Oligosaccharide binding to family 11 xylanases: both covalent intermediate and mutant product complexes display ^2,5B conformations at the active centre

The glycoside hydrolase sequence-based classification reveals two families of enzymes which hydrolyse the β-1,4-linked backbone of xylan, xylanases, termed families GH-10 and GH-11. Family GH-11 xylanases are intriguing in that catalysis is performed via a covalent intermediate adopting an unusual ^2,5B (boat) conformation, a conformation which also fulfils the stereochemical constraints of the oxocarbenium ion-like transition state. Here, the 1.9 Å structure of a nucleophile, E94A, mutant of the Xyn11 from Bacillus agaradhaerens in complex with xylotriose is presented. Intriguingly, this complex also adopts the ^2,5B conformation in the −1 subsite, with the vacant space provided by the Glu→Ala mutation allowing the sugar to adopt the α-configuration at C1. The structure of the covalent 2-deoxy-2-fluoroxylobiosyl-enzyme intermediate has been extended to atomic (1.1 Å) resolution.