Mechanism and Catalytic Site Atlas

Introduction

Binding of the alpha-sialic acid to NA, forming the Michaelis complex. Triad of arginine residues binds to and stabilises the carboxylate group of the substrate by strong ionic interactions with their guanidinium groups. Distortion of the substrate pyranose ring from chair to boat conformation. The glycosidic oxygen is raised from its equatotiral position to point vertically out of the active site and lie close to an Asp 151 carboxylate oxygen. The N-acetyl group of the substrate is firmly bound in the hydrophobic pocket of the enzyme active site, with the C2 carbon forced into planar conformation. The glycerol side chain is hydrogen bonded to Glu 275 through the C8 and C9 hydroxyls. Proton donation to the glycosidic oxygen from a neighbouring water molecule, activated by a hydrogen bond to the Asp 151 residue. Glu 277 activates Tyr 412 for nucleophilic attack by an acid/base interaction between the residues. Tyr 412 is close to, and nucleophilically attacks the anomeric carbon centre of the sialic acid residue. Formation and stabilisation of the endocyclic sialosyl cation transition state. Formation and release of the sialic acid as the alpha-monomer