Biocatalytic Asymmetric Michael Additions of Nitromethane to α,β-Unsaturated Aldehydes via Enzyme-bound Iminium Ion Intermediates

Abstract

The enzyme 4-oxalocrotonate tautomerase (4-OT) exploits an N-terminal proline as main catalytic residue to facilitate several promiscuous C–C bond-forming reactions via enzyme-bound enamine intermediates. Here we show that the active site of this enzyme can give rise to further synthetically useful catalytic promiscuity. Specifically, the F50A mutant of 4-OT was found to efficiently promote asymmetric Michael additions of nitromethane to various α,β-unsaturated aldehydes to give γ-nitroaldehydes, important precursors to biologically active γ-aminobutyric acids. High conversions, high enantiocontrol, and good isolated product yields were achieved. The reactions likely proceed via iminium ion intermediates formed between the catalytic Pro-1 residue and the α,β-unsaturated aldehydes. In addition, a cascade of three 4-OT(F50A)-catalyzed reactions followed by an enzymatic oxidation step enables assembly of γ-nitrocarboxylic acids from three simple building blocks in one pot. Our results bridge organo- and biocatalysis, and they emphasize the potential of enzyme promiscuity for the preparation of important chiral synthons.

Enantiocomplementary Epoxidation Reactions Catalyzed by an Engineered Cofactor‐Independent Non‐natural Peroxygenase

“…Communications group is focused on further improving the peroxygenase activity of this promising cofactor-independent enzyme, making use of a recently developed selective colorimetric "turn-on" probe for efficient engineering of iminium biocatalysis. [13] The non-natural peroxygenase activity of 4-OT, together with its previously reported promiscuous CÀC bondforming Michaelase [7,9,14] and aldolase [15] activities, emphasize the chemical versatility of the 4-OT protein scaffold.…”

In Situ Acetaldehyde Synthesis for Carboligation Reactions

“…[e] Determined by GC with chiral stationary phase or derivatized into a cyclic acetal and determined by HPLC with a chiral stationary phase. The absolute configuration was determined by comparison to literature . [f] Isolated yield compared to 6 .…”

“…4‐OT is unique in that it can use 5 as substrate for C−C bond‐forming Michael‐type additions to nitroolefin acceptors and for aldol condensations with benzaldehydes . Previously, we have demonstrated that 4‐OT can be combined with other biocatalysts and chemocatalysts to convert 6 and 8 into GABA analogues using two complementary one‐pot cascade reactions . However, both routes rely on 5 as a substrate.…”

Enabling protein-hosted organocatalytic transformations

“…7d). 115 Reduction of the intermediate iminium ion by sodium cyanoborohydride and subsequent mass spectrometry analysis provide evidence that supports the formation of the iminium intermediate. 73 Mutagenesis via a combined computational and experimental approach has led to the identication of enhanced variants.…”