ABCMdb

ABCC1 p.Trp459Ala

None has been submitted yet.

Publications

PMID: 12388549 [PubMed] Koike K et al: "Multiple membrane-associated tryptophan residues contribute to the transport activity and substrate specificity of the human multidrug resistance protein, MRP1."

No. Sentence Comment

48 Tryptophan substitutions were generated in the pGEM-3Z and pBluescriptSK(ϩ) plasmids above according to the manufacturer`s instructions with the following mutagenic primers (substituted nucleotides are underlined): MSD1 Trp mutants W40A (5Ј-G GTC CTC GTG GCC GTG CCT TG-3Ј, W47A (5Ј-GT TTT TAC CTC GCC GCC TGT TTC CCC-3Ј), W82A (5Ј-C TTG GGA TTT TTG CTG GCG ATC GTC TGC TGG GC-3Ј), W86A (5Ј-G CTG TGG ATC GTC TGC GCT GCA GAC CTC TTC TAC TC-3Ј), W94A (5Ј-C CTC TTC TAC TCT TTC GCG GAA AGA AGT CGG GGC-3Ј), W142A (5Ј-GGG ATC ATG CTC ACT TTC GCA CTG GTA GCC CTA ATG TG-3Ј), W142F (5Ј-G CTC ACT TTT TTC CTG GTA GCC C-3Ј); MSD2 Trp mutants W361A (5Ј-C ACG AAG GCC CCA GAT GCG CAG GGC TAC TTC TAC-3Ј), W445A (5Ј-G TAC ATT AAC ATG ATC GCG TCA GCC CCC CTG CAA G-3Ј), W445F (5Ј-CG TAC ATT AAC ATG ATC TTC TCA GCC CCC CTG CAA GTC-3Ј), W445Y (5Ј-CC ACG TAC ATT AAC ATG ATC TAC TCA GCG CCC CTG CAA GTC-3Ј), W459A (5Ј-GCT CTC TAC CTC CTG GCG CTG AAT CTG GGC CC-3Ј), W553A (5Ј-G GGC ACC TTC ACC GCG GTC TGC ACG CCC-3Ј), W553F (5Ј-G GGC ACC TTC ACC TTC GTC TGC ACG CCC-3Ј), W553Y (5Ј-GCC CTG GGC ACC TTC ACA TAT GTC TGC ACG CCC-3Ј; and MSD3 Trp mutants W1198A (5Ј-C GTG GCC AAC AGG GCG CTG GCC GTG CGG C-3Ј), W1198F (5Ј-GTG GCC AAC AGG TTC CTG GCC GTG CGG C-3Ј), W1198Y (5Ј-GTG GCC AAC AGG TAC CTG GCC GTG CGG C-3Ј). Login to comment
108 All four mutants generated (W361A, W445A, W459A, and W553A) were expressed at levels 60-90% those of wild-type MRP1, indicating that none of the mutations had a major effect on the expression levels of the protein. Login to comment
110 After 1 min, ATP-dependent [3 H]LTC4 uptake by the W445A and W553A MRP1 mutants was reduced by ϳ75 and 50%, respectively, whereas uptake by the W361A and W459A mutants was comparable with wild-type MRP1 (Fig. 3B). Login to comment
112 The ATP-dependent uptake rate of this substrate by the W459A MRP1 mutant, as for LTC4 uptake, was comparable with that of wild-type MRP1 (Fig. 3C). Login to comment
119 Similarly, GSH-stimulated E13SO4 uptake levels by the W445A and W553A mutants were just 30 and Ͻ10% of wild-type MRP1 levels, respectively, whereas uptake by the W361A mutant was similar to wild-type MRP1, and uptake by W459A MRP1 was reduced by just 25% (Fig. 4B). Login to comment
121 A, ATP-dependent uptake of [3 H]LTC4 was measured in membrane vesicles prepared from HEK293T cells transfected with empty vector pcDNA3.1(-) (E) and vectors containing wild-type MRP1 (f) and MSD2 Trp-Ala mutant MRP1 cDNAs (W361A, Œ; W445A, ; W459A, ࡗ; and W553A, q). Login to comment
124 C, the time course of ATP-dependent uptake of [3 H]E217betaG by wild-type MRP1 and MSD2 mutants W361A, W445A, W459A, and W553A was measured as described for A. D, relative levels of [3 H]E217betaG uptake at 1 min are shown and were determined from the time course shown in C as described for B. Login to comment
127 MTX uptake by the W361A mutant was also reduced (ϳ40%), whereas uptake by the W459A mutant was similar to that of wild-type MRP1. Login to comment
153 Relative uptake levels of 3 H-labeled organic anions by membrane vesicles enriched for wild-type MRP1 (solid bar) and W361A, W445A, W459A, and W553A mutant MRP1 proteins (shaded bars) were determined as described under "Experimental Procedures." Login to comment
180 The W459A mutant displayed a slightly different phenotype in that its LTC4, E217betaG, and MTX transport activities were similar to wild-type MRP1, but GSH and E13SO4 transport activities were reduced by 25-50%. Login to comment