Mycolactone is the sole virulence factor in the neglected tropical skin disease Buruli Ulcer, and an important model system for understanding host-pathogen interactions. Key outstanding questions are how this toxin invades host cells, is long-lived in the host vasculature, evades the secondary immune response, and is difficult to detect with antibodies. Each of these issues could be explained by interactions with lipophilic carriers, including membranes. Using state of the art enhanced sampling methods, we herein calculate permeation free energy profiles for this relatively large molecule, thereby revealing how mycolactone interacts strongly with model lipid membranes. Comparing atomistic to MARTINI CG simulations reveals the importance of atomistic modeling and properly accounting for hydration during permeation. This work sets the stage for understanding how mycolactone is trafficked and distributed in host systems, information that will be crucial in the development of effective diagnostics and adjunctive therapies, as well as understanding disease pathogenesis.