Third Ring, Chao Yang District, Beijing 100029, China.
authors: Zhenya
authors: Z
authors: Chen
authors: Department of Biotechnology, Beijing Polytechnic, No. 1 Taiyanggong Shaoyaoju, Chao Yang District, Beijing 100029, China. Electronic address: liyeyashi@163.com.
authors: Ye
authors: Y
authors: Li
authors: State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, No. 15 East Road of North Third Ring, Chao Yang District, Beijing 100029, China. Electronic address: yuanqp@mail.buct.edu.cn.
authors: Qipeng
authors: Q
authors: Yuan
mesh_terms: D055162:Biocatalysis; D019765:Chondroitin ABC Lyase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D004795:Enzyme Stability; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D058619:Maltose-Binding Proteins / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011514:Proteus vulgaris / Q000201:enzymology* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification* / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013696:Temperature
chemical_list: D058619:Maltose-Binding Proteins; D011993:Recombinant Fusion Proteins; D019765:Chondroitin ABC Lyase
keywords: Characterization; Expression; MBP-chondrotinase ABC I; Productivity; Thermostability