ABCB4 p.Tyr118Cys
| Predicted by SNAP2: | A: N (57%), C: N (66%), D: D (63%), E: D (63%), F: N (78%), G: N (61%), H: N (61%), I: N (82%), K: D (63%), L: N (82%), M: N (78%), N: N (53%), P: D (71%), Q: N (53%), R: D (59%), S: N (66%), T: N (61%), V: N (78%), W: N (66%), |
| Predicted by PROVEAN: | A: D, C: D, D: D, E: D, F: N, G: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: D, T: D, V: D, W: D, |
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[hide] Identification of residues within the drug-binding... J Biol Chem. 2000 Dec 15;275(50):39272-8. Loo TW, Clarke DM
Identification of residues within the drug-binding domain of the human multidrug resistance P-glycoprotein by cysteine-scanning mutagenesis and reaction with dibromobimane.
J Biol Chem. 2000 Dec 15;275(50):39272-8., [PMID:11013259]
Abstract [show]
P-glycoprotein (P-gp) can transport a wide variety of cytotoxic compounds that have diverse structures. Therefore, the drug-binding domain of the human multidrug resistance P-gp likely consists of residues from multiple transmembrane (TM) segments. In this study, we completed cysteine-scanning mutagenesis of all the predicted TM segments of P-gp (TMs 1-5 and 7-10) and tested for inhibition by a thiol-reactive substrate (dibromobimane) to identify residues within the drug-binding domain. The activities of 189 mutants were analyzed. Verapamil-stimulated ATPase activities of seven mutants (Y118C and V125C (TM2), S222C (TM4), I306C (TM5), S766C (TM9), and I868C and G872C (TM10)) were inhibited by more than 50% by dibromobimane. The activities of mutants S222C (TM4), I306C (TM5), I868C (TM10), and G872C (TM10), but not that of mutants Y118C (TM2), V125C (TM2), and S776C (TM9), were protected from inhibition by dibromobimane by pretreatment with verapamil, vinblastine, or colchicine. These results and those from previous studies (Loo, T. W. and Clarke, D. M. (1997) J. Biol. Chem. 272, 31945-31948; Loo, T. W. and Clarke, D. M. (1999) J. Biol. Chem. 274, 35388-35392) indicate that the drug-binding domain of P-gp consists of residues in TMs 4, 5, 6, 10, 11, and 12.
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No. Sentence Comment
183 DBBn also inhibited P-gp by reacting with cysteines in TM2 (Y118C and V125C) and TM8 (S766C).
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ABCB4 p.Tyr118Cys 11013259:183:60
status: NEW185 It is possible that some of these residues do not lie within the drug-binding site for these compounds because relatively high concentrations of dBBn were needed to inhibit 50% of the activity of mutants Y118C (TM2) and V125C (TM2) (740 and 870 M, respectively).
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ABCB4 p.Tyr118Cys 11013259:185:204
status: NEW186 Another possibility is that modification of mutant Y118C, V125C, or S766C blocks an essential conformational change during coupling of drug binding to ATPase activity.
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ABCB4 p.Tyr118Cys 11013259:186:51
status: NEW