Int J Biochem Mol Biol 2011;2(1):39-46
Original Article
Photoaffinity Labeling of the Multidrug Resistance Protein 2 (ABCC2/cMOAT) with a
Photoreactive Analog of LTC4
Mara L. Leimanis, Joel Karwatsky, Elias Georges
Institute of Parasitology, Macdonald Campus, McGill University, Montreal, Canada
Received December 1, 2010; accepted December 12, 2010; Epub December 15, 2010; published February 15, 2011
Abstract: Several studies have shown that the multidrug resistant protein MRP2 mediates the transport of chemotherapeutic drugs and normal
cell metabolites, including Leukotriene C (LTC4); however direct binding of the LTC4 to MRP2 has not been demonstrated. In this study, a
photoreactive analog of LTC4 (IAALTC4) was used to demonstrate its direct binding to MRP2. Our results show specific photoaffinity labeling
of MRP2 with IAALTC4 in plasma membranes from MDCKIIMRP2 cells. The photoaffinity labeling signal of MRP2 with IAALTC4 was much
lower than that of MRP1, consistent with previous studies whereby the measured Km values of MRP1 and MRP2 for LTC4 1 µM and 0.1 µM
LTC4, respectively. Competition of IAALTC4 photoaffinity labeling to MRP2 with MK571, a well characterized inhibitor of MRP2 function, showed
~75% reduction in binding in the presence of 50 µM excess MK571. Interestingly, unmodified LTC4 enhanced the photoaffinity labeling of
IAALTC4 to MRP2, whereas excess GSH and Quercetin had no significant effect. Mild tryptic digestion of photoaffinity labeled MRP2 revealed
several photoaffinity labeled peptides that localized the IAALTC4 binding to a 15 kDa amino acid sequence that contains transmembrane 16
and 17. Together these results provide the first demonstration of direct LTC4 binding to MRP2. (IJBMB1012001).
Keywords: Multi-drug resistance, multi-drug resistance protein 2, leukotriene C4, drug binding, photoaffinity labeling
Full Text PDF
Address all correspondence to:
Dr. Elias Georges
Institute of Parasitology
Macdonald Campus, McGill University
Ste. Anne de Bellevue (Montreal)
Canada, H9X-3V9;
Tel: 514-398-8137; Fax: 514-398-7857
E-mail: elias.georges@mcgill.ca
Original Article
Photoaffinity Labeling of the Multidrug Resistance Protein 2 (ABCC2/cMOAT) with a
Photoreactive Analog of LTC4
Mara L. Leimanis, Joel Karwatsky, Elias Georges
Institute of Parasitology, Macdonald Campus, McGill University, Montreal, Canada
Received December 1, 2010; accepted December 12, 2010; Epub December 15, 2010; published February 15, 2011
Abstract: Several studies have shown that the multidrug resistant protein MRP2 mediates the transport of chemotherapeutic drugs and normal
cell metabolites, including Leukotriene C (LTC4); however direct binding of the LTC4 to MRP2 has not been demonstrated. In this study, a
photoreactive analog of LTC4 (IAALTC4) was used to demonstrate its direct binding to MRP2. Our results show specific photoaffinity labeling
of MRP2 with IAALTC4 in plasma membranes from MDCKIIMRP2 cells. The photoaffinity labeling signal of MRP2 with IAALTC4 was much
lower than that of MRP1, consistent with previous studies whereby the measured Km values of MRP1 and MRP2 for LTC4 1 µM and 0.1 µM
LTC4, respectively. Competition of IAALTC4 photoaffinity labeling to MRP2 with MK571, a well characterized inhibitor of MRP2 function, showed
~75% reduction in binding in the presence of 50 µM excess MK571. Interestingly, unmodified LTC4 enhanced the photoaffinity labeling of
IAALTC4 to MRP2, whereas excess GSH and Quercetin had no significant effect. Mild tryptic digestion of photoaffinity labeled MRP2 revealed
several photoaffinity labeled peptides that localized the IAALTC4 binding to a 15 kDa amino acid sequence that contains transmembrane 16
and 17. Together these results provide the first demonstration of direct LTC4 binding to MRP2. (IJBMB1012001).
Keywords: Multi-drug resistance, multi-drug resistance protein 2, leukotriene C4, drug binding, photoaffinity labeling
Full Text PDF
Address all correspondence to:
Dr. Elias Georges
Institute of Parasitology
Macdonald Campus, McGill University
Ste. Anne de Bellevue (Montreal)
Canada, H9X-3V9;
Tel: 514-398-8137; Fax: 514-398-7857
E-mail: elias.georges@mcgill.ca
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