Beyond Iron: Iridium-Containing P450 Enzymes for Selective Cyclopropanations of Structurally Diverse Alkenes

Abstract

Enzymes catalyze organic transformations with exquisite levels of selectivity, including chemoselectivity, stereoselectivity, and substrate selectivity, but the types of reactions catalyzed by enzymes are more limited than those of chemical catalysts. Thus, the convergence of chemical catalysis and biocatalysis can enable enzymatic systems to catalyze abiological reactions with high selectivity. Recently, we disclosed artificial enzymes constructed from the apo form of heme proteins and iridium porphyrins that catalyze the insertion of carbenes into a C–H bond. We postulated that the same type of Ir(Me)-PIX enzymes could catalyze the cyclopropanation of a broad range of alkenes with control of multiple modes of selectivity. Here, we report the evolution of artificial enzymes that are highly active and highly stereoselective for the addition of carbenes to a wide range of alkenes. These enzymes catalyze the cyclopropanation of terminal and internal, activated and unactivated, electron-rich and electron-deficient, conjugated and nonconjugated alkenes. In particular, Ir(Me)-PIX enzymes derived from CYP119 catalyze highly enantio- and diastereoselective cyclopropanations of styrene with ±98% ee, >70:1 dr, >75% yield, and ∼10,000 turnovers (TON), as well as 1,2-disubstituted styrenes with up to 99% ee, 35:1 dr, and 54% yield. Moreover, Ir(Me)-PIX enzymes catalyze cyclopropanation of internal, unactivated alkenes with up to 99% stereoselectivity, 76% yield, and 1300 TON. They also catalyze cyclopropanation of natural products with diastereoselectivities that are complementary to those attained with standard transition metal catalysts. Finally, Ir(Me)-PIX P450 variants react with substrate selectivity that is reminiscent of natural enzymes; they react preferentially with less reactive internal alkenes in the presence of more reactive terminal alkenes. Together, the studies reveal the suitability of Ir-containing P450s to combine the broad reactivity and substrate scope of transition metal catalysts with the exquisite selectivity of enzymes, generating catalysts that enable reactions to occur with levels and modes of activity and selectivity previously unattainable with natural enzymes or transition metal complexes alone.

Exploiting and engineering hemoproteins for abiological carbene and nitrene transfer reactions

“…The CYP119- derived biocatalysts were also found to be active for the intermolecular carbene C–H insertion reaction with phthalan and EDA. Ir-substituted CYP119 variants were also shown to be active catalysts for the cyclopropanation of olefins currently not accepted by Fe-containing enzymes, including various internal, aliphatic and electron-deficient alkenes [34]. These artificial enzymes have the shortcomings of requiring a cofactor containing rare and expensive metal (Ir) and specialized in vitro reconstitution or expression protocols for their assembly.…”

An Artificial Heme Enzyme for Cyclopropanation Reactions

“…These enzymes have proven amenable to optimization by both genetic methods and co‐factor replacement 14, 15, 16, 17, 18, 19, 20. A common feature of these (designed) heme enzymes is that they contain a large hydrophobic substrate binding pocket orthogonal to the plane of the heme moiety.…”

Diverse engineered heme proteins enable stereodivergent cyclopropanation of unactivated alkenes

“…19,20,32 Use of an artificial iridium cofactor (Ir(Me)PIX) required the lysis, purification, and in vitro metalation of the apoprotein with the Ir(Me)PIX, all of which add time and cost to catalyst preparation. Though it may be possible to perform these metalations in vivo, 33 the synthetic, noble-metal catalyst is far more expensive than the native heme cofactor, which is manufactured by the cell and loaded into the catalyst during protein expression in vivo.…”

“…Unactivated, aliphatic alkenes are attractive feedstocks for chemical synthesis, but their transformation to higher-value chiral products is challenging due to their inert nature, high degree of conformational flexibility, and limited steric and electronic bias to guide stereocontrol. 17 State-of-the art methods for unactivated alkene cyclopropanation often rely on noble metals [18][19][20] (Supplemental Table S1); no iron-based catalyst for the enantioselective intermolecular cyclopropanation of unactivated alkenes has been reported. However, directed evolution of heme proteins has previously enabled biocatalysis to access reactions performed with noble-metal catalysts, such as carbonsilicon bond formation 21 and intermolecular CH amination 22 .…”