ABCB1 p.Val331Ala
| Predicted by SNAP2: | A: N (78%), C: N (57%), D: D (80%), E: D (75%), F: N (72%), G: D (75%), H: D (75%), I: N (87%), K: D (75%), L: N (93%), M: N (93%), N: D (75%), P: D (80%), Q: D (75%), R: D (80%), S: D (63%), T: N (72%), W: D (80%), Y: D (63%), |
| Predicted by PROVEAN: | A: N, C: D, D: D, E: D, F: D, G: D, H: D, I: N, K: D, L: N, M: N, N: D, P: D, Q: D, R: D, S: D, T: N, W: D, Y: D, |
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[hide] Studies of human MDR1-MDR2 chimeras demonstrate th... Mol Cell Biol. 1999 Feb;19(2):1450-9. Zhou Y, Gottesman MM, Pastan I
Studies of human MDR1-MDR2 chimeras demonstrate the functional exchangeability of a major transmembrane segment of the multidrug transporter and phosphatidylcholine flippase.
Mol Cell Biol. 1999 Feb;19(2):1450-9., [PMID:9891078]
Abstract [show]
P-glycoprotein (P-gp), encoded by the MDR1 gene, is a plasma membrane transporter which effluxes a large number of structurally nonrelated hydrophobic compounds. The molecular basis of the broad substrate recognition of P-gp is not well understood. Despite the 78% amino acid sequence identity of the MDR1 and MDR2 transporter, MDR2, which has been identified as a phosphatidylcholine transporter, does not transport most MDR1 substrates. The structural and functional differences between MDR1 and MDR2 provide an opportunity to identify the residues essential for the broad substrate spectrum of MDR1. Using an approach involving exchanging homologous segments of MDR1 and MDR2 and site-directed mutagenesis, we have demonstrated that MDR1 residues Q330, V331, and L332 in transmembrane domain 6 are sufficient to allow an MDR2 backbone in the N-terminal half of P-gp to transport several MDR1 substrates, including bisantrene, colchicine, vinblastine, and rhodamine-123. These studies help define some residues important for multidrug transport and indicate the close functional relationship between the multidrug transporter (MDR1) and phosphatidylcholine flippase (MDR2).
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No. Sentence Comment
49 MDR1NAM was created by introducing mutations Q330N, V331A, and L332M into MDR1 by PCR mutagenesis.
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ABCB1 p.Val331Ala 9891078:49:52
status: NEW169 To confirm the significance of Q330, V331, and L332 in TM6, we generated a MDR1 mutant containing the mutations Q330N, V331A, and L332M.
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ABCB1 p.Val331Ala 9891078:169:119
status: NEW226 One is that the substitutions Q330N, V331A, and L332M in MDR1 significantly decreased overall MDR1 multidrug transporter activity.
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ABCB1 p.Val331Ala 9891078:226:37
status: NEW241 Thus, the triple mutation Q330N, V331A, and L332M appeared to have a synergistic effect.
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ABCB1 p.Val331Ala 9891078:241:33
status: NEW262 Since the UIC2 epitope depends on a particular P-gp conformation (24), the decreased level of UIC2 recognition may indicate a conformational change in MDR1 caused by mutations Q330N, V331A, and L332M.
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ABCB1 p.Val331Ala 9891078:262:183
status: NEW[hide] The extracellular loop between TM5 and TM6 of P-gl... Arch Biochem Biophys. 1999 Jul 1;367(1):74-80. Zhou Y, Gottesman MM, Pastan I
The extracellular loop between TM5 and TM6 of P-glycoprotein is required for reactivity with monoclonal antibody UIC2.
Arch Biochem Biophys. 1999 Jul 1;367(1):74-80., [PMID:10375401]
Abstract [show]
P-glycoprotein (P-gp), encoded by the MDR1 gene, is a plasma membrane transporter which confers resistance to many chemotherapeutic drugs. Monoclonal antibodies raised against P-gp have been used as tools to study P-gp topology and activity. Monoclonal antibody UIC2 recognizes a functional conformation of P-gp on the cell surface and blocks P-gp-mediated drug transport. Knowledge about the UIC2 epitope and the mechanism of its inhibitory effects may be helpful for understanding P-gp structure and developing P-gp inhibitors. In the present work, using several chimeras of MDR1 and MDR2, we found that the native sequence of the predicted extracellular loop between transmembrane domains (TM) 5 and 6 of P-gp is necessary, but not sufficient, for UIC2 reactivity. In addition, UIC2 reactivity is also affected by mutations in TM6, a region known to be involved in interactions of P-gp with substrates. These observations suggest that residues in the extracellular loop between TM5 and TM6 are directly involved in the display of the UIC2 epitope. Since TM6 has been shown to be actively involved in drug transport process, the proximity of this region to TM6 may help to explain why UIC2 binding is sensitive to the functional state of P-gp and why binding of UIC2 inhibits P-gp-mediated drug transport.
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No. Sentence Comment
66 Those substitutions are T318S, L322I, G324K, S327T, Q330N, V331A, and L332M (Fig. 1).
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ABCB1 p.Val331Ala 10375401:66:59
status: NEW79 This possibility was examined by testing the UIC2 reactivity of a P-gp mutant, MDR1/ MDR2(330-332), which carried substitutions Q330N, V331A, and L332M in TM6.
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ABCB1 p.Val331Ala 10375401:79:135
status: NEW84 However, in the presence of cyclosporin A, cells expressing MDR1/MDR2(330-332) displayed a number of UIC2 binding sites more similar to cells expressing wild-type P-gp, indicating that the triple mutation (Q330N, V331A, and L332M) in TM6 did not directly affect the affinity of UIC2 for its epitope (Fig. 4).
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ABCB1 p.Val331Ala 10375401:84:213
status: NEW115 Since the substitutions T318S, L322I, G324K, and S327T had little effect on either MRK-16 labeling or multidrug transporter activity, it is less likely that these mutations produce any major changes in P-gp structure.
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ABCB1 p.Val331Ala 10375401:115:75
status: NEW116 UIC2 Reactivity Is Sensitive to Mutations in TM6 A triple mutation, Q330N, V331A, and L332M, in TM6 decreased overall multidrug transporter activity FIG. 3. FACS analysis of cell surface expression of MDR1 and MDR1/MDR2(330-332) using MRK-16 and UIC2.
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ABCB1 p.Val331Ala 10375401:116:75
status: NEW78 This possibility was examined by testing the UIC2 reactivity of a P-gp mutant, MDR1/ MDR2(330-332), which carried substitutions Q330N, V331A, and L332M in TM6.
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ABCB1 p.Val331Ala 10375401:78:135
status: NEW83 However, in the presence of cyclosporin A, cells expressing MDR1/MDR2(330-332) displayed a number of UIC2 binding sites more similar to cells expressing wild-type P-gp, indicating that the triple mutation (Q330N, V331A, and L332M) in TM6 did not directly affect the affinity of UIC2 for its epitope (Fig. 4).
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ABCB1 p.Val331Ala 10375401:83:213
status: NEW