Rational Enzyme Design without Structural Knowledge: A Sequence-Based Approach for Efficient Generation of Transglycosylases

David Teze*, Jiao Zhao, Mathias Wiemann, Zubaida G. A. Kazi, R. Lupo, Birgitte Zeuner, Marlène Vuillemin, Mette E. Rønne, Göran Carlström, Jens Ø. Duus, Yves-Henri Sanejouand, Michael J. O'Donohue, Eva Nordberg Karlsson, Régis Fauré, Henrik Stålbrand, Birte Svensson*

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review


Glycobiology is dogged by the relative scarcity of synthetic, defined oligosaccharides. Enzyme-catalysed glycosylation using glycoside hydrolases is feasible but is hampered by the innate hydrolytic activity of these enzymes. Protein engineering is useful to remedy this, but it usually requires prior structural knowledge of the target enzyme, and/or relies on extensive, time-consuming screening and analysis. Here, a straightforward strategy that involves rational rapid in silico analysis of protein sequences is described. The method pinpoints 6-12 single-mutant candidates to improve transglycosylation yields. Requiring very little prior knowledge of the target enzyme other than its sequence, the method is generic and procures catalysts for the formation of glycosidic bonds involving various d/l-, α/β-pyranosides or furanosides, and exo or endo action. Moreover, mutations validated in one enzyme can be transposed to others, even distantly related enzymes.
Original languageEnglish
JournalChemistry: A European Journal
Issue number40
Pages (from-to)10323-10334
Number of pages13
Publication statusPublished - 2021


  • Enzyme catalysis
  • Hydrolases
  • Multiple sequences alignment
  • Ogliosaccharide synthesis
  • Protein engineering
  • Transglycosylation


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