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タイトル: | In vivo FRET analyses reveal a role of ATP hydrolysis–associated conformational changes in human P-glycoprotein |
著者: | Futamata, Ryota Ogasawara, Fumihiko Ichikawa, Takafumi https://orcid.org/0000-0002-0490-4176 (unconfirmed) Kodan, Atsushi https://orcid.org/0000-0003-0317-9109 (unconfirmed) Kimura, Yasuhisa Kioka, Noriyuki https://orcid.org/0000-0002-2708-537X (unconfirmed) Ueda, Kazumitsu https://orcid.org/0000-0003-2980-6078 (unconfirmed) |
著者名の別形: | 二股, 良太 小笠原, 史彦 市川, 尚文 小段, 篤史 木村, 泰久 木岡, 紀幸 植田, 和光 |
キーワード: | ABC transporter ATPase multidrug transporter membrane protein fluorescence resonance energy transfer (FRET) conformational change ABCB1 MDR1 P-glycoprotein |
発行日: | 20-Apr-2020 |
出版者: | American Society for Biochemistry and Molecular Biology Elsevier BV |
誌名: | Journal of Biological Chemistry |
巻: | 295 |
号: | 15 |
開始ページ: | 5002 |
終了ページ: | 5011 |
抄録: | P-glycoprotein (P-gp; also known as MDR1 or ABCB1) is an ATP-driven multidrug transporter that extrudes various hydrophobic toxic compounds to the extracellular space. P-gp consists of two transmembrane domains (TMDs) that form the substrate translocation pathway and two nucleotide-binding domains (NBDs) that bind and hydrolyze ATP. At least two P-gp states are required for transport. In the inward-facing (pre-drug transport) conformation, the two NBDs are separated, and the two TMDs are open to the intracellular side; in the outward-facing (post-drug transport) conformation, the NBDs are dimerized, and the TMDs are slightly open to the extracellular side. ATP binding and hydrolysis cause conformational changes between the inward-facing and the outward-facing conformations, and these changes help translocate substrates across the membrane. However, how ATP hydrolysis is coupled to these conformational changes remains unclear. In this study, we used a new FRET sensor that detects conformational changes in P-gp to investigate the role of ATP binding and hydrolysis during the conformational changes of human P-gp in living HEK293 cells. We show that ATP binding causes the conformational change to the outward-facing state and that ATP hydrolysis and subsequent release of γ-phosphate from both NBDs allow the outward-facing state to return to the original inward-facing state. The findings of our study underscore the utility of using FRET analysis in living cells to elucidate the function of membrane proteins such as multidrug transporters. |
著作権等: | © 2020 Futamata et al. This is an Open Access article under the CC BY license. |
URI: | http://hdl.handle.net/2433/281618 |
DOI(出版社版): | 10.1074/jbc.RA119.012042 |
PubMed ID: | 32111736 |
出現コレクション: | 学術雑誌掲載論文等 |
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