Engineering CGTase to improve synthesis of alkyl glycosides

Kazi Zubaida Gulshan Ara, Javier A Linares-Pastén, Jonas Jönsson, Maria Viloria-Cols, Stefan Ulvenlund, Patrick Adlercreutz, Eva Nordberg Karlsson

Research output: Contribution to journalArticlepeer-review


Alkyl glycoside surfactants with elongated carbohydrate chains are useful in different applications due to their improved biocompatibility. Cyclodextrin glucanotransferases can catalyse the elongation process through the coupling reaction. However, due to the presence of a hydrophobic tail, the interaction between an alkyl glycoside acceptor and the active site residues is weaker than the interaction with maltooligosaccharides at the corresponding site. Here we report the mutations of F197, G263 and E266 near the acceptor subsites in the CGTase CspCGT13 from Carboxydocella sp. The results showed that substitutions of both F197 and G263 were important for the binding of acceptor substrate dodecyl maltoside during coupling reaction. The double mutant F197Y/G263A showed enhanced coupling activity and displayed a 2-fold increase of the primary coupling product using γ-cyclodextrin as donor when compared to wildtype CspCGT13. Disproportionation activity was also reduced, which was also the case for another double mutant (F197Y/E266A) that however not showed the corresponding increase in coupling. A triple mutant F197Y/G263A/E266A maintained the increase in primary coupling product (1.8-fold increase) using dodecyl maltoside as acceptor, but disproportionation was approximately at the same level as in the double mutants. In addition, hydrolysis of starch was slightly increased by the F197Y and G263A substitutions, indicating that interactions at both positions influenced the selectivity between glycosyl and alkyl moieties.

Original languageEnglish
Article numbercwaa109
Pages (from-to)603-612
Number of pages10
Issue number5
Early online date2020 Dec 2
Publication statusPublished - 2021 May 1

Subject classification (UKÄ)

  • Biocatalysis and Enzyme Technology

Free keywords

  • Acceptor subsites
  • Cyclodextrin glycosyltransferase
  • Coupling reaction
  • dodecyl-β-maltoside
  • γ-Cyclodextrin


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