Ganoderma lucidum is a saprotrophic white-rot fungus which contains a rich set of cellulolytic enzymes. Here, we screened an array of potential 1,4-beta-endoglucanases from G. lucidum based on the gene annotation library and found that one candidate gene, GlCel5A, exhibits CMC-hydrolyzing activity. The recombinant GlCel5A protein expressed in Pichia pastoris is able to hydrolyze CMC and beta-glucan but not xylan and mannan. The enzyme exhibits optimal activity at 60 degrees C and pH 3-4, and retained 50% activity at 80 and 90 degrees C for at least 15 and 10 min. The crystal structure of GlCel5A and its complex with cellobiose, solved at 2.7 and 2.86 angstrom resolution, shows a classical (beta/alpha)(8) TIM-barrel fold as seen in other members of glycoside hydrolase family 5. The complex structure contains a cellobiose molecule in the +1 and +2 subsites, and reveals the interactions with the positive sites of the enzyme. Collectively, the present work provides the first comprehensive characterization of an endoglucanase from G. lucidum that possesses properties for industrial applications, and strongly encourages further studying in the cellulolytic enzyme system of G. lucidum. (C) 2016 Published by Elsevier Inc.