title: Essential role of a family-32 carbohydrate-binding module in substrate recognition by Clostridium thermocellum mannanase CtMan5A.
abstract: The family-5 glycoside hydrolase domain (GH5) and the family-32 carbohydrate-binding module (CBM32) of Clostridium thermocellum mannanase CtMan5A, along with their genetically inactivated derivatives, were collectively or separately expressed. Their catalytic and substrate-binding abilities were measured to investigate importance of CBM32 in substrate recognition by CtMan5A. Characterization of the truncated derivatives of CtMan5A and isothermal calorimetry analysis of the interaction between the inactivated proteins and mannooligosaccharides suggested that GH5 and CBM32 collectively formed a substrate-binding site capable of accommodating a mannotetraose unit in CtMan5A. This suggested that CBM32 directly participated in the substrate recognition required for catalytic action.
authors.affiliation: Applied Microbiology Laboratory, Graduate School of Bioresources, Mie University, Tsu, Japan.
authors.forename: Kimiya
authors.initials: K
authors.lastname: Mizutani
authors.affiliation: Applied Microbiology Laboratory, Graduate School of Bioresources, Mie University, Tsu, Japan.
authors.forename: Makiko
authors.initials: M
authors.lastname: Sakka
authors.affiliation: Applied Microbiology Laboratory, Graduate School of Bioresources, Mie University, Tsu, Japan.
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Kimura
authors.affiliation: Applied Microbiology Laboratory, Graduate School of Bioresources, Mie University, Tsu, Japan. Electronic address: sakka@bio.mie-u.ac.jp.
authors.forename: Kazuo
authors.initials: K
authors.lastname: Sakka
mesh_terms: D000327:Adsorption; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites; D048013:Acetivibrio thermocellus / Q000201:enzymology*; D006868:Hydrolysis; D008361:Mannosidases / Q000737:chemistry*; D009844:Oligosaccharides / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary
chemical_list: D001426:Bacterial Proteins; D009844:Oligosaccharides; C048068:mannopentaose; D008361:Mannosidases
keywords: Carbohydrate-binding module; Clostridium thermocellum; Mannanase; Substrate recognition