title: Identification and characterization of a new acid-stable endoglucanase from a metagenomic library.
abstract: A new endoglucanase gene cel124 was cloned from a metagenomic library and expressed in Escherichiacoli. Catalytic triad analysis showed that the catalytic triad sites were different from the known endoglucanases. Cel124, a 34 kDa protein, exhibited a specific activity (29.08 U mg(-1)) toward 1% of sodium carboxymethyl cellulose and was stable at 50 °C for 30 min. The optimal temperature and pH for its catalytic activity were 50 °C and pH 5.5 respectively. Cel124 could hydrolyze soluble cellulose, but not insoluble cellulose or other polysaccharides. The kinetic parameters (5.63 mg ml(-1) for Km and 0.0397 mmol min(-1) mg(-1) for Vmax) were measured. 3M NaCl in the system could increase its activity by 2 fold. Site-directed mutation and circular dichroism spectra test suggested that the residue (Glu41) was essential for its activity, might be a potential active site. Based on our data, we proposed that Cel124 might represent a new type of endoglucanase.
authors.affiliation: The Collaborative Innovation Center of Bio-resource Green Transformation, The College of Life Sciences, Hubei University, Wuhan 430062, China.
authors.forename: La
authors.initials: L
authors.lastname: Xiang
authors.affiliation: The College of Life Sciences, Central China Normal University, Wuhan 430079, China.
authors.forename: Aiying
authors.initials: A
authors.lastname: Li
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Chaoguang
authors.initials: C
authors.lastname: Tian
authors.affiliation: The Collaborative Innovation Center of Bio-resource Green Transformation, The College of Life Sciences, Hubei University, Wuhan 430062, China.
authors.forename: Yuling
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: The Collaborative Innovation Center of Bio-resource Green Transformation, The College of Life Sciences, Hubei University, Wuhan 430062, China. Electronic address: zhangguimin@hubu.edu.cn.
authors.forename: Guimin
authors.initials: G
authors.lastname: Zhang
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Yanhe
authors.initials: Y
authors.lastname: Ma
mesh_terms: D000595:Amino Acid Sequence; D002480:Cellulase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002482:Cellulose / Q000378:metabolism; D003001:Cloning, Molecular / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics*; D015723:Gene Library; D016680:Genome, Bacterial; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D056186:Metagenomics / Q000379:methods*; D010802:Phylogeny; D012230:Rhizobiaceae / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D002482:Cellulose; D002480:Cellulase
keywords: Endoglucanase; Enzymatic assay; Metagenomic library; Prokaryotic expression