Menthol Binds to Extracellular and Transmembrane Domains of the Human A4β2 Nicotinic Receptor

Abstract

Menthol, a small organic compound, has long been used for its flavor as well as its local anesthetic and counterirritant properties. In the US, the 2009 Family Smoking Prevention and Tobacco Control Act by the Congress banned all additives to cigarettes with the exception of menthol. However, an abundance of scientific and clinical research characterize menthol as a non-inert molecule in the body. Smoking menthol cigarettes has been correlated with lower quitting rates and higher relapse rates and it has been shown that all cigarettes, even if labeled as nonmenthol, contain menthol. Unfortunately, at the molecular level not much is known about menthol's interaction with its sites of action, most prominently nicotinic acetylcholine receptors (nAChRs) in the brain. The published X-ray crystal structure of the human α;4β;2 nicotinic receptor - the most abundant nAChR in the human brain - provided an unprecedented opportunity for us to perform molecular dynamics simulations to investigate the interaction between menthol and a heteropentameric nAChR. In this study, we first used free energy perturbation calculations as well as flooding simulations to calculate menthol's water-membrane partition coefficient. The results from the flooding simulations were used to determine the penetration depth as well as the preferred orientation of menthol molecules in a POPC lipid bilayer. Umbrella sampling simulations were performed to derive the free energy profile of menthol across a POPC lipid bilayer. After concluding that menthol molecules partition in mammalian membranes in large numbers we performed flooding simulations in the presence of the human α;4β;2 nicotinic receptor. We demonstrate that menthol molecules bind to both the extracellular and transmembrane domains of the protein with larger numbers of menthol molecules binding to the transmembrane domain of the protein after partitioning into the membrane.