title: Overexpression and characterization of a novel endo-β-1,3(4)-glucanase from thermophilic fungus Humicola insolens Y1.
abstract: A novel endo-β-1,3(4)-glucanase gene, cel16A, was cloned from the fungus Humicola insolens Y1. The 988-bp full-length gene encoded a 286-residue polypeptide consisting of a putative signal peptide of 20 residues and a catalytic domain belonging to glycosyl hydrolase family 16. It was successfully overexpressed in Pichia pastoris GS115. The purified recombinant Cel16A exhibited highest specific activity toward barley β-glucan, followed by lichenan and laminarin, but not toward CMC-Na, birchwood xylan, Avicel and filter paper, indicating that Cel16A is an endo-β-1,3(4)-glucanases. Recombinant Cel16A had a pH optimum at 5.5 and a temperature optimum at 55 °C with a specific activity of 693 U/mg toward barley β-glucan. It exhibited good stability over pH 5.0-9.0 and at temperatures up to 50 °C, retaining over 80% maximum activity. The Kmand Vmaxvalues of Cel16A for barley β-glucan were 0.91 mg ml-1and 1530 μmol min-1·mg-1, respectively. All these favorable enzymatic properties of Cel16A make it a good candidate for applications in various industries.
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, PR China.
authors.forename: Jinyang
authors.initials: J
authors.lastname: Li
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, PR China.
authors.forename: Xinxin
authors.initials: X
authors.lastname: Xu
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, PR China.
authors.forename: Pengjun
authors.initials: P
authors.lastname: Shi
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, PR China.
authors.forename: Bo
authors.initials: B
authors.lastname: Liu
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, PR China.
authors.forename: Yuhong
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, PR China. Electronic address: zhangwei02@caas.cn.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
mesh_terms: D000595:Amino Acid Sequence; D020134:Catalytic Domain; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D044644:Endo-1,3(4)-beta-Glucanase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D057075:Enzyme Assays; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015870:Gene Expression; D005822:Genetic Vectors / Q000737:chemistry / Q000378:metabolism; D005936:Glucans / Q000378:metabolism; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D010843:Pichia / Q000235:genetics* / Q000378:metabolism; D021382:Protein Sorting Signals / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016415:Sequence Alignment; D020037:Sordariales / Q000737:chemistry* / Q000201:enzymology; D013379:Substrate Specificity; D047071:beta-Glucans / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D005936:Glucans; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D047071:beta-Glucans; C008247:laminaran; D044644:Endo-1,3(4)-beta-Glucanase; C015968:lichenin
keywords: Glycosyl hydrolase family 16; Humicola insolens Y1; β-1,3(4)-Glucanase