Project Summary Cu (along with Fe) enzymes play dominant roles in O2 activation in Nature. They are important in melanogenesis, neurochemistry, metal ion metabolism, proton pumping, bioremediation, ROS generation, a range of disease states, and can be utilized in biofuel cells for implantable devices. On a molecular level these can reduce O2 by 1,2, possibly 3 and 4 electrons to accomplish a range of functions, including H-atom abstraction from both weak and strong C-H bonds, electrophilic activation by 2 as well as 1 Cu center, and the coupling of O2 reduction to water to Fe metabolism by the multicopper oxidases and proton pumping by the heme-copper oxidases. Oxygen- activating Cu enzymes are divided into five classes based on structure and type of reaction with the mononuclear Cu class further divided based on the redox active states of the Cu and the oxygen. Our studies use the combination of kinetics on the enzymes to obtain intermediates, application of a wide range of spectroscopies to define these, parallel model studies and the coupling of the results of these kinetic, spectroscopic and model studies to electronic structure calculations to define frontier molecular orbitals and reaction coordinates in catalysis. Innovative aspects of this research effort include the development of new spectroscopic methods, the analyses of the unique spectral features often exhibited by Cu/O2 intermediates in terms of their geometric and electronic structures and the coupling of these data to electronic structure calculations for development of experimentally validated reaction coordinates. These studies define the reaction mechanisms of each class and subclass on a molecular level (important in the design of drugs, devices and catalysts) and by parallel studies over the classes develop general insight into structure/function correlations over oxygen utilization in Nature.