Abstract
P-glycoprotein (P-gp) is an ATP-binding cassette transporter that confers multidrug resistance in cancer cells1,2. It also affects the absorption, distribution and clearance of cancer-unrelated drugs and xenobiotics. For these reasons, the structure and function of P-gp have been studied extensively for decades3. Here we present biochemical characterization of P-gp from Caenorhabditis elegans and its crystal structure at a resolution of 3.4 ångströms. We find that the apparent affinities of P-gp for anticancer drugs actinomycin D and paclitaxel are approximately 4,000 and 100 times higher, respectively, in the membrane bilayer than in detergent. This affinity enhancement highlights the importance of membrane partitioning when a drug accesses the transporter in the membrane4. Furthermore, the transporter in the crystal structure opens its drug pathway at the level of the membrane’s inner leaflet. In the helices flanking the opening to the membrane, we observe extended loops that may mediate drug binding, function as hinges to gate the pathway or both. We also find that the interface between the transmembrane and nucleotide-binding domains, which couples ATP hydrolysis to transport, contains a ball-and-socket joint and salt bridges similar to the ATP-binding cassette importers5, suggesting that ATP-binding cassette exporters and importers may use similar mechanisms to achieve alternating access for transport. Finally, a model of human P-gp derived from the structure of C. elegans P-gp not only is compatible with decades of biochemical analysis6,7,8,9,10,11,12, but also helps to explain perplexing functional data regarding the Phe335Ala mutant13,14. These results increase our understanding of the structure and function of this important molecule.
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Acknowledgements
We thank the beamline staff of the GM/CA CAT at the Advanced Photon System for assistance with data collection, Y.-K. Cho for assistance with protein purification and A. Davidson for comments on the manuscript. We also thank the MacKinnon laboratory for reagents and advice on the P. pastoris expression system. This work was supported by Howard Hughes Medical Institute (J.C.), Purdue Center for Cancer Research (NCI CCSG CA23168), and postdoctoral fellowships from the National Research Foundation of Korea and the International Human Frontier Science Program (M.S.J.).
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All authors helped design the study and analysed the data. M.S.J. and M.L.O. determined the crystal structure. M.S.J. and Q.Z. performed the biochemical experiments. M.S.J., M.L.O. and J.C. wrote the manuscript.
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Jin, M., Oldham, M., Zhang, Q. et al. Crystal structure of the multidrug transporter P-glycoprotein from Caenorhabditis elegans. Nature 490, 566–569 (2012). https://doi.org/10.1038/nature11448
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DOI: https://doi.org/10.1038/nature11448
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