ABCB1 p.Val53Arg
| Predicted by SNAP2: | A: N (87%), C: N (97%), D: D (66%), E: D (59%), F: N (93%), G: N (82%), H: N (72%), I: N (97%), K: D (63%), L: N (97%), M: N (93%), N: D (59%), P: D (66%), Q: D (59%), R: D (59%), S: N (82%), T: N (87%), W: D (63%), Y: N (72%), |
| Predicted by PROVEAN: | A: N, C: N, D: D, E: D, F: N, G: D, H: D, I: N, K: D, L: N, M: N, N: D, P: D, Q: D, R: D, S: N, T: N, W: D, Y: N, |
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[hide] Arginines in the first transmembrane segment promo... J Biol Chem. 2008 Sep 5;283(36):24860-70. Epub 2008 Jul 2. Loo TW, Bartlett MC, Clarke DM
Arginines in the first transmembrane segment promote maturation of a P-glycoprotein processing mutant by hydrogen bond interactions with tyrosines in transmembrane segment 11.
J Biol Chem. 2008 Sep 5;283(36):24860-70. Epub 2008 Jul 2., 2008-09-05 [PMID:18596043]
Abstract [show]
A key goal is to correct defective folding of mutant ATP binding cassette (ABC) transporters, as they cause diseases such as cystic fibrosis. P-glycoprotein (ABCB1) is a useful model system because introduction of an arginine at position 65 of the first transmembrane (TM) segment could repair folding defects. To determine the mechanism of arginine rescue, we first tested the effects of introducing arginines at other positions in TM1 (residues 52-72) of a P-glycoprotein processing mutant (G251V) that is defective in folding and trafficking to the cell surface (20% maturation efficiency). We found that arginines introduced into one face of the TM1 helix (positions 52, 55, 56, 59, 60, 62, 63, 66, and 67) inhibited maturation, whereas arginines on the opposite face of the helix promoted (positions 64, 65, 68, and 71) or had little effect (positions 61, and 69) on maturation. Arginines at positions 61, 64, 65, and 68 appeared to lie close to the drug binding sites as they reduced the apparent affinity for drug substrates such as vinblastine and verapamil. Therefore, arginines that promoted maturation may face an aqueous drug translocation pathway, whereas those that inhibited maturation may face the lipid bilayer. The highest maturation efficiencies (60-85%) were observed with the Arg-65 and Arg-68 mutants. Mutations that removed hydrogen bond acceptors (Y950F/Y950A or Y953F/Y953A) in TM11 predicted to lie close to Arg-65 or Arg-68 inhibited maturation but did not affect maturation of the G251V parent. Therefore, arginine may rescue defective folding by promoting packing of the TM segments through hydrogen bond interactions.
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No. Sentence Comment
118 Mutants G251V, V53R/G251V (inhibited maturation) and M68R/G251V (enhanced maturation) were subjected endoglycosidase H or F digestion to determine whether the alterations in mobility of the mutant P-gps were indeed due to changes in the glycosylation state of the protein.
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ABCB1 p.Val53Arg 18596043:118:15
status: NEW138 Whole cell SDS extracts of HEK 293 cells expressing A52-tagged mutants G251V, V53R/G251V, or M68R/G251V were treated with endoglycosidase Hf (H),peptideN-glycosidaseF(F)ornoendoglycosidase(-).Equivalentamountsweresubjectedtoimmunoblot analysis.
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ABCB1 p.Val53Arg 18596043:138:78
status: NEW140 V53R, Fig. 2A) were not grossly misfolded, then it should be possible to promote their maturation by carrying out expression in the presence of drug substrates.
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ABCB1 p.Val53Arg 18596043:140:0
status: NEW151 By contrast, 12 of the mutants (V53R, G54R, T55R, L56R, A57R, A58R, I59R, I60R, G62R, A63R, P66R, and F72R) showed little or no rescue in the presence of drug substrates.
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ABCB1 p.Val53Arg 18596043:151:32
status: NEW165 TABLE 1 Effects of drug substrates on the maturation of TM1 arginine mutants containing the G251V mutation Mutation (G251V ؉) No drug Cyclosporin A Verapamil Vinblastine Rhodamine None - 111a,c 111 111 11 V52R -a,b 111 111 111 11 V53R 2 - - - - G64R 2 - - - - T55R 2 - - - - L56R 2 - - - - A57R 2 - - - - A58R 2 - - - - I59R 2 - - - - I60R 2 - - - - H61R - 111 1 1 1 G62R 2 - - - - A63R 2 - - - - G64R 1 111 1 1 11 L65R 11 111 11 11 11 P66R 2 1 - - - L67R - 111 111 111 11 M68R 111 111 111 111 111 M69R - 111 - 11 111 L70R - 111 111 111 11 V71R 1 111 111 111 11 F72R 2 1 1 1 1 a Change in the amount of mature (170 kDa) protein in the presence of drug substrate relative to that in the absence of drug substrate.
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ABCB1 p.Val53Arg 18596043:165:236
status: NEW