title: Characterization of a multi-function processive endoglucanase CHU_2103 from Cytophaga hutchinsonii.
abstract: Cytophaga hutchinsonii is a Gram-negative gliding bacterium which can efficiently degrade crystalline cellulose by an unknown strategy. Genomic analysis suggests the C. hutchinsonii genome lacks homologs to an obvious exoglucanase that previously seemed essential for cellulose degradation. One of the putative endoglucanases, CHU_2103, was successfully expressed in Escherichia coli JM109 and identified as a processive endoglucanase with transglycosylation activity. It could hydrolyze carboxymethyl cellulose (CMC) into cellodextrins and rapidly decrease the viscosity of CMC. When regenerated amorphous cellulose (RAC) was degraded by CHU_2103, the ratio of the soluble to insoluble reducing sugars was 3.72 after 3 h with cellobiose and cellotriose as the main products, indicating that CHU_2103 was a processive endoglucanase. CHU_2103 could degrade cellodextrins of degree of polymerization ≥3. It hydrolyzed p-nitrophenyl β-D-cellodextrins by cutting glucose or cellobiose from the non-reducing end. Meanwhile, some larger-molecular-weight cellodextrins could be detected, indicating it also had transglycosylation activity. Without carbohydrate-binding module (CBM), CHU_2103 could bind to crystalline cellulose and acted processively on it. Site-directed mutation of CHU_2103 demonstrated that the conserved aromatic amino acid W197 in the catalytic domain was essential not only for its processive activity, but also its cellulose binding ability.
authors.affiliation: State Key Laboratory of Microbial Technology, School of Life Science, Shandong University, Jinan, 250100, China.
authors.forename: Cong
authors.initials: C
authors.lastname: Zhang
authors.forename: Ying
authors.initials: Y
authors.lastname: Wang
authors.forename: Zhe
authors.initials: Z
authors.lastname: Li
authors.forename: Xiangru
authors.initials: X
authors.lastname: Zhou
authors.forename: Weican
authors.initials: W
authors.lastname: Zhang
authors.forename: Yue
authors.initials: Y
authors.lastname: Zhao
authors.forename: Xuemei
authors.initials: X
authors.lastname: Lu
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002480:Cellulase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002482:Cellulose / Q000031:analogs & derivatives / Q000378:metabolism; D003589:Cytophaga / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D003912:Dextrins / Q000378:metabolism; D004795:Enzyme Stability; D007700:Kinetics; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D003912:Dextrins; D002482:Cellulose; C068994:cellodextrin; D002480:Cellulase