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Outward-facing P-glycoprotein bound to drug displays nucleotide-dependent drug egress mechanism

Han, S. B.; Prince, S. M.; Warwicker, J.; Fan, H.

2025-12-07 molecular biology
10.64898/2025.12.04.692479 bioRxiv
Show abstract

ABCB1/P-glycoprotein requires coordination between ligand movement in the trans-membrane cavity and ATP binding and hydrolysis at the nucleotide binding domains, yet the sequence of these events remains incompletely resolved. Here, atomistic simulations with paclitaxel and verapamil across outward facing P-glycoprotein ensembles in various nucleotide states were integrated with analysis of transmembrane domain contacts, geometric solvent/ligand pathways, extracellular gating distances, and nucleotide coordi-nation characterization. Collectively, our simulations delineate a mechanistic framework for P-glycoprotein in which ATP binding organizes an outward-facing ensemble and sequential hydrolysis at the two nucleotide-binding sites drives substrate progression toward egress. Asymmetry in the nucleotide state is correlated to distinct motions within the transmembrane domains, which suggests a possible transient extracellular gating mechanism that enables staged release and limits re-entry of the ligand. The resulting model connects nucleotide chemistry to transmembrane domain motions that regulate gate dynamics and may guide experiments and development of modulators for ABC exporter activity in physiological and pharmacological contexts.

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