ABCB1 p.Glu256Arg
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PMID: 23634976
[PubMed]
Loo TW et al: "A salt bridge in intracellular loop 2 is essential for folding of human p-glycoprotein."
No.
Sentence
Comment
33
If a salt bridge between Glu256 and Arg276 were important for the maturation of human P-gp, it would be expected that an E256R/R276E mutant would still mature.
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ABCB1 p.Glu256Arg 23634976:33:121
status: NEW34 Accordingly, we reversed the positions of the glutamic acid and arginine residues (E256R/R276E) and tested the mutant`s ability to undergo maturation.
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ABCB1 p.Glu256Arg 23634976:34:83
status: NEW35 The mature 170 kDa P-gp protein was the major product in mutant E256R/R276E (Figure 2A).
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ABCB1 p.Glu256Arg 23634976:35:64
status: NEW37 The E256R or R276E mutation, however, inhibited maturation of P-gp (Figure 2A).
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ABCB1 p.Glu256Arg 23634976:37:4
status: NEW38 For example, Figure 2B shows that the 150 kDa E256R protein was sensitive to treatment with endoglycosidase H or PNGase F. Similar results were obtained when mutant R276E was treated with endoglycosidase H or PNGase F (data not shown).
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ABCB1 p.Glu256Arg 23634976:38:46
status: NEW40 To test if mutant E256R/R276E yielded an active protein, the histidine-tagged mutant was expressed in HEK293 cells, isolated by nickel chelate chromatography, and assayed for verapamil-stimulated ATPase activity.
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ABCB1 p.Glu256Arg 23634976:40:18
status: NEW41 Verapamil was used because it is transported by P-gp14 and strongly stimulates (>10-fold) the ATPase activity of human P-gp.15 The E256R/ R276E mutant showed verapamil-stimulated ATPase activity that was similar to that of the wild-type protein (Figure 2C).
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ABCB1 p.Glu256Arg 23634976:41:131
status: NEW61 (A) Whole cell sodium dodecyl sulfate (SDS) extracts of wild-type P-gp and mutants E256R, R276E, and E256R/R276E.
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ABCB1 p.Glu256Arg 23634976:61:83
status: NEWX
ABCB1 p.Glu256Arg 23634976:61:101
status: NEW