Mechanistic studies of two iron-containing enzymes that catalyze unusual chemical transformations

Enzymes are biological catalysts which trigger chemically inert reaction or accelerate the rates of chemical reactions, oftentimes by many orders of magnitude compared to uncatalyzed reactions. The remarkable catalytic ability afforded by enzymes derives not only from the structure and chemical properties of the enzyme active sites, but sometimes involves redox active metal ions, which allow enzymes to selectively bind to their substrates and to stabilize high energy chemical species along the reaction coordiante. To enhance their catalytic ability, many enzymes have also evolved to require metal ions for activity. Metal ions adapted by enzymes often provide crucial chemical functionality and/or reactivity that are not accessible by the twenty canonical amino acids. Metal ion-containing enzymes serve to greatly broaden the diversity of chemical reactions that can be mediated by enzymes.

The work described herein focuses on mechanistic studies of two enzymes that use iron to catalyze two distinctive reactions. In the first part of this work, studies will focus on the (S)-2-hydroxylpropylphosphonate epoxidase (HppE), a mono-nuclear non-heme iron containing enzyme that is an essential enzyme in fosfomycin biosynthesis, and employs an unidentified reduction system for catalysis. In biological systems, mono-nuclear non-heme iron containing enzymes mediate C-H bond activation and further diverse to various outcomes. The HppE catalyzed reaction discovered herein involves oxidative carbon-phosphorous bond migration, raising questions as chemical mechanism(s) can account for such an unusual transformation. The chemical mechanism of HppE will be interrogated with a combination of organic synthesis and biochemical techniques. Our current results suggest that the HppE may first employ a novel mode of non-heme iron containing enzymes catalysis involving hydrogen atom removal and followed by a carbocation-triggered C-P bond migration. In the second part of this dissertation, the focus is to elucidate the mechanism of 1-hydroxy-2-methyl-2-(E)-butenyl-4-diphosphate reductase (IspH), an enzyme that plays a role in regulating the production of the isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). IspH is a [4S-4Fe] enzyme that catalyzes a reductive deoxygenation reaction requiring the addition of two electrons during turnover. Although extensive efforts have been devoted to the study of this transformation, the mechanism of this intriguing reaction remains elusive. Our current data provide experimental evidence indicating the role(s) of iron-sulfur cluster and has mechanistic implication for this unusual reaction. Taken together, our studies of HppE and IspH help to illustrate the catalytic diversity of non-heme iron containing enzymes, and provide mechanistic insights into these atypical reactions.