My research focuses on understanding the activation of dioxygen and the mechanism of oxygen binding and reactivity in the noncoupled binuclear copper enzymes dopamine β-monooxygenase (DβM) and peptidylglycine α-hydroxylating monooxygenase (PHM).  DβM converts dopamine to norepinephrine and is important in the regulation of these neurotransmitters.  PHM is part of the bifunctional peptidylglycine α-amidating monooxygenase (PAM) and catalyzes the hydroxylation of the glycine α-carbon of glycine-extended peptides, an important step in the formation of bioactive peptides (Figure 1).  The active site of each enzyme is occupied by two copper atoms separated by a large (11Å for PHM) solvent-filled cleft (Figure 2).  These copper atoms are termed “noncoupled” as they exhibit no observable magnetic interaction with each other. Each of these enzymes activates dioxygen at a single copper center (CuM) to form a CuMII-O2 intermediate that abstracts a hydrogen atom from the substrate.  To complete the reaction, an electron is transferred to CuM from the second, distant copper site (CuH).

Figure 1: H-atom abstraction reactions of DβM and PHM

My studies on DβM and PHM include the spectroscopic investigations of the resting enzymes, intermediate species along the enzymes’ reaction pathways, and relevant model complexes.  Spectroscopic techniques including Magnetic Circular Dichroism (MCD), Electric Paramagnetic Resonance (EPR), Absorption, Resonance Raman, and X-Ray Absorption Spectroscopy (XAS) are coupled to Density Functional Theoretical (DFT) calculations to obtain detailed electronic and structural information relevant to the catalytic mechanisms of these enzymes.

Figure 2: The noncoupled binuclear copper active site of PHM and DβM

In addition to PHM and DβM I also study the mechanism of cofactor biogenesis in the mononuclear copper enzyme galactose oxidase (GO).  GO catalyzes the oxidation of primary alcohols to their corresponding aldehydes and hydrogen peroxide.  Remarkably, in its first turnover after expression, GO employs its copper center as a catalyst for the formation of a necessary redox cofactor, topaquinone (TPQ).  In a process termed cofactor biogenesis, GO generates TPQ intramolecularly through a post-translation modification of a Tyr residue within the enzyme.  MCD, EPR, and rR spectroscopies are used to investigate the cofactor biogenesis mechanism.