Research Summary The goal of the proposed research is to develop fundamental aspects of copper coordination chemistry relevant to its essential role in the biological processing of O2 and nitrogen oxides (NOx). Research subprojects are: (1) Design & implementation of ligands which H-bond to proximal and/or distal O atoms in LCuII(O2·–) cupric-superoxide complexes will be carried out; initial studies show such elements add to complex stability yet enhance reactivity. Structural, spectroscopic and substrate reactivity consequences will be explored. The recent realization of the importance of acid promoted reactivity leads to new parallel investigations using Bronsted/Lewis acids and substrates with a range of O–H and C–H bond strengths. (2) New research will emphasize the thermodynamics involved in Cu/O2 chemistry. For a (O2·–) superoxide bound to a Cu2 center, the reduction potential (E°) and Marcus λi parameter were determined. Analogs will be studied in order to learn how E° depends on Cu-ligation. Complementary pKa determination for Cun-–OOH hydroperoxo complexes, will provide CunOO–H bond dissociation energies; these parameters correlate to the oxidizing power of Cun-(O2·–) precursors, representing critical new fundamental information. (3) New means for generating & characterizing high-valent Cu-complexes will include study of (a) H+ induced CuII-OOR (R = H, acyl) reductive cleavage studies, a first in Cu (bio)chemistry. We will deduce the role of R, acid pKa and reaction mechanism (i.e., homolysis vs heterolysis). DFT calculations will also be performed and cryo-spectroelectrochemistry will be employed to characterize high-valent dicopper complexes. (b) A new approach will be used to probe the formation and properties of the elusive CuII-O· species. Dimethylaniline (DMA) N-oxides will be used to O-atom transfer to ligand-Cu(I) complexes, where the DMA also serves as a substrate for mechanistic studies. (c) New ligands mimicking the `histidine-brace' motif observed in certain Cu monooxygenases will be employed for study of their CuI/CuII chemistry and O2 or H2O2 reactivity. (4) Copper complexes may participate in formation of deleterious peroxynitrite (PN) through Cu/O2/·NO chemistry. Application of resonance Raman spectroscopic characterization and identification of new intermediates will be carried out. New ligands, including one with an H-bonding moiety, will be added to the studies. The reactivity of Cu-PN complexes will be tested with biologically relevant substrates, CO2, phenols, and indoles; CO2 activates oxidative or nitrative reactivity in biological milieu. Overall, the proposed studies contribute to a broader understanding of copper biochemistry, other metalloprotein (e.g., heme or non-heme iron, Mn, Ni) activation/reduction of O2 and NOx in biology, and associated disease states. Potential long-term applications of this basic research include development of enzyme inhibitors and relevant disease therapeutic strategies.