title: Cloning and expression of thermo-alkali-stable laccase of Bacillus licheniformis in Pichia pastoris and its characterization.
abstract: A thermo-alkali-stable laccase gene from Bacillus licheniformis was cloned and expressed in Pichia pastoris. The recombinant laccase was secreted into the culture medium with a maximum activity of 227.9 U/L. The purified laccase is a monomeric glycoprotein, and its molecular weight was estimated to be 65 kDa on SDS-PAGE after deglycosylation. Optimal enzyme activity was observed at pH 6.2 and 70°C with syringaldazine as substrate. The recombinant laccase was highly stable in the pH range 7-9 after 10 days at 30°C. The enzyme displayed remarkable thermostability at 50-70°C, with a half-life of inactivation at 70°C of 6.9 h. It also exhibited high tolerance to NaCl and organic solvents like the native spore laccase. The purified laccase could rapidly decolorize reactive blue 19, reactive black 5 and indigo carmine in the presence of acetosyringone. More than 93% of the tested dyes were decolorized in 4 h at pH 9.0.
authors.affiliation: College of Life Sciences, Northeast Forestry University, Harbin 150040, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Lu
authors.forename: Tian-Nv
authors.initials: TN
authors.lastname: Wang
authors.forename: Teng-Fei
authors.initials: TF
authors.lastname: Xu
authors.forename: Jing-Yao
authors.initials: JY
authors.lastname: Wang
authors.forename: Chun-Lei
authors.initials: CL
authors.lastname: Wang
authors.forename: Min
authors.initials: M
authors.lastname: Zhao
mesh_terms: D001407:Bacillus / Q000187:drug effects / Q000201:enzymology*; D003001:Cloning, Molecular; D003116:Color; D004396:Coloring Agents / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability / Q000187:drug effects; D005798:Genes, Bacterial / Q000235:genetics; D006835:Hydrazones / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D042845:Laccase / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D009930:Organic Chemicals / Q000494:pharmacology; D010843:Pichia / Q000235:genetics*; D011994:Recombinant Proteins / Q000378:metabolism; D012997:Solvents / Q000494:pharmacology; D013171:Spores, Bacterial / Q000187:drug effects / Q000201:enzymology; D013379:Substrate Specificity / Q000187:drug effects; D013696:Temperature; D013997:Time Factors
chemical_list: D004396:Coloring Agents; D004791:Enzyme Inhibitors; D006835:Hydrazones; D009930:Organic Chemicals; D011994:Recombinant Proteins; D012997:Solvents; C009370:syringaldazine; D042845:Laccase