The high turnover Drosophila multidrug resistance-associated protein shares the biochemical features of its human orthologues.
DMRP, an ABC transporter encoded by the dMRP/CG6214 gene, is the Drosophila melanogaster orthologue of the "long" human multidrug resistance-associated proteins (MRP1/ABCC1, MRP2/ABCC2, MRP3/ABCC3, MRP6/ABCC6, and MRP7/ABCC10). In order to provide a detailed biochemical characterisation we expressed DMRP in Sf9 insect cell membranes. We demonstrated DMRP as a functional orthologue of its human counterparts capable of transporting several human MRP substrates like beta-estradiol 17-beta-D-glucuronide, leukotriene C4, calcein, fluo3 and carboxydichlorofluorescein. Unexpectedly, we found DMRP to exhibit an extremely high turnover rate for the substrate transport as compared to its human orthologues. Furthermore, DMRP showed remarkably high basal ATPase activity (68-75 nmol Pi/mg membrane protein/min), which could be further stimulated by probenecid and the glutathione conjugate of N-ethylmaleimide. Surprisingly, this high level basal ATPase activity was inhibited by the transported substrates. We discussed this phenomenon in the light of a potential endogenous substrate (or activator) present in the Sf9 membrane.
Szeri
Flóra
F
Institute of Enzymology, Hungarian Academy of Sciences, Budapest, P.O. Box 7, H-1518, Hungary. szeri@enzim.hu
Iliás
Attila
A
Pomozi
Viola
V
Robinow
Steven
S
Bakos
Eva
E
Váradi
András
A
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D004331:Drosophila melanogaster / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018411:Spodoptera; D013379:Substrate Specificity
D000251:Adenosine Triphosphatases; D000255:Adenosine Triphosphate; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; C000715087:ABCC2 protein, human