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Evaluation of fluoropyruvate as nucleophile in reactions catalysed by N-acetyl neuraminic acid lyase variants: scope, limitations and stereoselectivity

DOI: 10.1039/C5OB02037A DOI Help
PMID: 26537532 PMID Help

Authors: Jennifer Stockwell (University of Leeds) , Adam D. Daniels (University of Leeds) , Claire Windle (University of Leeds) , Thomas A. Harman (University of Leeds) , Thomas Woodhall (University of Leeds) , Tomas Lebl , Chi Hung Trinh (Institute of Molecular and Cellular Biology, University of Leeds) , Keith Mulholland (AstraZeneca) , Arwen Pearson (University of Leeds) , Alan Berry (University of Leeds) , Adam Nelson (University of Leeds)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Organic & Biomolecular Chemistry

State: Published (Approved)
Published: December 2015
Diamond Proposal Number(s): 8367

Abstract: The catalysis of reactions involving fluoropyruvate as donor by N-acetyl neuraminic acid lyase (NAL) variants was investigated. Under kinetic control, the wild-type enzyme catalysed the reaction between fluoropyruvate and N-acetyl mannosamine to give a 90 : 10 ratio of the (3R,4R)- and (3S,4R)-configured products; after extended reaction times, equilibration occurred to give a 30 : 70 mixture of these products. The efficiency and stereoselectivity of reactions of a range of substrates catalysed by the E192N, E192N/T167V/S208V and E192N/T167G NAL variants were also studied. Using fluoropyruvate and (2R,3S)- or (2S,3R)-2,3-dihydroxy-4-oxo-N,N-dipropylbutanamide as substrates, it was possible to obtain three of the four possible diastereomeric products; for each product, the ratio of anomeric and pyranose/furanose forms was determined. The crystal structure of S. aureus NAL in complex with fluoropyruvate was determined, assisting rationalisation of the stereochemical outcome of C–C bond formation.

Subject Areas: Biology and Bio-materials

Instruments: I04-Macromolecular Crystallography