ABCMdb

ABCC7 p.Glu1104Arg

ClinVar:	c.3310G>T , p.Glu1104* D , Pathogenic
CF databases:	c.3310G>T , p.Glu1104* D , CF-causing c.3310G>A , p.Glu1104Lys (CFTR1) ? ,
Predicted by SNAP2:	A: D (53%), C: D (53%), D: N (82%), F: D (71%), G: D (71%), H: D (80%), I: D (63%), K: D (85%), L: D (71%), M: D (59%), N: D (66%), P: D (80%), Q: D (63%), R: D (85%), S: D (59%), T: D (53%), V: D (63%), W: D (85%), Y: D (75%),
Predicted by PROVEAN:	A: D, C: D, D: N, F: D, G: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: N, R: D, S: D, T: D, V: D, W: D, Y: D,

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Publications
[hide] Cystic fibrosis-associated mutations at arginine 3... J Biol Chem. 1999 Feb 26;274(9):5429-35. Cotten JF, Welsh MJ
Cystic fibrosis-associated mutations at arginine 347 alter the pore architecture of CFTR. Evidence for disruption of a salt bridge.
J Biol Chem. 1999 Feb 26;274(9):5429-35., 1999-02-26 [PMID:10026154]

Abstract [show]
Arginine 347 in the sixth transmembrane domain of cystic fibrosis transmembrane conductance regulator (CFTR) is a site of four cystic fibrosis-associated mutations. To better understand the function of Arg-347 and to learn how mutations at this site disrupt channel activity, we mutated Arg-347 to Asp, Cys, Glu, His, Leu, or Lys and examined single-channel function. Every Arg-347 mutation examined, except R347K, had a destabilizing effect on the pore, causing the channel to flutter between two conductance states. Chloride flow through the larger conductance state was similar to that of wild-type CFTR, suggesting that the residue at position 347 does not interact directly with permeating anions. We hypothesized that Arg-347 stabilizes the channel through an electrostatic interaction with an anionic residue in another transmembrane domain. To test this, we mutated anionic residues (Asp-924, Asp-993, and Glu-1104) to Arg in the context of either R347E or R347D mutations. Interestingly, the D924R mutation complemented R347D, yielding a channel that behaved like wild-type CFTR. These data suggest that Arg-347 plays an important structural role in CFTR, at least in part by forming a salt bridge with Asp-924; cystic fibrosis-associated mutations disrupt this interaction.

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Sentences [show]
No. Sentence Comment
154 We studied the conductance properties of the following double mutants: R347D/D924R, R347D/D993R, and R347E/E1104R. Login to comment
155 The R347D/D993R and R347E/E1104R mutants each had two conductance states with pHc-dependent behavior (Fig. 5). Login to comment
157 Accordingly, for R347D/D993R and R347E/E1104R the current variance in the open state increased with decreasing pHc (Fig. 5B). Login to comment
158 Qualitatively, the lifetimes of the OL and OB conductance states in the R347D/D993R and R347E/E1104R were similar to that of the R347D and R347E mutants, respectively. The amplitude of the OL state was larger for both of these double mutants as compared with the single mutants (Figs. Login to comment
160 We also observed an infrequent, additional small conductance state in the R347E/E1104 mutant (see amplitude histogram in Fig. 5A); this is likely due to the E1104R mutation itself. Login to comment
179 A, single-channel current tracings from excised, inside-out membrane patches containing R347E/E1104R, R347D/D924R, and R347D/D993R. Login to comment
181 B, current variance of R347E/E1104R, R347D/D924R, and R347D/D993R at the indicated pHc was collected as in Fig. 3. Login to comment
209 Consistent with this, mutation of D993R and E1104R in MSD2 increased the relative amplitude of the OL conductance state in the context of Arg-347 mutations. Login to comment

[hide] Mutations at arginine 352 alter the pore architect... J Membr Biol. 2008 Mar;222(2):91-106. Epub 2008 Apr 18. Cui G, Zhang ZR, O'Brien AR, Song B, McCarty NA
Mutations at arginine 352 alter the pore architecture of CFTR.
J Membr Biol. 2008 Mar;222(2):91-106. Epub 2008 Apr 18., [PMID:18421494]

Abstract [show]
Arginine 352 (R352) in the sixth transmembrane domain of the cystic fibrosis transmembrane conductance regulator (CFTR) previously was reported to form an anion/cation selectivity filter and to provide positive charge in the intracellular vestibule. However, mutations at this site have nonspecific effects, such as inducing susceptibility of endogenous cysteines to chemical modification. We hypothesized that R352 stabilizes channel structure and that charge-destroying mutations at this site disrupt pore architecture, with multiple consequences. We tested the effects of mutations at R352 on conductance, anion selectivity and block by the sulfonylurea drug glipizide, using recordings of wild-type and mutant channels. Charge-altering mutations at R352 destabilized the open state and altered both selectivity and block. In contrast, R352K-CFTR was similar to wild-type. Full conductance state amplitude was similar to that of wild-type CFTR in all mutants except R352E, suggesting that R352 does not itself form an anion coordination site. In an attempt to identify an acidic residue that may interact with R352, we found that permeation properties were similarly affected by charge-reversing mutations at D993. Wild-type-like properties were rescued in R352E/D993R-CFTR, suggesting that R352 and D993 in the wild-type channel may interact to stabilize pore architecture. Finally, R352A-CFTR was sensitive to modification by externally applied MTSEA+, while wild-type and R352E/D993R-CFTR were not. These data suggest that R352 plays an important structural role in CFTR, perhaps reflecting its involvement in forming a salt bridge with residue D993.

