This study investigates the mechanism of O-O bond cleavage in heme-copper oxidase (HCO) enzymes, combining experimental and computational insights from enzyme intermediates and synthetic models. It is determined that HCOs undergo a proton-initiated O-O cleavage where a single water molecule in the active site enables proton transfer (PT) from the cross-linked tyrosine to a peroxo ligand bridging the heme Fe-III and Cu-II, and multiple H-bonding interactions lower the tyrosine pK(a). Due to sterics within the active site, the proton must either transfer initially to the O(Fe) (a high-energy intermediate), or from another residue over a similar to 10 angstrom distance to reach the O(Cu) atom directly. While the distance between the H+ donor (Tyr) and acceptor (O(Cu)) results in a barrier to PT, this separation is critical for the low barrier to O-O cleavage as it enhances backbonding from Fe into the O-2(2-) sigma* orbital. Thus, PT from Tyr precedes O-O elongation and is rate-limiting, consistent with available kinetic data. The electron transfers from tyrosinate after the barrier via a superexchange pathway provided by the cross-link, generating intermediate P-M. PM is evaluated using available experimental data. The geometric structure contains an Fe-IV=O that is H-bonded to the Cu-II-OH. The electronic structure is a singlet, where the Fe-IV and Cu-II are antiferromagnetically coupled through the H-bond between the oxo(Fe) and hydroxo(Cu) ligands, while the Cu-II and Tyr(center dot) are ferromagnetically coupled due their delocalization into orthogonal magnetic orbitals on the cross-linked His residue. These findings provide critical insights into the mechanism of efficient O-2 reduction in HCOs, and the nature of the P-M intermediate that couples this reaction to proton pumping.