UC Irvine

Nature performs challenging chemical transformations with the help of metalloenzymes. The efficiency and selectivity of such reactions has been attributed to the control of the covalent as well as non-covalent interactions around the metal center. In synthetic systems, it has been difficult to emulate the long range non-covalent interactions that are found in native enzymes. To incorporate such non-covalent interactions into the modeling of the active sites of metalloenzymes, biotin-streptavidin (Sav) technology has been utilized to immobilize metal complexes within the protein scaffold. This dissertation describes my efforts in engineering artificial metalloproteins (ArMs) using biotin-Sav technology to install non-heme iron active sites that are capable of binding to small molecules. Chapter 2 describes the structural characterization of an FeII ArM that binds to nitric oxide to form Fe-nitrosyl species that has been characterized by electronic absorption, electron paramagnetic resonance, and Mössbauer spectroscopy as well as by X-ray crystallography. Chapter 3 describes the challenges that lability of Fe ions posed in our attempts to study dioxygen activation in a FeII ArM. A previously reported structure of the FeII ArM was re-refined with a new model that fits the electron density better. Addition of the substrate, phenylglyoxylic acid (PGA), to the FeII ArM led to leaching of the Fe ions from the ArM. The leaching of the Fe ions from the ArM was monitored spectroscopically by electronic absorption and Mössbauer spectroscopy. Chapter 4 describes the design and characterization of a μ-oxo-di-FeIII center within Sav. The μ-oxo-di-FeIII center was characterized spectroscopically as well as structurally by X-ray crystallography. The FeIIIFeIII center is capable of binding to external ligands like acetate and azide ions. Oxidation and reduction of the μ-oxo-di-FeIII center has also been studied by solution spectroscopy.