ABCC1 p.Asp360Lys
| Predicted by SNAP2: | A: N (82%), C: N (78%), E: N (93%), F: N (57%), G: N (82%), H: N (72%), I: N (72%), K: N (78%), L: N (72%), M: N (61%), N: N (93%), P: N (66%), Q: N (87%), R: N (72%), S: N (93%), T: N (93%), V: N (72%), W: D (75%), Y: D (63%), |
| Predicted by PROVEAN: | A: N, C: N, E: N, F: N, G: N, H: N, I: N, K: N, L: N, M: N, N: N, P: N, Q: N, R: N, S: N, T: N, V: N, W: D, Y: N, |
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[hide] Mutations of charged amino acids in or near the tr... Mol Pharmacol. 2004 Jun;65(6):1375-85. Haimeur A, Conseil G, Deeley RG, Cole SP
Mutations of charged amino acids in or near the transmembrane helices of the second membrane spanning domain differentially affect the substrate specificity and transport activity of the multidrug resistance protein MRP1 (ABCC1).
Mol Pharmacol. 2004 Jun;65(6):1375-85., [PMID:15155831]
Abstract [show]
Multidrug resistance protein 1 (MRP1) belongs to the ATP-binding cassette superfamily of transport proteins. In addition to drugs, MRP1 mediates the active transport of many conjugated and unconjugated organic anions. MRP1 consists of two membrane-spanning domains (MSD2 and MSD3) each followed by a nucleotide binding domain plus a third NH2-terminal MSD1. MSD2 contains transmembrane (TM) helices 6 through 11, and previously, we identified two charged residues in TM6 as having important but markedly different roles in MRP1 transport activity and substrate specificity by characterizing mutants containing nonconservative substitutions of Lys332 and Asp336. We have now extended these studies and found that the same-charge TM6 mutant K332R, like the nonconservatively substituted Lys332 mutants, exhibits a selective decrease in leukotriene C4 (LTC4) transport, associated with substantial changes in both Km and Vmax and LTC4 binding. The overall organic anion transport activity of the same-charge mutant of Asp336 (D336E) also remained very low, as observed for D336R. In addition, nonconservative substitutions of TM6-associated Lys319 and Lys347 resulted in a selective decrease in GSH transport. Of eight other charged residues in or proximal to TM7 to TM11 that were investigated, nonconservative substitutions of three of them [Lys396 (TM7), Asp436 (TM8), and Arg593 (TM11)] caused a substantial and global reduction in transport activity. However, unlike TM6 Asp336, wild-type transport activity could be reestablished in these MRP1 mutants by conservative substitutions. We conclude that MSD2-charged residues in or proximal to TM6, TM7, TM8, and TM11 play critical but differential roles in MRP1 transport activity and substrate specificity.
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No. Sentence Comment
48 The sequences of the individual sense strands (with the corresponding amino acid changes indicated in parentheses, the altered codons underlined, and silent mutations introducing new restriction sites italicized) were as follows: K(D)332K, 5Ј-G AGC TTC TTC TTC AAG GCC ATC CAC GAC CTG-3Ј; K332R, 5Ј-G AGC TTC TTC TTC AGG GCC ATC CAC GAC CTG-3Ј; D(K)336E, 5Ј-C AAG GCC ATC CAC GAG CTC ATG ATG TTT TCC-3Ј; D336K, 5Ј-C AAG GCC ATC CAC AAG CTT ATG ATG TTT TCC-3Ј; K332D/D336K, 5Ј-GC TTC TTC TTC GAC GCC ATC CAC AAA CTG ATG ATG-3Ј; K319D, 5Ј-G TTT AAG GTG TTA TAC GAC ACG TTT GGG CCC-3Ј; K347D, 5Ј-GGG CCG CAG ATA TTA GAC TTG CTC ATC AAG-3Ј; K347L, 5Ј-GGG CCG CAA ATC TTA CTT TTG CTC ATC AAG-3Ј; D360K, 5Ј-GAC ACG AAG GCG CCA AAG TGG CAG GGC TAC-3Ј; R394D, 5Ј-C GTC AGT GGC ATG GAG ATC AAG ACC GCT GTC-3Ј; R394I, 5Ј-C GTC AGT GGC ATG ATC ATC AAG ACC GCT GTC-3Ј; K396E, 5Ј-GT GGC ATG AGG ATC GAG ACC GCT GTC ATT GGG-3Ј; K396I, 5Ј-GT GGC ATG AGG ATC ATC ACC GCT GTC ATT GGG-3Ј; K(E)396R, 5Ј-GGC ATG AGG ATC AGG ACC GCT GTC ATT GGG GC-3Ј; D430K, 5Ј-C AAC CTC ATG TCT GTG AAG GCT CAG AGG Fig. 1.
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ABCC1 p.Asp360Lys 15155831:48:787
status: NEW142 Asp360 was replaced with an oppositely charged Lys residue (D360K), whereas Arg394 and Lys396 were substituted with Asp and Glu, respectively, to introduce the opposite charge (R394D and K396E) and with a nonpolar neutral amino acid (Ile) to eliminate charge (R394I and K396I).
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ABCC1 p.Asp360Lys 15155831:142:60
status: NEW148 MRP1 Mutant % Wild-Type MRP1 Transport Activity LTC4 GSH E217betaG E13SO4 MTX TM6 K332D Ͻ10 Ͻ10 100 100 100 K332R 40 20 100 100 100 D336K Ͻ10 Ͻ10 Ͻ10 15 45 D(K)336E 20 Ͻ10 25 25 45 K332D/D336K Ͻ10 Ͻ10 30 25 50 K319D 100 50 100 100 100 K347D 100 45 100 100 100 TM7 D360K 100 100 100 100 100 R394D 100 100 100 100 100 K396E 25 50 25 25 35 K(E)396R 100 100 100 100 100 TM8 D436K 40 30 20 30 55 D(K)436E 100 100 100 100 100 ECL5/TM11 D572R 100 100 100 100 100 E573R 100 100 100 100 100 D578R 100 100 100 100 100 R593E 35 30 Ͻ10 Ͻ10 40 R(E)593K 100 100 100 100 100 or substrate specificity of MRP1.
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ABCC1 p.Asp360Lys 15155831:148:312
status: NEW