title: Crystal structures of A. acidocaldarius endoglucanase Cel9A in complex with cello-oligosaccharides: strong -1 and -2 subsites mimic cellobiohydrolase activity.
abstract: Alicyclobacillus acidocaldarius endoglucanase Cel9A (AaCel9A) is an inverting glycoside hydrolase with beta-1,4-glucanase activity on soluble polymeric substrates. Here, we report three X-ray structures of AaCel9A: a ligand-free structure at 1.8 A resolution and two complexes at 2.66 and 2.1 A resolution, respectively, with cellobiose obtained by co-crystallization and with cellotetraose obtained by the soaking method. AaCel9A forms an (alpha/alpha)(6)-barrel like other glycoside hydrolase family 9 enzymes. When cellobiose is used as a ligand, three glucosyl binding subsites are occupied, including two on the glycone side, while with cellotetraose as a ligand, five subsites, including four on the glycone side, are occupied. A structural comparison showed no conformational rearrangement of AaCel9A upon ligand binding. The structural analysis demonstrates that of the four minus subsites identified, subsites -1 and -2 show the strongest interaction with bound glucose. In conjunction with the open active-site cleft of AaCel9A, this is able to reconcile the previously observed cleavage of short-chain oligosaccharides with cellobiose as main product with the endo mode of action on larger polysaccharides.
authors.affiliation: Laboratoire d'Enzymologie et Biochimie Structurales, CNRS, Avenue de la Terrasse, 91198 Gif-sur-Yvette, France.
authors.forename: Kelvin
authors.initials: K
authors.lastname: Eckert
authors.forename: Armelle
authors.initials: A
authors.lastname: Vigouroux
authors.forename: Leila
authors.initials: L
authors.lastname: Lo Leggio
authors.forename: Solange
authors.initials: S
authors.lastname: Moréra
mesh_terms: D001105:Archaea / Q000201:enzymology*; D019843:Archaeal Proteins / Q000737:chemistry*; D001665:Binding Sites; D055162:Biocatalysis; D002475:Cellobiose / Q000737:chemistry*; D002482:Cellulose / Q000031:analogs & derivatives* / Q000737:chemistry; D043366:Cellulose 1,4-beta-Cellobiosidase / Q000737:chemistry* / Q000378:metabolism; D018360:Crystallography, X-Ray; D008024:Ligands; D008670:Metals / Q000378:metabolism; D018716:Molecular Mimicry; D017433:Protein Structure, Secondary; D055672:Static Electricity; D013379:Substrate Specificity; D013780:Tetroses / Q000737:chemistry*
chemical_list: D019843:Archaeal Proteins; D008024:Ligands; D008670:Metals; D013780:Tetroses; D002475:Cellobiose; C048468:cellotetraose; D002482:Cellulose; D043366:Cellulose 1,4-beta-Cellobiosidase