Sculpting Metal-binding Environments in De Novo Designed Three-helix Bundles

De novo protein design is a biologically relevant approach used to study the active centers of native metalloproteins. In this review, we will first discuss the design process in achieving α₃D, a de novo designed three-helix bundle peptide with a well-defined fold. We will then cover our recent work in functionalizing the α₃D framework by incorporating a tris(cysteine) and tris(histidine) motif. Our first design contains the thiol-rich sites found in metalloregulatory proteins that control the levels of toxic metal ions (Hg, Cd, and Pb). The latter design recapitulates the catalytic site and activity of a natural metalloenzyme carbonic anhydrase. The review will conclude with future design goals aimed at introducing an asymmetric metal-binding site in the α₃D framework.