Engineering the Donor Selectivity of D‐Fructose‐6‐Phosphate Aldolase for Biocatalytic Asymmetric Cross‐Aldol Additions of Glycolaldehyde |
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| Authors: | Dr. Anna Szekrenyi Anna Soler Dr. Xavier Garrabou Dr. Christine Guérard‐Hélaine Dr. Teodor Parella Dr. Jesús Joglar Prof. Dr. Marielle Lemaire Dr. Jordi Bujons Prof. Dr. Pere Clapés |
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| Affiliation: | 1. Biotransformation and Bioactive Molecules Group, Instituto de Química Avanzada de Catalu?a (IQAC‐CSIC), Jordi Girona 18‐26, 08034 Barcelona (Spain), Fax: (+34)?932045904;2. Present address: Laboratory of Organic Chemistry, HCI F322, ETH H?nggerberg, 8093 Zurich (Switzerland);3. Clermont Université, Université Blaise Pascal, ICCF, BP 10448, 63000 Clermont‐Ferrand (France);4. CNRS, UMR 6296, ICCF, BP 80026, 63171 Aubière (France);5. Dept Química, Servei de Ressonància Magnètica Nuclear, Universitat Autònoma de Barcelona, Bellaterra (Spain) |
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| Abstract: | D ‐Fructose‐6‐phosphate aldolase (FSA) is a unique catalyst for asymmetric cross‐aldol additions of glycolaldehyde. A combination of a structure‐guided approach of saturation mutagenesis, site‐directed mutagenesis, and computational modeling was applied to construct a set of FSA variants that improved the catalytic efficiency towards glycolaldehyde dimerization up to 1800‐fold. A combination of mutations in positions L107, A129, and A165 provided a toolbox of FSA variants that expand the synthetic possibilities towards the preparation of aldose‐like carbohydrate compounds. The new FSA variants were applied as highly efficient catalysts for cross‐aldol additions of glycolaldehyde to N‐carbobenzyloxyaminoaldehydes to furnish between 80–98 % aldol adduct under optimized reaction conditions. Donor competition experiments showed high selectivity for glycolaldehyde relative to dihydroxyacetone or hydroxyacetone. These results demonstrate the exceptional malleability of the active site in FSA, which can be remodeled to accept a wide spectrum of donor and acceptor substrates with high efficiency and selectivity. |
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| Keywords: | aldol reaction carbohydrates enzyme catalysis enzyme models protein engineering |
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