Biocatalysis

The direct, catalytic functionalization of C-H bonds presents a long withstanding challenge for synthetic chemists due, in part, to their inert nature and ubiquity in organic compounds, making it difficult to achieve selective transformations. Transition-metal catalysts have been proven time and again to be effective towards the activation of these bonds; however, their use presents a myriad of issues pertaining to the chemo-, regio-, and stereoselectivity in activating a target C-H bond. Therefore, the usage of C-H functionalization strategies is often situationally-limited depending on the synthetic route. For example, directing groups are commonly employed to dictate the position of the target C-H bond towards the metal center via coordination-assisted control. Though highly effective, this method can be limited, requiring tactical design of the substrate. Within the many strategies available to activate C-H bonds, direct functionalization via metal-catalyzed group transfer reactions is an attractive approach in organic synthesis to directly generate C-O, C-N, or C-C bonds.[1] Synthetic metalloporphyrin catalysts have been extensively studied for these reactions due, in large part, from inspiration driven by the native C-H hydroxylation activity of heme-enzymes such as Cytochrome P450s.[2]

Carbene transfer reactions are of particular interest in the quickly developing field of organometallic biocatalysis. These reactions can produce new C-C, C-N, and C-S bonds, and useful organic products such as cyclopropanes, amines and thioethers, among others. A number of transition metal complexes catalyze the decomposition of diazo reagents to form an electrophilic metal-carbene unit that is then attacked by a nucleophilic reagent such as an olefin (for cyclopropanation) or an amine (for N-H insertion). In particular, a variety of metalloporphyrin complexes including metals such as Fe, Co, Ru, Rh, and Ir have been effective towards catalyzing carbene transfer reactions.[3] These complexes catalyze thousands of turnovers, and the second and third row transition metal complexes typically have long catalyst lifetimes, but also exhibit the aforementioned drawbacks of small molecule catalysts where stereoselectivity is difficult to control and reactions are often performed in environmentally detrimental organic solvents. The Arnold and Fasan groups have demonstrated that heme proteins such as Cytochrome P450BM3 and myoglobin (Mb) can be modified in order to produce very active and stereoselective carbene transfer catalysts.[4-6] Multiple rounds of active-site mutagenesis and reactivity screening through directed evolution (invented by the Arnold group) can lead to improved product yield and stereoselectivity, though at the expense of gaining a more fundamental understanding of how these modifications improve catalysis.[1] Rational design of a catalyst through active-site mutations and cofactor modification is a more time-intensive approach, but can also lead to increased activity and selectivity of the system, and a better understanding of how the second coordination sphere influences reactivity.

An alternative strategy to using the naturally occuring heme cofactor in biocatalysis is to incorporate non-natural hemes into heme proteins. Two strategies have evolved here, which are the use of non-biological metals like Ru, Rh, Ir, etc.,[7,8] the use of non-natural heme derivatives (like porphycenes), and a combination of the two. We are interested in both of these approaches, using Mb as the protein matrix of choice.