title: Cloning and characterization of a cold-adapted endo-1,5-α-L-arabinanase from Paenibacillus polymyxa and rational design for acidic applicability.
abstract: AbnZ1, with optimal pH of 6.0 and optimal temperature of 40 °C, is a cold-adapted endo-1,5-α-L-arabinanase encoded by the gene abnZ1 from Paenibacillus polymyxa Z6. The specific activity of AbnZ1 remained 54.1% of maximum at 5 °C. To apply AbnZ1 in acidic conditions, three basic hsitidine (His) residues, His(48), His(218), and His(297), around the catalytic domain were selected as mutation sites, which were replaced with Asp, Glu, Arg, and Lys, respectively, to yield 12 mutants, H48D/E/R/K, H218D/E/R/K, and H297D/E/R/K. The optimum pH of mutant H218D shifted toward the acidic direction by 0.5 unit, and the relative activity was enhanced from 20.4 to 55.7% at pH 5.0. Furthermore, the specific activity of H218D in optimal conditions was 82.6 U/mg versus that of wild type, 73.4 U/mg, and the K(m) decreased from 11.9 to 7.1 mg/mL. This work provided an arabinanase candidate for juice clarification and pectin extraction.
authors: State Key Laboratory of Bioreactor Engineering, Department of Food Science and Technology, School of Biotechnology, East China University of Science and Technology , Shanghai 200237, People's Republic of China.
authors: Shaohua
authors: S
authors: Wang
authors: Ying
authors: Y
authors: Yang
authors: Rujian
authors: R
authors: Yang
authors: Jian
authors: J
authors: Zhang
authors: Min
authors: M
authors: Chen
authors: Shingo
authors: S
authors: Matsukawa
authors: Jingli
authors: J
authors: Xie
authors: Dongzhi
authors: D
authors: Wei
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D001628:Beverages / Q000032:analysis; D003001:Cloning, Molecular; D003080:Cold Temperature; D004795:Enzyme Stability; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D056507:Paenibacillus / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D016415:Sequence Alignment
chemical_list: D001426:Bacterial Proteins; D006026:Glycoside Hydrolases; C082526:arabinan endo-1,5-alpha-L-arabinosidase
keywords: arabinanase; cold-adapted; molecular modeling; pH stability; site-directed mutagenesis