Engineering aldolases for asymmetric synthesis

Formation of carbon-carbon bonds is central to synthetic chemistry. The aldol reaction provides the chemistry to fuse a nucleophilic enolate with an electrophilic aldehyde to form a new CC bond between two newly formed asymmetric centers. A major challenge in the reaction is steering the stereochemistry of the product. Aldolases are lyases that catalyze aldol reactions as well as the retro-aldol cleavage, and are abundant in cellular metabolism. Due to the often exquisite stereoselectivity in aldolase catalyzed carboligation reactions, these enzymes are gaining increased interest as potentially important tools in asymmetric synthesis of new useful compounds. Fructose 6-phosphate aldolase from Escherichia coli (FSA) is of special interest because of its very unusual independence of phosphorylated reactant substrates. The current text describes the protein engineering of FSA, applying principles of directed evolution, for the generation, production and characterization of new aldolase variants. A range of new enantiopure polyhydroxylated compounds were produced applying isolated FSA variants.