De novo proteins provide a unique opportunity for investigating the structure-function relationships of metalloproteins in a minimalistic and easily controlled scaffold. Beginning with the de novo designed four-helix bundle di-iron carboxylate protein DFsc, we successfully created a protein with catalytic two-electron oxidase activity. We then rationally reprogrammed this protein to catalyze the selective N-hydroxylation of arylamines through the introduction of a third iron-binding His ligand in the active site. Through a combination of detailed kinetic and spectroscopic studies, we have obtained molecular level insights into the differential reactivities of these two model proteins. Our results suggest that these proteins all form end-on hydroperoxy intermediates which then perform either oxidase or oxygenase chemistry depending on the ability of substrate to bind directly to one of the active site iron atoms.

Differential oxidase and oxygenase reactivities in de novo Due Ferri proteins

251st National Meeting (2016)

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