Investigating the binding of beta-1,4-galactan to Bacillus licheniformis beta-1,4-galactanase by crystallography and computational modeling.

Microbial β-1,4-galactanases are glycoside hydrolases belonging to family 53, which degrade galactan and arabinogalactan side chains in the hairy regions of pectin, a major plant cell wall component. They belong to the larger clan GH-A of glycoside hydrolases, which cover many different poly- and oligosaccharidase specificities. Crystallographic complexes of Bacillus licheniformis β-1,4-galactanase and its inactive nucleophile mutant have been obtained with methyl-β(1→4)-galactotetraoside, providing, for the first time, information on substrate binding to the aglycone side of the β-1,4-galactanase substrate binding groove. Using the experimentally determined subsites as a starting point, a β(1→4)-galactononaose was built into the structure and subjected to molecular dynamics simulations giving further insight into the residues involved in the binding of the polysaccharide from subsite −4 to +5. In particular, this analysis newly identified a conserved β-turn, which contributes to subsites −2 to +3. This β-turn is unique to family 53 β-1,4-galactanases among all clan GH-A families that have been structurally characterized and thus might be a structural signature for endo-β-1,4-galactanase specificity. Proteins 2009. © 2008 Wiley-Liss, Inc.