ABCC7 p.Glu217Cys
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PMID: 24086355
[PubMed]
Rahman KS et al: "Modeling the conformational changes underlying channel opening in CFTR."
No.
Sentence
Comment
144
To provide experimental confirmation of the closed and open structures, and the transitions between the two, we asked whether cysteines engineered at these two positions in the double-mutant, R334C/E217C-CFTR, could be functionally crosslinked.
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ABCC7 p.Glu217Cys 24086355:144:198
status: NEW146 The traces in Figure 7 show that R334C/E217C-CFTR can repeatedly be activated by stimulation of the co-expressed beta2-adrenergic receptor using isoproterenol, without substantial decrement in peak current prior to exposure to the crosslinker MTS-2-MTS.
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ABCC7 p.Glu217Cys 24086355:146:39
status: NEW147 When the same cell was exposed to MTS-2-MTS in the absence of isoproterenol (Figure 7B), when most of the channels should be closed, subsequent exposure to isoproterenol failed to activate CFTR channels to the same degree as prior to MTS-2-MTS; these results are consistent with the notion that R334C and E217C are positioned very near each other in the channel closed state.
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ABCC7 p.Glu217Cys 24086355:147:305
status: NEW152 In control experiments, exposure to MTS-2-MTS did not have similar effects on the single mutants R334C-CFTR and E217C-CFTR, with respect to the ability to re-open channels after MTS-2-MTS exposure (Figure S6).
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ABCC7 p.Glu217Cys 24086355:152:112
status: NEW154 Inspection of the C0- and O-CFTR models indicates that the side chains of R334C and E217C are, indeed, close enough to be crosslinked by MTS-2-MTS only in the closed state (Figure S8).
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ABCC7 p.Glu217Cys 24086355:154:84
status: NEW173 Crosslinking R334C to E217C locks CFTR channels into the closed state.
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ABCC7 p.Glu217Cys 24086355:173:22
status: NEW176 B: Representative trace (left) and summary data (right) for macroscopic currents measured from R334C/E217C-CFTR with addition of the crosslinker MTS-2-MTS in the absence of isoproterenol (ISO), used to activate channels.
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ABCC7 p.Glu217Cys 24086355:176:101
status: NEW178 C: Representative trace and summary data for currents measured from R334C/E217C-CFTR with the crosslinker MTS-2-MTS added in the continuing presence of isoproterenol.
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ABCC7 p.Glu217Cys 24086355:178:74
status: NEW266 (TIF) Figure S6 Effects of 1 mM MTS2-2MTS on R334C-CFTR and E217C-CFTR channels. Representative traces (left) and summary data (right) for macroscopic currents measured from R334C- (A) and E217C-CFTR (B) by two-electrode voltage clamp.
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ABCC7 p.Glu217Cys 24086355:266:60
status: NEWX
ABCC7 p.Glu217Cys 24086355:266:189
status: NEW270 MTS-2-MTS also appeared to covalently decrease macroscopic current at R334C-CFTR, but not E217C-CFTR, perhaps due to alteration of charge or side-chain volume.
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ABCC7 p.Glu217Cys 24086355:270:90
status: NEW275 (TIF) Figure S7 Effects of MTSET (ET+) and MTSES (ES-) on R334C/E217C-CFTR channels. Representative traces (left) and summary data (right) for macroscopic currents measured from R334C/E217C-CFTR by two-electrode voltage clamp with addition of the 1 mM monofunctional MTS reagents MTSET+ (ET+ ) or MTSES2 (ES2 ) in the presence of isoproterenol (ISO).
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ABCC7 p.Glu217Cys 24086355:275:64
status: NEWX
ABCC7 p.Glu217Cys 24086355:275:184
status: NEW277 Both ET+ and ES2 covalently bound to R334C/E217C-CFTR.
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ABCC7 p.Glu217Cys 24086355:277:43
status: NEW284 (TIF) Figure S8 Distances between residues in R334C-E217C Double Mutant.
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ABCC7 p.Glu217Cys 24086355:284:52
status: NEW
PMID: 25024266
[PubMed]
Cui G et al: "Three charged amino acids in extracellular loop 1 are involved in maintaining the outer pore architecture of CFTR."
No.
Sentence
Comment
321
This is in contrast to the ability of MTS-2-MTS to lock R352C/D993C-CFTR into the open state or to lock R334C/E217C-CFTR into the closed state (Cui et al., 2013; Rahman et al., 2013).
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ABCC7 p.Glu217Cys 25024266:321:110
status: NEW