Understanding protein structure-function relationships in family 47 α-1,2-mannosidases through computational docking of ligands

Family 47 alpha-1,2-mannosidases are crucial enzymes involved in N-glycan maturation in the endoplasmic reticula and Golgi apparati of eukaryotic cells. High-resolution crystal structures of the human and yeast endoplasmic reticulum alpha-1,2-mannosidases have been recently determined, the former complexed with the inhibitors 1-deoxymannojirimycin and kifunensine, both of which bind in its active site in the unusual C-1(4) conformation. However, unambiguous identification of the catalytic proton donor and nucleophile involved in glycoside bond hydrolysis was not possible from this structural information. In this work, alpha-D-galactose, alpha-D-glucose, and alpha-D-mannose were computationally docked in the active site in the energetically stable C-4(1) conformation as well as in the C-1(4) conformation to compare their interaction energetics. From these docked structures, a model for substrate and conformer selectivity based on the dimensions of the active site was proposed. alpha-D-Galactopyranosyl(1-->2)-alpha-D-mannopyranose, alpha-D-glucopyranosyl(1-->2)-alpha-D-mannopyranose, and a-D-mannopyranosyl(1-->2)-alpha-D-mannopyranose were also docked into the active site with their nonreducing-end residues in the C-1(4) and E-4 (representing the transition state) conformations. Based on the docked structure of alpha-D-mannopyranosyl-E-4-(1-->2)-alpha-D-mannopyranose, the catalytic acid and base are Glu132 and Glu435, respectively. (C) 2002 Wiley-Liss, Inc.