An Artificial Metalloenzyme Based on a Copper Heteroscorpionate Enables sp³ C–H Functionalization via Intramolecular Carbene Insertion

Abstract

The selective functionalization of sp 3 C–H bonds is a versatile tool for the diversification of organic compounds. Combining attractive features of homogeneous and enzymatic catalysts, artificial metalloenzymes offer an ideal means to selectively modify these inert motifs. Herein, we report on a copper(I) heteroscorpionate complex embedded within streptavidin that catalyzes the intramolecular insertion of a carbene into sp 3 C–H bonds. Target residues for genetic optimization of the artificial metalloenzyme were identified by quantum mechanics/molecular mechanics simulations. Double-saturation mutagenesis yielded detailed insight on the contribution of individual amino acids on the activity and the selectivity of the artificial metalloenzyme. Mutagenesis at a third position afforded a set of artificial metalloenzymes that catalyze the enantio- and regioselective formation of β- and γ-lactams with high turnovers and promising enantioselectivities.

Non-Native Site-Selective Enzyme Catalysis

“…Artificial metalloenzymes (ArMs) 17 represent a particularly interesting source of enzymes for siteselective catalysis since they could be used to impart site selectivity to synthetic catalysts, thus avoiding the blocking and protecting group strategies noted in the introduction. To date, there have been relatively few examples of this capability, 404,405 and these involved the generation of carbene intermediates. The ArM scaffold is primarily involved in binding and orienting substrates relative to these intermediates to impart selectivity, which mimics the mechanisms of P450s, FeDOs, FDHs, and other enzymes that generate intermediates capable of reacting with a range of functional groups and C-H bonds in organic substrates (e.g.…”

“…Sitesaturation mutagenesis of hotspots within the streptavidin scaffold resulted in two variants for selective formation of the 4-and 5-membered lactam (Scheme 60B). 405 Scheme 60. Site-selective alkylation via carbene transfer by artificial metalloenzymes.…”

“…404 B) Lactam formation via intramolecular cyclization by engineered artificial copper-streptavidin metalloenzymes with major products displayed with the corresponding percent of product pool. 405…”

Non-Native Site-Selective Enzyme Catalysis

“…Site saturation mutagenesis of hotspots within the streptavidin scaffold resulted in two variants for selective formation of the 4-and 5-membered lactam (Scheme 60B). 407…”

“…Specifically, a streptavidin variant containing a biotinylated copper­(I) heteroscorpionate complex catalyzed cyclization of aryl amides resulting in the formation of β- and γ-lactams. Site saturation mutagenesis of hotspots within the streptavidin scaffold resulted in two variants for selective formation of the 4- and 5-membered lactam (Scheme B) …”