ABCMdb

ABCC1 p.Arg1138Glu

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Publications

PMID: 16230346 [PubMed] Conseil G et al: "Functional importance of three basic residues clustered at the cytosolic interface of transmembrane helix 15 in the multidrug and organic anion transporter MRP1 (ABCC1)."

No. Sentence Comment

47 Substitutions of charged residues were generated in the pGEM-3Z- XmaI/MRP1 plasmid according to the manufacturer`s instructions with the following mutagenic primers (Integrated DNA Technologies, Inc., Coralville, IA), which also introduced an additional silent restriction site (substituted nucleotides are underlined): R1138E, 5Ј-CGT GGC TTC CTC CGA GCA ACT GAA GCG CCT CGA G-3Ј; R1138K, 5Ј-CGT GGC TTC CTC CAA GCA GCT TAA GCG CCT CGA G-3Ј; K1141E, 5Ј-CGG CAG CTG GAG CGC CTG GAG TCG GTC AGC-3Ј; K1141R, 5Ј-CCC GGC AGC TGC GGC GCC TCG AGT CGG-3Ј; R1142E, 5Ј-CCC GGC AGC TGA AGG AGC TCG AGT CGG TCA GCC G-3Ј;R1142K, 5Ј-CCC GGC AGC TGA AGA AGC TTG AGT CGG TCA GCC G-3Ј. Login to comment
118 A, membrane vesicles (1 ␮g of protein) prepared from HEK293T cells transfected with wild-type (WT-MRP1), and mutant (R1138E/K, K1141E/R, R1142E/K) MRP1 cDNAs were immunoblotted, and MRP1 was detected with monoclonal antibody QCRL-1. Login to comment
122 B, HEK293T cells were transfected with expression vectors encoding WT-MRP1 and MRP1 mutants R1138E, R1138K, K1141E, K1141R, R1142E, and R1142K, and 48 h later, cells were probed with monoclonal antibody QCRL-3 and processed for confocal fluorescence microscopy using Alexa Fluor 488 conjugated secondary antibody. Login to comment
135 Finally, the effects of the mutations on MTX transport were relatively minor and ranged from no decrease to a maximum decrease of 35% (R1138E) when compared with wild-type MRP1 (Fig. 3D). Login to comment
136 Photolabeling of Arg1138 , Lys1141 , and Arg1142 Mutants of MRP1 with [3 H]LTC4-To ascertain whether the decrease in LTC4 uptake by the R1138E/K, K1141E, and R1142E/K mutants was related to a decrease in substrate binding, membrane vesicles enriched for wild-type or mutant MRP1s were photolabeled with [3 H]LTC4. Login to comment
137 As shown in Fig. 4, LTC4 labeling of R1138E and K1141E was decreased by 50% or more, after correcting for differences in protein expression levels, consistent with their decreased LTC4 transport activity. Login to comment
139 Therefore, reduced substrate binding appeared to explain, at least in part, the loss of LTC4 transport for the R1138E and K1141E mutants but not the R1138K, R1142E, and R1142K mutants. Login to comment
149 ATP-dependent uptake of 3 H-labeled organic anions was measured in membrane vesicles prepared from HEK293T cells transfected with wild-type MRP1 (WT-MRP1) and Arg1138 , His1141 and Arg1142 mutant MRP1 (R1138E/K, K1141E/R, R1141E/K) cDNAs (shown in Fig. 2). Login to comment
163 In contrast, when the catalytic activity of MRP1 was measured at 37 °C by the orthovanadate-induced trapping of 8-azido-[␣32 P]ADP in the post-hydrolysis state (Fig. 6B), five of the six mutants (R1138E/K, R1142E/K, and K1141E) showed substantially reduced nucleotide trapping (ϳ40-60% that of wild-type MRP1). Login to comment
200 The reduction in GSH and LTC4 transport appeared largely attributable to a substantial decrease in substrate binding in the case of R1138E but not R1138K. Login to comment

PMID: 21143116 [PubMed] He SM et al: "Structural and functional properties of human multidrug resistance protein 1 (MRP1/ABCC1)."

No. Sentence Comment

739 In contarst, the mutations showed minor to moderate effect on MTX transport, with a maximum decrease for Arg1138Glu). Login to comment
744 The Arg1138Glu and Lys1141Glu mutants, but not the Arg1138Lys, Arg1142Glu, and Arg1142Lys mutants, showed a >50% decrease in binding to [3 H]LTC4. Login to comment