Enantioselective Single and Dual a-C–H Bond Functionalization of Cyclic Amines via Enzymatic Carbene Transfer

17 October 2022, Version 1
This content is a preprint and has not undergone peer review at the time of posting.


Cyclic amines are ubiquitous structural motifs found in pharmaceuticals and biologically active natural products, making methods for their elaboration via direct C–H functionalization of considerable synthetic value. Herein, we report the development of an iron-based biocatalytic strategy for enantioselective a-C–H functionalization of pyrrolidines via a carbene transfer reaction with diazoacetone. Currently unreported for organometallic catalysts, this transformation can be accomplished in high yields, high catalytic activity and high stereoselectivity (up to 99:1 e.r. and 20,350 TON) using engineered variants of cytochrome P450 CYP119 from Sulfolobus solfataricus. This methodology was further extended to enable enantioselective a-C–H functionalization in the presence of ethyl diazoacetate as carbene donor (up to 89:11 e.r. and 8,920 TON), and the two strategies were combined to achieve a one-pot as well as a tandem dual C–H functionalization of the cyclic amine substrate with enzyme-controlled diastereo- and enantiodivergent selectivity. This biocatalytic approach is amenable to gram-scale synthesis and can be applied to drug scaffolds for late-stage C–H functionalization. This work provides an efficient and tunable method for direct asymmetric a-C–H functionalization of saturated N-heterocycles which should offer new opportunities for the synthesis, discovery, and optimization of bioactive molecules.


C-H functionalization
Carbene Transferase
Directed evolution
Enantioselective and stereodivergent synthesis
Late-stage functionalization

Supplementary materials

Supplementary Information
Supplementary Information for the manuscript


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