論文

査読有り
2020年4月10日

In vivo FRET analyses reveal a role of ATP hydrolysis–associated conformational changes in human P-glycoprotein

Journal of Biological Chemistry
  • Ryota Futamata
  • ,
  • Fumihiko Ogasawara
  • ,
  • Takafumi Ichikawa
  • ,
  • Atsushi Kodan
  • ,
  • Yasuhisa Kimura
  • ,
  • Noriyuki Kioka
  • ,
  • Kazumitsu Ueda

295
15
開始ページ
5002
終了ページ
5011
記述言語
掲載種別
研究論文(学術雑誌)
DOI
10.1074/jbc.ra119.012042
出版者・発行元
American Society for Biochemistry & Molecular Biology (ASBMB)

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.

リンク情報
DOI
https://doi.org/10.1074/jbc.ra119.012042
URL
https://syndication.highwire.org/content/doi/10.1074/jbc.RA119.012042

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