title: A novel family 9 beta-1,3(4)-glucanase from thermoacidophilic Alicyclobacillus sp. A4 with potential applications in the brewing industry.
abstract: An endo-beta-1,3(4)-glucanase gene, Agl9A, was cloned from Alicyclobacillus sp. A4 and expressed in Pichia pastoris. Its deduced amino acid sequence shared the highest identity (48%) with an endo-beta-1,4-glucansae from Alicyclobacillus acidocaldarius that belongs to family 9 of the glycoside hydrolases. The purified recombinant Agl9A exhibited relatively wide substrate specificity, including lichenan (109%), barley beta-glucan (100%), CMC-Na (15.02%), and laminarin (6.19%). The optimal conditions for Agl9A activity were pH 5.8 and 55 degrees C. The enzyme was stable over a broad pH range (>60% activity retained after 1-h incubation at pH 3.8-11.2) and at 60 degrees C (>70% activity retained after 1-h incubation). Agl9A was highly resistant to various neutral proteases (e.g., trypsin, alpha-chymotrypsin, and collagenase) and Neutrase 0.8L (Novozymes), a protease widely added to the mash. Under simulated mashing conditions, addition of Agl9A (20 U/ml) or a commercial xylanase (200 U/ml) reduced the filtration rate (26.71% and 20.21%, respectively) and viscosity (6.12% and 4.78%, respectively); furthermore, combined use of Agl9A (10 U/ml) and the xylanase (100 U/ml) even more effectively reduced the filtration rate (31.73%) and viscosity (8.79%). These characteristics indicate that Agl9A is a good candidate to improve glucan degradation in the malting and brewing industry.
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: Yingguo
authors.initials: Y
authors.lastname: Bai
authors.forename: Jianshe
authors.initials: J
authors.lastname: Wang
authors.forename: Zhifang
authors.initials: Z
authors.lastname: Zhang
authors.forename: Pengjun
authors.initials: P
authors.lastname: Shi
authors.forename: Huiying
authors.initials: H
authors.lastname: Luo
authors.forename: Huoqing
authors.initials: H
authors.lastname: Huang
authors.forename: Chunliang
authors.initials: C
authors.lastname: Luo
authors.forename: Bin
authors.initials: B
authors.lastname: Yao
mesh_terms: D056450:Alicyclobacillus / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D044644:Endo-1,3(4)-beta-Glucanase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004795:Enzyme Stability; D005285:Fermentation; D005936:Glucans / Q000378:metabolism; D006358:Hot Temperature; D007218:Industrial Microbiology; D007700:Kinetics; D008969:Molecular Sequence Data; D005810:Multigene Family; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D005936:Glucans; D044644:Endo-1,3(4)-beta-Glucanase