To shed further light on the PFK amyloid fibrils' structural features and putative catalytic mechanism, we carried out computational and cryogenic electron microscopy (cryo-EM) analyses (Figure ). Specifically, free molecular dynamics (MD) simulations were performed, illuminating the docking and interactions between PFK and nitrocefin (Figure ), constituting the initial step of the β-lactam hydrolysis reaction. The calculations utilized as a starting point a naive model of PFK dodecamer, predicted in AlphaFold3 , with the nitrocefin molecule placed 1.4 nm above the PFK surface. This configuration ensured no biasing of nitrocefin and PFK to a specific initial structure, allowing the system to better scan bound nitrocefin conformations.