<i>De Novo</i> Design of a Metalloprotein with a Synthetically Inspired Dinuclear Paddlewheel Coordination Motif

A metal-centered design approach was used to design a tetrameric protein assembly, PW1, with a new-to-nature metal center inspired by the structures of transition metal paddlewheel complexes such as di-Cu II tetraacetate. The structure of PW1 matches the design prediction with atomic accuracy, preorganizing four Glu residues to form the desired dinuclear Cu II center. Notably, Cu-PW1 has a reduction potential of −0.085 V at pH 7.0, which is more negative than any natural or engineered Cu redox protein. In addition to Cu II, PW1 can also scaffold Rh II ions in the dinuclear paddlewheel geometry in crystallo as well as trivalent lanthanide ions in a mononuclear fashion. These results demonstrate the feasibility of designing new protein architectures around synthetically inspired, nonbiological metal centers, thus expanding the scope of artificial metalloproteins beyond those constructed around pre-existing protein scaffolds and opening new avenues to discover structure–function relationships outside the constraints of natural evolution.