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Engineering CGTase to improve synthesis of alkyl glycosides

Ara, Kazi Zubaida Gulshan ; Linares-Pastén, Javier A LU orcid ; Jönsson, Jonas ; Viloria-Cols, Maria LU ; Ulvenlund, Stefan LU ; Adlercreutz, Patrick LU orcid and Karlsson, Eva Nordberg LU orcid (2021) In Glycobiology 31(5). p.603-612
Abstract

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... (More)

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.

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author
; ; ; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Acceptor subsites, Cyclodextrin glycosyltransferase, Coupling reaction, dodecyl-β-maltoside, γ-Cyclodextrin
in
Glycobiology
volume
31
issue
5
article number
cwaa109
pages
10 pages
publisher
Oxford University Press
external identifiers
  • scopus:85107902497
  • pmid:33270133
ISSN
1460-2423
DOI
10.1093/glycob/cwaa109
language
English
LU publication?
yes
id
099f666d-4b26-476f-a351-a746f99dc689
date added to LUP
2020-12-06 21:57:58
date last changed
2024-06-13 01:42:32
@article{099f666d-4b26-476f-a351-a746f99dc689,
  abstract     = {{<p>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 <i>Carboxydocella</i> 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.</p>}},
  author       = {{Ara, Kazi Zubaida Gulshan and Linares-Pastén, Javier A and Jönsson, Jonas and Viloria-Cols, Maria and Ulvenlund, Stefan and Adlercreutz, Patrick and Karlsson, Eva Nordberg}},
  issn         = {{1460-2423}},
  keywords     = {{Acceptor subsites; Cyclodextrin glycosyltransferase; Coupling reaction; dodecyl-β-maltoside; γ-Cyclodextrin}},
  language     = {{eng}},
  month        = {{05}},
  number       = {{5}},
  pages        = {{603--612}},
  publisher    = {{Oxford University Press}},
  series       = {{Glycobiology}},
  title        = {{Engineering CGTase to improve synthesis of alkyl glycosides}},
  url          = {{http://dx.doi.org/10.1093/glycob/cwaa109}},
  doi          = {{10.1093/glycob/cwaa109}},
  volume       = {{31}},
  year         = {{2021}},
}