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Substrate binding to a cyclodextrin glycosyltransferase and mutations increasing the gamma-cyclodextrin production
G Parsiegla1, A K Schmidt, G E Schulz
1Institut für Organische Chemie und Biochemie, Freiburg im Breisgau, Germany.
European Journal of Biochemistry
|September 17, 1998
Summary
Researchers engineered bacterial cyclodextrin glycosyltransferases to enhance production of gamma-cyclodextrins. These modified enzymes shift product spectrum, enabling efficient synthesis of valuable cyclodextrins for diverse applications.
Area of Science:
- Biochemistry
- Enzymology
- Carbohydrate Chemistry
Background:
- Bacterial cyclodextrin glycosyltransferases (CGTases) catalyze starch conversion into cyclodextrins.
- Cyclodextrins are cyclic maltooligosaccharides with diverse industrial applications.
- The enzyme from Bacillus circulans produces primarily beta-cyclodextrin.
Purpose of the Study:
- To investigate substrate binding modes of Bacillus circulans CGTase.
- To engineer CGTase variants for altered cyclodextrin production spectra.
- To enhance the yield of gamma-cyclodextrins, which are valuable for encapsulating larger molecules.
Main Methods:
- Performed binding studies with various substrates and analogues using the wild-type enzyme.
- Designed and created two specific mutations in the CGTase based on binding study insights.
- Analyzed the cyclodextrin production spectrum of the mutant enzymes.
Main Results:
- Observed detailed binding modes consistent with homologous enzymes.
- Successfully modified the enzyme's production spectrum through targeted mutations.
- Shifted the predominant product from beta-cyclodextrin towards gamma-cyclodextrin.
Conclusions:
- Enzyme engineering can effectively alter cyclodextrin product distribution.
- Mutant CGTases offer a pathway to increased gamma-cyclodextrin production.
- This research facilitates the synthesis of rare cyclodextrins for advanced applications.