title: Functional interactions between nucleotide binding domains and leukotriene C4 binding sites of multidrug resistance protein 1 (ABCC1).
abstract: Multidrug resistance protein 1 (MRP1) is a member of the "C" branch of the ATP-binding cassette transporter superfamily. The NH(2)-proximal nucleotide-binding domain (NBD1) of MRP1 differs functionally from its COOH-proximal domain (NBD2). NBD1 displays intrinsic high-affinity ATP binding and little ATPase activity. In contrast, ATP binding to NBD2 is strongly dependent on nucleotide binding by NBD1, and NBD2 is more hydrolytically active. We have demonstrated that occupancy of NBD2 by ATP or ADP markedly decreased substrate binding by MRP1. We have further explored the relationship between nucleotide and substrate binding by examining the effects of various ATP analogs and ADP trapping, as well as mutations in conserved functional elements in the NBDs, on the ability of MRP1 to bind the photoactivatable, high-affinity substrate cysteinyl leukotriene C(4) (LTC(4))(.) Overall, the results support a model in which occupancy of both NBD1 and NBD2 by ATP results in the formation of a low-affinity conformation of the protein. However, nonhydrolyzable ATP analogs (beta,gamma-imidoadenosine 5'-triphosphate and adenylylmethylene diphosphonate) failed to substitute for ATP or adenosine 5'-O-(thiotriphosphate) (ATPgammaS) in decreasing LTC(4) photolabeling. Furthermore, mutations of the signature sequence in either NBD that had no apparent effect on azido-ATP binding abrogated the formation of a low-affinity substrate binding state in the presence of ATP or ATPgammaS. We suggest that the effect of these mutations, and possibly the failure of some ATP analogs to decrease LTC(4) binding, may be attributable to an inability to elicit a conformational change in the NBDs that involves interactions between the signature sequence and the gamma-phosphate of the bound nucleotide.
authors.affiliation: Division of Camcer Biology and Genetics, Cancer Research Institute, Queen's University, 10 Stuart Street, Kingston, Ontario, K7L 3N6 Canada.
authors.forename: Lea
authors.initials: L
authors.lastname: Payen
authors.forename: Mian
authors.initials: M
authors.lastname: Gao
authors.forename: Christopher
authors.initials: C
authors.lastname: Westlake
authors.forename: Ashley
authors.initials: A
authors.lastname: Theis
authors.forename: Susan P C
authors.initials: SP
authors.lastname: Cole
authors.forename: Roger G
authors.initials: RG
authors.lastname: Deeley
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000502:physiology; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D007313:Insecta; D047908:Intracellular Signaling Peptides and Proteins; D017997:Leukotriene C4 / Q000378:metabolism*; D008969:Molecular Sequence Data; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D017997:Leukotriene C4; D027425:ATP-Binding Cassette, Sub-Family C Proteins; C089765:NUBP1 protein, human; C421109:multidrug resistance-associated protein 1