Biocatalytic Knoevenagel reaction using alkaline protease from Bacillus licheniformis

BLAP (alkaline protease from Bacillus licheniformis) was used as a biocatalyst in the Knoevenagel condensations of aromatic, hetero-aromatic and alpha;beta-unsaturated aldehydes with less reactive acetylacetone or ethyl acetoacetate. The reactions were performed in organic solvent in the presence of water. The functionalized trisubstituted alkenes and alpha,beta,gamma,delta-unsaturated carbonyl compounds could be obtained in moderate to good yields with E/Z selectivities up to > 99:1. This biocatalytic reaction provided an alternative pathway for Knoevenagel condensation, which also demonstrates a novel case of unnatural activity of existing enzymes.