The use of enzyme-catalyst interfaces is underexplored in the field of biocatalysis, particularly in studies on enabling novel reactivity of enzymes. For this thesis, the HaloTag® protein tagging platform was proposed as a bioconjugation method for a pinacol coupling reaction using lipases, as a model for novel reactivities proceeding via ketyl radical intermediates and hydrogen-bonding-facilitated redox attenuation. After an initial lipase screening of 9 lipases, one lipase (Candida rugosa) was found to perform the pinacol coupling of p-anisaldehyde under standard conditions (fluorescein and 530nm light, 3% yield). Based on a retrosynthetic analysis for the photocatalyst-incorporated HaloTag® linker, the intermediates haloamine 1 and aldehyde 6 were synthesized. Further experiments are underway or planned to complete linker synthesis and conduct pinacol coupling experiments with a bioconjugated system. This project underscores the promising biocatalytic promiscuity of lipases for performing reactions proceeding through ketyl radical intermediates, as well as the underdeveloped potential of incorporating bioengineering principles like bioconjugation into biocatalysis to overcome kinetic barriers to electron transfer and optimize biocatalytic reactions.

Despite comprising a variety of bioactive compounds that can be utilized as effective synthetic precursors, the construction of halogenated arenes often relies on hazardous reagents and conditions that pose regioselectivity issues in complex systems. Halodecarboxylation using vanadium-dependent haloperoxidases (VHPOs) has emerged as a sustainable alternative for the synthesis of halogenated arenes. In the Biegasiewicz group, we recently discovered that VHPOs can furnish 3-bromooxindoles from 3-carboxyindoles through a decarboxylation event, followed by oxidation. While this tandem process was exciting, the intermediates of this process, 3- bromoindoles are independently valuable reagents, which necessitated further investigation. Herein we examine the biocatalytic access to bromoindoles for which we addressed the major challenge of undesired oxidation event. The first preventative approach acylated the indole nitrogen, resulting in 1-acetylindole-3-CO2H. This could then be subjected to optimized enzymatic bromination conditions to produce 1-acetyl-3-bromoindole in 98% yield with CiVCPO. The second preventative approach was to modify the reaction conditions, furnishing 1-methyl-3-bromoindole in 73% yield from 1-methylindole-3- CO2H with AmVBPO.

The development of novel aqueous cross-coupling strategies has emerged as a rapidly expanding area of research within organic synthesis. However, many of these cross-coupling reactions require the pre-formation of an organohalide substrate, which often involves toxic halogenating reagents and harsh reaction conditions. This work details the development of a tandem halogenation/cross-coupling procedure in which an electron-rich arene or heteroarene is brominated through an enzymatic halogenation reaction catalyzed by a vanadium dependent haloperoxidase (VHPO) and then used without workup in a subsequent aqueous Suzuki cross-coupling reaction. This sequential process allows the arylated product to be accessed in a single pot from the unfunctionalized substrate via the brominated intermediate. Optimization of the enzymatic halogenation step was performed for three different substrates, resulting in the discovery of conditions for the bromination of 2,3-dihydrobenzofuran, chromane, and anisole in high yield (>95%). The scope of the reaction was then investigated for a range of electron-rich arene and heteroarene substrates. Next, Suzuki cross-coupling conditions were developed in a reaction mixture of pH 5 citrate buffer and acetonitrile and applied to the arylation of 2,3-dihydrobenzofuran utilizing an array of arylboronic acid coupling partners. Finally, the two procedures were combined to perform a tandem enzymatic halogenation/aqueous Suzuki cross-coupling of 2,3-dihydrobenzofuran to give the arylated product in 74% yield.