The X-ray crystal structure of the Trichoderma reesei family 12 endoglucanase 3, Cel12A, at 1.9 A resolution

J Mol Biol. 2001 Apr 27;308(2):295-310. doi: 10.1006/jmbi.2001.4583.


We present the three-dimensional structure of Trichoderma reesei endoglucanase 3 (Cel12A), a small, 218 amino acid residue (24.5 kDa), neutral pI, glycoside hydrolase family 12 cellulase that lacks a cellulose-binding module. The structure has been determined using X-ray crystallography and refined to 1.9 A resolution. The asymmetric unit consists of six non-crystallographic symmetry-related molecules that were exploited to improve initial multiple isomorphous replacement phasing, and subsequent structure refinement. The enzyme contains one disulfide bridge and is glycosylated at Asp164 by a single N-acetyl glucosamine residue. The protein has the expected fold for a glycoside hydrolase clan-C family 12 enzyme. It contains two beta-sheets, of six and nine strands, packed on top of one another, and one alpha-helix. The concave surface of the nine-stranded beta-sheet forms a large substrate-binding groove in which the active-site residues are located. In the active site, we find a carboxylic acid trio, similar to that of glycoside hydrolase families 7 and 16. The strictly conserved Asp99 hydrogen bonds to the nucleophile, the invariant Glu116. The binding crevice is lined with both aromatic and polar amino acid side-chains which may play a role in substrate binding. The structure of the fungal family 12 enzyme presented here allows a complete structural characterization of the glycoside hydrolase-C clan.

MeSH terms

  • Amino Acid Sequence
  • Bacterial Proteins / chemistry
  • Binding Sites
  • Cellulase / chemistry*
  • Crystallography, X-Ray
  • Disulfides / metabolism
  • Glycosylation
  • Hydrogen Bonding
  • Models, Molecular
  • Molecular Sequence Data
  • Protein Folding
  • Protein Structure, Secondary
  • Sequence Alignment
  • Trichoderma / enzymology*


  • Bacterial Proteins
  • Disulfides
  • Cellulase

Associated data

  • PDB/1H8V