Structural Features of a Bacteroidetes-Affiliated Cellulase Linked with a Polysaccharide Utilization Locus.

ABSTRACT: Previous gene-centric analysis of a cow rumen metagenome revealed the first potentially cellulolytic polysaccharide utilization locus, of which the main catalytic enzyme (AC2aCel5A) was identified as a glycoside hydrolase (GH) family 5 endo-cellulase. Here we present the 1.8 Å three-dimensional structure of AC2aCel5A, and characterization of its enzymatic activities. The enzyme possesses the archetypical (?/?)8-barrel found throughout the GH5 family, and contains the two strictly conserved catalytic glutamates located at the C-terminal ends of ?-strands 4 and 7. The enzyme is active on insoluble cellulose and acts exclusively on linear ?-(1,4)-linked glucans. Co-crystallization of a catalytically inactive mutant with substrate yielded a 2.4 Å structure showing cellotriose bound in the -3 to -1 subsites. Additional electron density was observed between Trp178 and Trp254, two residues that form a hydrophobic "clamp", potentially interacting with sugars at the +1 and +2 subsites. The enzyme's active-site cleft was narrower compared to the closest structural relatives, which in contrast to AC2aCel5A, are also active on xylans, mannans and/or xyloglucans. Interestingly, the structure and function of this enzyme seem adapted to less-substituted substrates such as cellulose, presumably due to the insufficient space to accommodate the side-chains of branched glucans in the active-site cleft.

SUBMITTER: Naas AE 

PROVIDER: S-EPMC4488959 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature