ABCMdb

ABCC1 p.Lys332Asp

[switch to full view]
Comments [show]

None has been submitted yet.

Publications

PMID: 12186871 [PubMed] Haimeur A et al: "Charged amino acids in the sixth transmembrane helix of multidrug resistance protein 1 (MRP1/ABCC1) are critical determinants of transport activity."

No. Sentence Comment

47 The sequences of the individual sense strands, with the altered codons underlined and the corresponding changes in amino acids indicated in parentheses were as follows: (K332D) 5Ј-CTC ATG AGC TTC TTC TTC GAC GCC ATC CAC GAC CTG-3Ј; (K332L) 5Ј-CTC ATG AGC TTC TTC TTC CTG GCC ATC CAC GAC CTG-3Ј; (H335E) 5Ј-GC TTC TTC TTC AAG GCC ATC GAG GAC CTG ATG ATG-3Ј; (H335L) 5Ј-GC TTC TTC TTC AAG GCC ATC TTG GAC CTG ATG ATG-3Ј; (H335Q) 5Ј-GC TTC TTC TTC AAG GCC ATC CAG GAC CTG ATG ATG-3Ј; (D336R) 5Ј-C AAG GCC ATC CAC CGG CTG ATG ATG TTT TCG-3Ј; (D336L) 5Ј-C AAG GCC ATC CAC CTG CTG ATG ATG TTT TCG-3Ј. Login to comment
104 Representative confocal micrographs of cells expressing GFP-tagged wild-type MRP1 and mutants K332D, H335E, and D336R are shown in Fig. 2. Login to comment
106 In the case of the MRP1-Lys332 mutants K332D and K332L (Fig. 3B) and the MRP1-Asp336 mutants D336L and D336R (Fig. 3D), LTC4 uptake was reduced to levels that were indistinguishable from those observed with vesicles prepared from empty vector-transfected control cells. Login to comment
131 In contrast, LTC4 had very little effect (Ͻ15%) on E217betaG uptake by MRP1 mutants K332D and K332L, indicating that loss of LTC4 transport in these mutants is associated with a loss of binding of this substrate. On the other hand, LTC4 was still able to inhibit E217betaG uptake by MRP1 mutants H335E, H335L, and H335Q, which is consistent with only a partial reduction in LTC4 transport activity observed with these mutants (Fig. 6B). Login to comment
140 B, wild-type MRP1 (f), MRP1 mutants K332D (Œ) and K332L (‚), and control empty pcDNA3.1(-) vector (E). Login to comment
152 However, this band is not detectable in [3 H]LTC4 photolabeled proteins from cells expressing comparable levels of the MRP1 mutants K332D and K332L or mutants D336L and D336R, indicating that these mutations abrogate photolabeling and hence binding of this compound. Login to comment
160 The relative levels of [3 H]GSH uptake by the MRP1 K332D and K332L mutants were less than 15% of wild-type MRP1 (Fig. 7A) after subtracting basal [3 H]GSH transport by membrane vesicles from the empty vector-transfected control cells. Login to comment
163 Membrane vesicles prepared from transfected cells were incubated at 37 °C with 400 nM [3 H]E217betaG in transport buffer for the times indicated. A, time courses of ATP-dependent [3 H]E217betaG uptake by membrane vesicles prepared from HEK293T cells transfected with wild-type MRP1 (f), TM6 mutants K332D (Œ) and K332L (‚), and the empty pcDNA3.1(-) vector (E). Login to comment
170 A, ATP-dependent [3 H]E217betaG uptake in membrane vesicles from cells expressing wild-type MRP1 (WT-MRP1) (open bars), TM6 mutants K332D and K332L (shaded bars), and the empty pcDNA3.1(-) vector control (solid bars). Login to comment
183 Substitution of Lys332 with a neutral (K332L) or negatively charged (K332D) amino acid had no effect on MTX uptake by MRP1. Login to comment
190 Thus, replacing Lys332 with either Leu or Asp eliminated the ability of MRP1 to transport LTC4 (and markedly reduced GSH transport) without affecting the transport of other organic anions including E217betaG, estrone 3-sulfate, and MTX. Login to comment
204 Membrane vesicles were preincubated with acivicin and then incubated with [3 H]GSH in the presence of 30 ␮M apigenin in transport buffer for 20 min at 37 °C. A, K332D and K332L; B, H335E, H335L, and H335Q; C, D336L and D336R. Login to comment
206 D, WT-MRP1 (f); K332D (Œ); K332L (‚); vector control (E). Login to comment

PMID: 12731862 [PubMed] Leslie EM et al: "Functional and structural consequences of cysteine substitutions in the NH2 proximal region of the human multidrug resistance protein 1 (MRP1/ABCC1)."

No. Sentence Comment

208 This contrasts with other amino acid substitutions of MRP1 (such as when Lys332 is replaced by Asp or Leu) that completely eliminate LTC4 transport without disrupting plasma membrane trafficking (43). Login to comment

PMID: 15155831 [PubMed] Haimeur A et al: "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)."

