O-/N-/S-specificity in glycosyltransferase catalysis: From mechanistic understanding to engineering

David Tezé*, Joan Coines, Folmer Fredslund, Kshatresh D. Dubey, Gonzalo N. Bidart, Paul D. Adams, John E. Dueber, Birte Svensson, Carme Rovira*, Ditte Hededam Welner*

*Corresponding author for this work

Research output: Contribution to journalLetterpeer-review

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Glycosyltransferases (GTs) catalyze the formation of glycosidic bonds in carbohydrates and glycoconjugates, with various outcomes depending not only on the acceptor molecules they bind but also on the type of glycosidic bond they form (C−O, C−N, C−S, or C−C). Here we show that the glucosyltransferase UGT1 from the indigo plant Polygonum tinctorium catalyzes either N-, O-, or S-glycosylation with similar rates. We solve the structure of the enzyme in complex with its donor and acceptor substrates and elucidate the molecular basis of N-, O-, and S-specificities using experimental mutagenesis and QM/MM simulations, revealing distinct mechanisms for N-, O-, and S-glycosylation. We also show that the active site can be engineered to increase or favor one of the three glycosylation activities over another. These results will foster the design of more active and specific enzyme variants for production of glycosides.
Original languageEnglish
JournalACS Catalysis
Issue number3
Pages (from-to)1810-1815
Publication statusPublished - 2021


  • Enzymes
  • O-glycosylation
  • N-gycosylation
  • S-glycosylation
  • Carbohydrates
  • Glycosyltransferases
  • Quantum mechanics/molecular mechanics


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