ABCB1 p.Lys1264Ala
| Predicted by SNAP2: | A: D (63%), C: N (61%), D: D (71%), E: N (66%), F: D (71%), G: N (72%), H: N (57%), I: D (63%), L: D (66%), M: D (63%), N: N (82%), P: D (63%), Q: N (78%), R: N (87%), S: D (53%), T: D (59%), V: D (66%), W: D (75%), Y: D (71%), |
| Predicted by PROVEAN: | A: D, C: D, D: D, E: N, F: D, G: D, H: D, I: D, L: D, M: D, N: D, P: D, Q: N, R: N, S: D, T: D, V: D, W: D, Y: D, |
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[hide] The dileucine motif at the COOH terminus of human ... J Biol Chem. 2005 Jan 28;280(4):2522-8. Epub 2004 Nov 12. Loo TW, Bartlett MC, Clarke DM
The dileucine motif at the COOH terminus of human multidrug resistance P-glycoprotein is important for folding but not activity.
J Biol Chem. 2005 Jan 28;280(4):2522-8. Epub 2004 Nov 12., 2005-01-28 [PMID:15542593]
Abstract [show]
P-glycoprotein (P-gp, ABCB1) actively transports a broad range of cytotoxic compounds out of the cell. The COOH terminus of P-gp contains a dileucine motif (Leu(1260)-Leu(1261)) and a conserved phenylalanine (Phe(1268)). Similar residues in SUR1 (ABCC8) were reported to be important plasma membrane-targeting signals (Sharma, N., Crane, A., Clement, J. P. t., Gonzalez, G., Babenko, A. P., Bryan, J., and Aguilar-Bryan, L. (1999) J. Biol. Chem. 274, 20628-20632). Here, we used alanine-scanning mutagenesis to test whether these residues were essential for trafficking of P-gp to the cell surface. Mutant L1260A expressed a 150-kDa immature protein that did not reach the cell surface and was sensitive to digestion by Endo H(f). By contrast, mutants L1261A, F1268A, and wild-type P-gps expressed the 170-kDa mature proteins at the cell surface. Mutation of Leu(1260) to Gly, Ile, Trp, Lys, or Glu also resulted in the expression of the 150-kDa immature protein. All of the mutants, however, expressed the 170-kDa protein in the presence of the drug substrate/specific chemical chaperone cyclosporin A. Mutant L1260A P-gp exhibited drug-stimulated ATPase activities similar to that of wild-type enzyme after rescue with cyclosporin A. Deletion of the last 22 amino acids (Q(1259)-Q(1280)) also caused misprocessing. The mutant, however, was rescued by expression in the presence of cyclosporin A and conferred resistance to colchicine in transfected cells. These results show that the dileucine motif is not a plasma membrane targeting signal. The COOH terminus is required for proper folding of P-gp but not for activity.
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No. Sentence Comment
90 By contrast, the major product in wild-type P-gp and in mutants L1261A, K1264A, and F1268A was the 170-kDa protein.
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ABCB1 p.Lys1264Ala 15542593:90:72
status: NEW106 Accordingly, wild-type P-gp and mutants L1260A, L1261A, K1264A, Y1267A, and F1268A were expressed in HEK 293 cells in the presence or absence of the chemical chaperone/drug substrate cyclosporin A. Whole cell SDS extracts were then subjected to immunoblot analysis. Fig. 2A shows that the cyclosporin A was able to rescue mutants L1260A and Y1267A, because a 170-kDa protein was then detected (Fig. 2A).
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ABCB1 p.Lys1264Ala 15542593:106:56
status: NEW