The function of metalloproteins can be directly linked to the local environment around the metallo-cofactor. This environment includes direct covalent interactions of amino acid residues and the metal ion, as well as non-covalent interactions proximal to the metal center. These interactions include hydrogen bonds (H-bonds) from amino acids as well as extended H-bonding networks that include structural water molecules. Synthetic systems have been designed that include non-covalent interactions within their secondary coordination sphere, however it is often difficult to control and predict these interactions. We are employing an approach to model the active sites of metalloproteins that utilizes bio-conjugates of metal complexes that are immobilized within a protein host. Using biotin-streptavidin (Sav) technology, artificial metalloproteins have been prepared that leverage the attributes of protein chemistry with design versatility of synthetic chemistry. Using this approach, we have demonstrated control over the primary coordination sphere by developing model artificial proteins for Type I Cu centers found in cupredoxins. We have further shown that the secondary coordination sphere can be controlled by altering the H-bonding interactions to ligands bound to the Cu center. This control was demonstrated for the stabilization of Cu(II)–OOH species in which the intramolecular H-bonds are derived from Sav. Cu(II)–OOH species are known to be highly unstable at room temperature but confining it within WT Sav rendered it stable with a half-life &gt; 1 day. We have also generated this Cu(II)–OOH species <i>in crystallo </i>and the structure was solved to 1.62 Å resolution to reveal an O–O ligand coordinated to the Cu center. In addition, the Cu(II)–OOH unit is involved in an H-bonding network: the proximal O-atom is H-bonded to a structural water molecule and the distal O-atom is H-bonded to the N49 residue. To our knowledge, this is the only example of a structurally characterized Cu-(hydro)peroxido species that has an H-bond to the distal oxygen. Engineered changes to the H-bonding network lead to control of stability and reactivity of the Cu(II)–OOH species. This presentation will highlight this work and describe the underlying design principles used to prepare these unusual artificial metalloproteins. <br/>

Building copper active sites in artificial metalloproteins

255th National Meeting (2018)

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