The neutron scattering length density map indicates that both Molecule A and Molecule B contain the amino terminal in the -ND2 protonation state. Fo-Fc omit difference maps calculated confirm the -ND2 state in both Molecule A and Molecule B (Fig. ). The role of possible deprotonation of the N-terminal histidine amino group came to the forefront when it was proposed to promote reactive intermediate formation and stabilization based on findings from a small molecule copper complex. Structural and spectroscopic studies of a substrate-bound LsLPMO9A suggested that the two protons of the N-terminus are exposed to different chemical environments, with one being involved in a hydrogen bond network, which potentially promotes N-terminal deprotonation during LPMO catalysis. Our neutron protein crystallography structures show that the N-terminal amine is not involved in protonation upon initial Cu 2+ reduction and molecular dioxygen activation. However, a mechanistic role for the N-terminal amine in other contexts cannot be conclusively ruled out by our experiments.