A Single Point Mutation Converts GH84 O-GlcNAc Hydrolases into Phosphorylases: Experimental and Theoretical Evidence

David Teze*, Joan Coines, Lluís Raich, Valentina Kalichuk, Claude Solleux, Charles Tellier, Corinne André-Miral, Birte Svensson, Carme Rovira

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review


Glycoside hydrolases and phosphorylases are two major classes of enzymes responsible for the cleavage of glycosidic bonds. Here we show that two GH84 O-GlcNAcase enzymes can be converted to efficient phosphorylases by a single point mutation. Noteworthy, the mutated enzymes are over 10-fold more active than naturally occurring glucosaminide phosphorylases. We rationalize this novel transformation using molecular dynamics and QM/MM metadynamics methods, showing that the mutation changes the electrostatic potential at the active site and reduces the energy barrier for phosphorolysis by 10 kcal·mol-1. In addition, the simulations unambiguously reveal the nature of the intermediate as a glucose oxazolinium ion, clarifying the debate on the nature of such a reaction intermediate in glycoside hydrolases operating via substrate-assisted catalysis.
Original languageEnglish
JournalJournal of the American Chemical Society
Issue number5
Pages (from-to)2120-2124
Number of pages5
Publication statusPublished - 2020

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