Metalloenzymes catalyze a variety of reactions using a limited number of natural amino acids and metallocofactors. Therefore, the environment beyond the primary coordination sphere must play an important role in both conferring and tuning their phenomenal catalytic properties, enabling active sites with otherwise similar primary coordination environments to perform a diverse array of biological functions. However, since the interactions beyond the primary coordination sphere are numerous and weak, it has been difficult to pinpoint structural features responsible for the tuning of activities of native enzymes. Designing artificial metalloenzymes (ArMs) offers an excellent basis to elucidate the roles of these interactions and to further develop practical biological catalysts. In this review, we highlight how the secondary coordination spheres of ArMs influence metal binding and catalysis, with particular focus on the use of native protein scaffolds as templates for the design of ArMs by either rational design aided by computational modeling, directed evolution, or a combination of both approaches. In describing successes in designing heme, nonheme Fe, and Cu metalloenzymes, heteronuclear metalloenzymes containing heme, and those ArMs containing other metal centers (including those with non-native metal ions and metallocofactors), we have summarized insights gained on how careful controls of the interactions in the secondary coordination sphere, including hydrophobic and hydrogen bonding interactions, allow the generation and tuning of these respective systems to approach, rival, and, in a few cases, exceed those of native enzymes. We have also provided an outlook on the remaining challenges in the field and future directions that will allow for a deeper understanding of the secondary coordination sphere a deeper understanding of the secondary coordintion sphere to be gained, and in turn to guide the design of a broader and more efficient variety of ArMs.

Ap rotocol for palladium-catalyzed dearomative functionalization of simple,nonactivated arenes with Grignard reagents has been established. This one-pot method features avisible-light-mediated [4+ +2] cycloaddition between an arene and an arenophile,and subsequent palladium-catalyzed allylic substitution of the resulting cycloadduct with aG rignard reagent. Av ariety of arenes and Grignard reagents can participate in this process,f orming carboaminated products with exclusive syn-1,4-selectivity.M oreover,t he dearomatized products are amenable to further elaborations,p roviding functionalizeda licyclic motifs and pharmacophores.F or example,n aphthalene was converted into sertraline,o ne of the most prescribed antidepressants,i no nly four operations. Finally,t his process could also be conducted in an enantioselective fashion, as demonstrated with the desymmetrization of naphthalene.

Engineering enzymes with novel reactivity and applying them in metabolic pathways to produce valuable products are quite challenging due to the intrinsic complexity of metabolic networks and the need for high in vivo catalytic efficiency. Triacetic acid lactone (TAL), naturally generated by 2-pyrone synthase (2PS), is a platform molecule that can be produced via microbial fermentation and further converted into valueadded products. However, these conversions require extra synthetic steps under harsh conditions. We herein report a biocatalytic system for direct generation of TAL derivatives under mild conditions with controlled chemoselectivity by rationally engineering the 2PS active site and then rewiring the biocatalytic pathway in the metabolic network of E. coli to produce high-value products, such as kavalactone precursors, with yields up to 17 mg/L culture. Computer modeling indicates sterics and hydrogen-bond interactions play key roles in tuning the selectivity, efficiency and yield.

A protocol for palladium‐catalyzed dearomative functionalization of simple, nonactivated arenes with Grignard reagents has been established. This one‐pot method features a visible‐light‐mediated [4+2] cycloaddition between an arene and an arenophile, and subsequent palladium‐catalyzed allylic substitution of the resulting cycloadduct with a Grignard reagent. A variety of arenes and Grignard reagents can participate in this process, forming carboaminated products with exclusive syn‐1,4‐selectivity. Moreover, the dearomatized products are amenable to further elaborations, providing functionalized alicyclic motifs and pharmacophores. For example, naphthalene was converted into sertraline, one of the most prescribed antidepressants, in only four operations. Finally, this process could also be conducted in an enantioselective fashion, as demonstrated with the desymmetrization of naphthalene.