title: High-level expression of a novel Penicillium endo-1,3(4)-β-D-glucanase with high specific activity in Pichia pastoris.
abstract: A novel endo-1,3(4)-β-D-glucanase gene (bgl16C1) from Penicillium pinophilum C1 was cloned and sequenced. The 945-bp full-length gene encoded a 315-residue polypeptide consisting of a putative signal peptide of 18 residues and a catalytic domain belonging to glycosyl hydrolase family 16. The deduced amino acid sequence showed the highest identity (82%) with the putative endo-1,3(4)-β-glucanase from Talaromyces stipitatus ATCC 10500 and 60% identity with the characterized β-1,3(4)-glucanase from Paecilomyces sp. FLH30. The gene was successfully overexpressed in Pichia pastoris. Recombinant Bgl16C1 constituted 95% of total secreted proteins (2.61 g l⁻¹) with activity of 28,721 U ml⁻¹ in a 15-l fermentor. The purified recombinant Bgl16C1 had higher specific activity toward barley β-glucan (12,622 U mg⁻¹) than all known glucanases and also showed activity against lichenan and laminarin. The enzyme was optimally active at pH 5.0 and 55°C and exhibited good stability over a broad acid and alkaline pH range (>85% activity at pH 3.0-7.0 and even 30% at pH 11.0). All these favorable enzymatic properties make it attractive for potential applications in various industries.
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, Beijing 100081, People's Republic of China.
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Chen
authors.forename: Kun
authors.initials: K
authors.lastname: Meng
authors.forename: Pengjun
authors.initials: P
authors.lastname: Shi
authors.forename: Yingguo
authors.initials: Y
authors.lastname: Bai
authors.forename: Huiying
authors.initials: H
authors.lastname: Luo
authors.forename: Huoqing
authors.initials: H
authors.lastname: Huang
authors.forename: Tiezheng
authors.initials: T
authors.lastname: Yuan
authors.forename: Peilong
authors.initials: P
authors.lastname: Yang
authors.forename: Bin
authors.initials: B
authors.lastname: Yao
mesh_terms: D000595:Amino Acid Sequence; D057005:Bioengineering; D003001:Cloning, Molecular; D044644:Endo-1,3(4)-beta-Glucanase / Q000096:biosynthesis* / Q000302:isolation & purification* / Q000378:metabolism; D005936:Glucans / Q000378:metabolism; D008969:Molecular Sequence Data; D010143:Paecilomyces / Q000201:enzymology; D010407:Penicillium / Q000201:enzymology*; D010843:Pichia / Q000235:genetics*; D011134:Polysaccharides / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D047071:beta-Glucans / Q000378:metabolism
chemical_list: D005936:Glucans; D011134:Polysaccharides; D011994:Recombinant Proteins; D047071:beta-Glucans; C008247:laminaran; D044644:Endo-1,3(4)-beta-Glucanase; C015968:lichenin