Cooperative Photometallobiocatalysis: Nonheme Fe Enzyme-Catalyzed Enantioconvergent Radical Decarboxylative Azidation, Thiocyanation, and Isocyanation of Redox-Active Esters

Abstract

Cooperative catalysis with an enzyme and a small-molecule photocatalyst has recently emerged as a potentially general activation mode to advance novel biocatalytic reactions with synthetic utility. Herein, we report cooperative photobiocatalysis involving an engineered nonheme Fe enzyme and a tailored photoredox catalyst to achieve enantioconvergent decarboxylative azidation, thiocyanation, and isocyanation of redox-active esters via a radical mechanism. We repurposed and further evolved metapyrocatechase (MPC), a nonheme Fe extradiol dioxygenase not previously studied in new-to-nature biocatalysis, for the enantioselective C─N₃, C─SCN, and C─NCO bond formation via an enzymatic Fe─X intermediate (X═N₃, NCS, and NCO). A range of primary, secondary, and tertiary alkyl radical precursors were effectively converted by our engineered MPC, allowing the syntheses of organic azides, thiocyanates, and isocyanates with good to excellent enantiocontrol. Further derivatization of these products furnished valuable compounds including enantioenriched amines, triazoles, ureas, and SCF₃-containing products. DFT and MD simulations shed light on the mechanism as well as the binding poses of the alkyl radical intermediate in the enzyme active site and the π-facial selectivity in the enantiodetermining radical rebound. Overall, cooperative photometallobiocatalysis with nonheme Fe enzymes provides a means to develop challenging asymmetric radical transformations eluding small-molecule catalysis.

Keywords: Enzymes; Iron; Photometallobiocatalysis; Reaction mechanisms; Synthetic methods.

Scheme 1

Nonheme Fe enzyme‐catalyzed enantioconvergent decarboxylative radical azidation, thiocyanation, and isocyanation of redox‐active esters enabled by cooperative photometallobiocatalysis. PyMol illustration of metapyrocatechase (MPC), a nonheme Fe extradiol dioxygenase, is made from PDB ID: 1MPY. FL: fluorescein, a photocatalyst. The red sphere is an aryl substituent.

Figure 1

PpMPC's substrate tunnel identified by CAVER^{[ 94 ]} and hydrophobic residues flanking the tunnel entrance. PyMol illustration of MPC is made from PDB ID: 1MPY. Beneficial mutation sites are colored in marine. Other hydrophobic residues are colored in wheat.

Scheme 2

Photometallobiocatalytic enantioconvergent decarboxylative isocyanation/unsymmetrical urea formation. Reaction conditions: 1a (6.67 mM), 6 (66.7 mM, 10.0 equiv), aniline (13.3 mM, 2.0 equiv), 2.0 mol% PpMPC I291L L155F I204L (133.4 mM), 3 mol% fluorescein sodium salt (0.20 mM), 3 mol% (NH₄)₂Fe(II)(SO₄)₂ (0.20 mM), hν (525 nm), 10 mM HEPES buffer, pH = 7.4, rt, and 12 h.

Scheme 3

Derivatization of enantioenriched secondary alkyl azides and thiocyanates.

Figure 2

DFT‐computed reaction energy profile of the azide radical rebound with a model Fe(III)–N₃ complex 14. Im = imidazole.

Figure 3

Representative structures from 500 ns MD simulations of the near‐attack conformations (NAC) that expose the (Re)‐face of the benzylic radical 15 (shown in purple) to the Fe(III)─N₃ intermediate. The forming C─N bond distance with the (A) internal and (B) terminal azide N atoms was restrained. Key active site residues around the azide and the alkyl radical are shown in pink. See Figures S19 and S20 for radical binding poses at longer forming C─N bond distances.