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. Login to comment
100 We showed previously that nonconservative substitutions of Lys332 with either Asp (K332D) or Leu (K332L) led to a selective loss of transport of GSH and the GSH conjugate, LTC4 (Haimeur et al., 2002). Login to comment
102 When the Asp residue of the K332D mutant was mutated back to the wild-type Lys residue to create K(D)332K, LTC4 and GSH transport was restored as expected (Fig. 3, A and B). Login to comment
104 Unlike the K332D mutant, the conservatively substituted K332R had a significant level of LTC4 transport activity (approximately 40% of wild-type MRP1 levels). Login to comment
113 A, time course of [3 H]LTC4 uptake by wild-type MRP1 (f), mutants K332D (), K(D)332K (F), K332R (Œ), and empty pcDNA3.1(-) vector control (E). Login to comment
114 B, [3 H]GSH uptake at 20 min by wild-type MRP1 (f) and mutants K332R, K332D, and K(D)332K (u), and empty pcDNA3.1(-) vector control (Ⅺ). Login to comment
115 C, [3 H]E217betaG uptake at 1 min by wild-type (WT) MRP1 (f) and mutants K332D () and K332R (Œ) was measured in the presence of three different concentrations (300, 600, and 900 nM) of LTC4. Login to comment
119 The dose-response curves shown in Fig. 3C indicate that LTC4 had a greater inhibitory effect on E217betaG uptake by the K332R mutant than by the K332D mutant, but this effect was significantly less (50-70%) than the effect of LTC4 on E217betaG uptake by wild-type MRP1. Login to comment
121 The same-charge mutant K332R, like the K332D and K332L mutants described previously (Haimeur et al., 2002), exhibited transport levels of the conjugated estrogens E217betaG and E13SO4 and the antifolate MTX that were comparable with wild-type MRP1 (Table 2); however, GSH transport by K332R was very low compared with wild-type MRP1 and similar to that which we reported previously for the K332D/L mutants (Fig. 3B). Login to comment
130 To test this idea, the K332D/D336K double mutant in which these two residues were exchanged was created. Login to comment
132 However, as shown in Fig. 4B, LTC4 transport by K332D/ D336K was not detectable. Login to comment
148 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. Login to comment
176 B, time course of [3 H]LTC4 uptake by wild-type MRP1 (f), TM6 double mutant K332D/D336K (ࡗ), and empty pcDNA3.1(-) vector control (E). Login to comment
194 For these experiments, GFP-tagged constructs encoding wild-type MRP1 and the mutant MRP1 proteins (D336E, K332D/D336K, K396E, D436K, and R593E) were generated and transfected into HEK 293T cells (Koike et al., 2002). Login to comment
211 However, the inactivity of the double-exchange mutant K332D/D336K suggests that this is unlikely to be the case (Fig. 2). Login to comment
272 HEK 293T cells were transfected with the wild-type pcDNA3.1(-)-MRP1-GFP and mutant pcDNA3.1(-)-MRP1- D336E-GFP, pcDNA3.1(-)-MRP1-K332D/D336K-GFP, pcDNA3.1(-)- MRP1-K396E-GFP, pcDNA3.1(-)-MRP1-D436K-GFP, and pcDNA3.1(-)- MRP1-R593E-GFP expression vectors as indicated, and cells were viewed 48 h later under the confocal microscope. Login to comment

PMID: 17295059 [PubMed] Chang XB et al: "A molecular understanding of ATP-dependent solute transport by multidrug resistance-associated protein MRP1."

No. Sentence Comment

109 P343A, K332L and K332D mutations in TM6 resulted in significantly reduced transport of some organic anion substrates [75, 76]. Login to comment
154 Mutations in TM6, such as K332D or K332R, significantly reduced ATP-dependent LTC4 or GSH transport, but did not have any effect on ATP-dependent E217βG or E13SO4 transport [94]. Login to comment

PMID: 18775981 [PubMed] Grant CE et al: "Structural determinants of substrate specificity differences between human multidrug resistance protein (MRP) 1 (ABCC1) and MRP3 (ABCC3)."

No. Sentence Comment

305 In addition, photolabeling with [3 H]LTC4 of K332D and K332L mutant proteins was severely reduced compared with wild-type MRP1. Login to comment

PMID: 19949927 [PubMed] Chang XB et al: "Molecular mechanism of ATP-dependent solute transport by multidrug resistance-associated protein 1."

No. Sentence Comment

104 Mutations of C43S in TM1 (112); P343A, K332L and K332D in TM6 (113, 114); W445A and P448A in TM8 (113, 115); T550A, T556A and P557A in TM10 (113, 116); N590A, F594A, P595A, N597A, S604A and S605A in TM11 (113, 117, 118); E1089Q, E1089A, E1089L, E1089N, K1092, S1097 and N1100 in TM14 (119, 120); R1197K in TM16 (121); Y1236F, T1241A, T1242A, T1242C, T1242S, T1242L, Y1243F, N1245A, W1246C, W1246A, W1246F, W1246Y or R1249K in TM17 (121-124) significantly affect MRP1 function. Login to comment