Nucleophile Promiscuity of Engineered Class II Pyruvate Aldolase from E. Coli, YfaU

Pyruvate-dependent aldolases exhibit a stringent selectivity for pyruvate, limiting their synthetic potential application, a drawback shared with other existing aldolases. Structure-guided rational protein engineering rendered a 2-keto-3-deoxy-L-rhamnonate aldolase variant, fused with maltose binding protein (MBP-YfaU W23V/L216A), capable to efficiently convert larger pyruvate analogs, e.g. having linear and branched aliphatic chains, in aldol addition reactions. Combination of these nucleophiles with N-Cbz-alaninal and N-Cbz-prolinal electrophiles gave access to chiral building blocks, e.g. derivatives of (2S,3S,4R)-4-amino-3-hydroxy-2-methylpentanoic acid (68%, dr 90:10) and the enantiomer of Dolaproine (33%, dr 94:6) as well as a collection of unprecedented α-amino acid derivatives of the proline and pyrrolizidine type, with conversions varying between 6-93% and diasteromeric ratios from 50:50 to 95:5 depending on the nucleophilic and electrophilic components
This project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement no. 635595 (CarbaZymes), the Ministerio de Economía y Competitividad (MINECO), the Fondo Europeo de Desarrollo Regional (FEDER) (grant no. CTQ2015-63563-R to P.C. and CTQ2015-64436-P to T.P.), and COST action CM1303 Systems Biocatalysis.