Non-Native Chemistry of Metalloenzymes

Citation

Goldberg, Nathaniel Wood (2022) Non-Native Chemistry of Metalloenzymes. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/6sec-zx89. https://resolver.caltech.edu/CaltechTHESIS:02232022-224045858

Abstract

Metalloenzymes are important catalysts in biochemistry, but the scope of their naturally occurring activities is dwarfed by the range of chemistry achieved by synthetic transition-metal catalysts. To date, efforts to expand the catalytic repertoire of metalloproteins beyond their native activities have focused almost exclusively on heme-binding proteins, which have been engineered to catalyze a wide variety of carbene- and nitrene-transfer chemistry. Heme-binding proteins represent only a limited subset of the vast diversity of metalloproteins that exists in Nature, and the non-native chemistry of the rest of the metalloproteome remains largely unexplored. This thesis details the discovery and engineering of non-native catalytic abilities of non-heme metalloproteins. Chapter 1 introduces metalloproteins as biocatalysts in synthetic chemistry, and various approaches to expand their catalytic activities. Chapter 2 describes efforts towards enzyme-catalyzed hydrosilylation, including the curation and development of a diverse library of non-heme metalloproteins. In Chapter 3, a non-heme iron-dependent dioxygenase (Pseudomonas savastanoi ethylene-forming enzyme, PsEFE) is found to catalyze nitrene-transfer chemistry, and is engineered by directed evolution to improve this non-native activity. The nitrene transfer activity and selectivity of PsEFE can be modulated by small-molecule metal-coordinating ligands. Chapter 4 describes the discovery and development of a PsEFE-catalyzed olefin aminoarylation reaction, a previously unknown reaction of sulfonyl azides and olefins. This reaction is unprecedented in the existing chemical literature, and displays a number of unusual mechanistic features. Together, the work described here represents the expansion of non-native chemistry to a new class of metalloenzymes, enabling the discovery of previously unknown catalytic activities.

Thesis Files