Fluorescent modified phosphatidylcholine floppase activity of reconstituted multidrug resistance-associated protein MRP1

Zhenhua Huang, Xiu-Bao D Chang, John R. Riordan, Youguo Huang

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

Multidrug resistance-associated protein (MRP1) may function as a floppase in human red blood cells to translocate phosphatidylserine and/or phosphatidylcholine from inner membrane leaflet to outer leaflet. Here we report that the purified and reconstituted MRP1 protein into asolectin proteoliposomes is mainly in an inside-out configuration and possesses the ability to flop a fluorescent labeled phosphatidylcholine (NBD-PC) from outer leaflet (protoplasmic) to inner leaflet (extrocytoplasmic). The reconstituted MRP1 protein retains endogenous ATPase activity. ATP hydrolysis is required for the flopping since removal of ATP and/or Mg2+ inhibits the translocation of NBD-PC. Further evidence to support this conclusion is that the translocation of NBD-PC is inhibited by vanadate, which traps ATP hydrolysis product ADP in the nucleotide binding domains. In addition, the translocation of NBD-PC by proteoliposomes containing MRP1 protein is in a glutathione-dependent manner, similar to the process of translocating anticancer drugs such as daunorubicin. verapamil, vincristine, vinblastine, doxorubicin and oxidized glutathione partially inhibited the translocation of NBD-PC, whereas MK 571, an inhibitor of MRP1 protein, inhibited the translocation almost completely. Taken together, the purified and reconstituted MRP1 protein possesses the ability to flop NBD-PC from outer to inner leaflet of the proteoliposomes.

Original languageEnglish (US)
Pages (from-to)155-163
Number of pages9
JournalBiochimica et Biophysica Acta - Biomembranes
Volume1660
Issue number1
DOIs
StatePublished - Jan 28 2004

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Multidrug Resistance-Associated Proteins
Phosphatidylcholines
Adenosine Triphosphate
verlukast
Proteins
Hydrolysis
Daunorubicin
Vanadates
Glutathione Disulfide
Vinblastine
Phosphatidylserines
Vincristine
Protein Transport
Verapamil
Doxorubicin
Adenosine Diphosphate
Glutathione
Adenosine Triphosphatases
1-acyl-2-(12-((7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)dodecanoyl)phosphatidylcholine
Blood

Keywords

  • Floppase
  • MRP1
  • NBD-PC

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Biophysics

Cite this

Fluorescent modified phosphatidylcholine floppase activity of reconstituted multidrug resistance-associated protein MRP1. / Huang, Zhenhua; Chang, Xiu-Bao D; Riordan, John R.; Huang, Youguo.

In: Biochimica et Biophysica Acta - Biomembranes, Vol. 1660, No. 1, 28.01.2004, p. 155-163.

Research output: Contribution to journalArticle

Huang, Zhenhua ; Chang, Xiu-Bao D ; Riordan, John R. ; Huang, Youguo. / Fluorescent modified phosphatidylcholine floppase activity of reconstituted multidrug resistance-associated protein MRP1. In: Biochimica et Biophysica Acta - Biomembranes. 2004 ; Vol. 1660, No. 1. pp. 155-163.
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