Structural Studies on the Multidrug Transporter P-glycoprotein Suggest a Mechanism for Polyspecificity

Authors

• MM Gottesman
• L Esser
• S Shukla
• F Zhou
• G Frank
• S Ambudkar
• S Subramaniam
• D Xia

Abstract

The human ATP-dependent multidrug transporter (MDR1, ABCB1), P-glycoprotein (P-gp), was identified and cloned in the Laboratory of Cell Biology in 1986. P-gp has been shown to contribute to multidrug resistance in cancer and to be a key element in pharmacokinetics and organ distribution of many drugs in current use. Intensive efforts in many laboratories have attempted to answer two major questions related to structure and function of P-gp: (1) How does the energy of ATP drive drug transport?; and (2) How does this transporter recognize so many different drugs? Using single particle cryo-EM analysis of human P-gp it is possible to observe structures of P-gp at high enough resolution (up to 12A) to determine that in the apo form, P-gp exists in both open (ATP binding sites far apart) and closed (ATP sites together allowing ATP binding) conformations in dynamic equilibrium. Binding and hydrolysis of ATP (determined in the presence of the ATPase inhibitor vanadate) stabilizes the closed conformation, whereas ADP binding favors the open conformation. X-ray crystallographic analysis of mouse P-gp to 3A resolution provides molecular detail of P-gp with deletions of the linker that connects the two halves of P-gp. As the distance between two ATP binding sites changes, the pitch of helices that bind drugs within transmembrane domains changes, and different residues are exposed for drug binding. Combined with the cryo-EM data, these results argue that the polyspecificity of P-gp results from dynamic twisting of transmembrane helices as the two ATP sites move closer and further apart.

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