Nature employs metalloproteins to mediate chemical transformations with rates and selectivities not yet achieved in synthetic systems. The efficiency of metalloproteins has been attributed to the specific control over the local environment surrounding their active sites. In synthetic systems, it is difficult to establish the extensive hydrogen bonding (H-bonding) networks and long-range noncovalent interactions that are essential for function. To overcome these structural limitations, we have designed artificial metalloproteins in which the local environments around synthetic metal complexes can be modulated with a protein host. Within this context, biotin-Sav technology is used as the basis to construct artificial metalloproteins. The high binding affinity of Sav for biotin (Kd ~ 10-15 M) provides a reproducible mechanism for placing artificial biotinylated metallocofactors within the protein host. Additionally, site-directed mutagenesis of the metal complex binding site is used to install amino acid residues that can: 1) act as ligands and coordinate to the metal center or 2) modulate local environments. Using this approach, biotinylated metal complexes have been synthesized and inserted into several variants of Sav to study the effects of varying the local environment around the metal center. This presentation will show results of new artificial metalloproteins in which the structures of immobilized metal sites determined from X-ray diffraction methods are compared to spectroscopic and reactivity studies done in solution.

Design of artificial metalloproteins using Biotin-Streptavidin Technology

255th National Meeting (2018)

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