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Structure‐guided probing of the leukotriene C4 binding site in human multidrug resistance protein 1 (MRP1; ABCC1)

The human multidrug resistance protein 1 (hMRP1) transporter is implicated in cancer multidrug resistance as well as immune responses involving its physiologic substrate, glutathione (GSH)‐conjugated leukotriene C4 (LTC4). LTC4 binds a bipartite site on hMRP1, which a recent cryoelectron microscopy structure of LTC4‐bound bovine Mrp1 depicts as composed of a positively charged pocket and a hydrophobic (H) pocket that binds the GSH moiety and surrounds the fatty acid moiety, respectively, of LTC4. Here, we show that single Ala and Leu substitutions of H‐pocket hMRP1‐Met1093 have no effect on LTC4 binding or transport. Estrone 3‐sulfate transport is also unaffected, but both hMRP1‐Met1093 mutations eliminate estradiol glucuronide transport, demonstrating that these steroid conjugates have binding sites distinct from each other and from LTC4. To eliminate LTC4 transport by hMRP1, mutation of 3 H‐pocket residues was required (W553/M1093/W1246A), indicating that H‐pocket amino acids are key to the vastly different affinities of hMRP1 for LTC4 vs. GSH alone. Unlike organic anion transport, hMRP1‐mediated drug resistance was more diminished by Ala than Leu substitution of Met1093. Although our findings generally support a structure in which H‐pocket residues bind the lipid tail of LTC4, their critical and differential role in the transport of conjugated estrogens and anticancer drugs remains unexplained.—Conseil, G., Arama‐Chayoth, M., Tsfadia, Y., Cole, S. P. C. Structure‐guided probing of the leukotriene C4 binding site in human multidrug resistance protein 1 (MRP1; ABCC1). FA8EB J. 33, 10692–10704 (2019). www.fasebj.org