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Sentences [show]
No. Sentence Comment
162 Slope conductances are summarized in Table 1 Table 1 Slope conductancea (in pS) of the f state of WT-CFTR and multiple single and double mutants CFTR n Negative VM Positive VM WT 7 6.82 ± 0.03 6.97 ± 0.06 R352A 6 6.80 ± 0.06 7.85 ± 0.07*, ** R352Q 6 5.29 ± 0.02* 6.28 ± 0.05*, ** R352K 5 6.87 ± 0.03 6.86 ± 0.01 R352E 5 3.78 ± 0.01* 6.03 ± 0.01*, ** R352E/E873R 6 3.84 ± 0.01* 5.64 ± 0.01*, ** R352E/ E1104R 6 4.36 ± 0.01* 5.86 ± 0.02*, ** R352E/D993R 5 5.90 ± 0.02* 6.44 ± 0.01*, ** D993R 7 8.27 ± 0.05* 7.13 ± 0.07** a Slope conductance indicates single-channel conductance calculated from 0 to +100 mV (positive VM) or to -100 mV (negative VM) by linear regression * P B 0.001 compared to the equivalent slope conductance in WT-CFTR, ** P B 0.001 compared to the slope conductance in the same mutant at negative VM reflects the loss of anion binding properties within the core of the permeation pathway, which contributes to the tight binding of SCN (Smith et al. 1999). Login to comment
199 We studied the conductance properties of CFTR channels bearing the following mutations: R352E, R352E/E873R, R352E/ D993R and R352E/E1104R. Login to comment
201 Three of these mutants, R352E-, R352E/E873R- and R352E/E1104R-CFTR, exhibited instability of the open state, in which the amplitudes of the s1, s2 and f conductance states were very similar between the three mutants. Login to comment
204 R352E-, R352E/ E873R- and R352E/E1104R-CFTR exhibited significant outward rectification, while WT-CFTR did not (Table 1). Login to comment
208 If D993 served as the interaction partner of R352, we would expect that block of R352E/D993R-CFTR would be similar to that 0.4 pA 2 s 0.4 pA 2 s 0.2 pA 2 s 0.2 pA 2 s c s1 s2 f c s1 s2 f c s1 s2 f c f R352E R352E/E873R R352E/E1104R R352E/D993R 0 4000 #ofevents 0.0 -0.4 -0.8 0.0 -0.4 -0.8 3000 #ofevents 0 #ofevents 0.0 -0.4 -0.8 3000 0 Currents (pA) 0.0 -0.4 0 2500#ofevents -0.8 fc s1 s2 s1 s2 s1 s2 0.4 pA 2 s 0.4 pA 2 s 0.2 pA 2 s 0.2 pA 2 s c s1 s2 f c s1 s2 f c s1 s2 f c f R352E R352E/E873R R352E/E1104R R352E/D993R 0.4 pA 2 s 0.4 pA 2 s 0.4 pA 2 s 0.4 pA 2 s 0.2 pA 2 s 0.2 pA 2 s 0.2 pA 2 s 0.2 pA 2 s c s1 s2 f c s1 s2 f c s1 s2 f c f R352E R352E/E873R R352E/E1104R R352E/D993R B C D A 0 4000 #ofevents 0.0 -0.4 -0.8 0 4000 #ofevents 0.0 -0.4 -0.8 0.0 -0.4 -0.8 3000 #ofevents 0 0.0 -0.4 -0.8 3000 #ofevents 0 #ofevents 0.0 -0.4 -0.8 3000 0 Currents (pA) #ofevents 0.0 -0.4 -0.8 3000 0 #ofevents 0.0 -0.4 -0.8 3000 0 Currents (pA) 0.0 -0.4 0 2500#ofevents -0.8 fc s1 s2 s1 s2 s1 s2 Fig. 7 Single-channel current tracings of R352E-CFTR and double mutants from excised inside-out patches (left) and resulting all-points amplitude histograms (right) under the same experimental conditions as in Fig. 1. Login to comment
232 Hence, it is likely that MTSEA+ modified one (or more) of the endogenous cysteines, which B WT-CFTR R352E/D993R R352E R352E/E873R R352E/E1104R WT-CFTR R352E/D993R R352E R352E/E873R R352E/E1104R mV -100 -50 50 100 -0.8 -0.4 0.4 0.8 pA 100 ms 0.2 nA A WT-CFTR R352E/D993R R352E R352E/E873R R352E/E1104R WT-CFTR R352E/D993R R352E R352E/E873R R352E/E1104R mV -100 -50 50 100 -0.8 -0.4 0.4 0.8 pA WT-CFTR R352E/D993R R352E R352E/E873R R352E/E1104R WT-CFTR R352E/D993R R352E R352E/E873R R352E/E1104R WT-CFTR R352E/D993R R352E R352E/E873R R352E/E1104R WT-CFTR R352E/D993R R352E R352E/E873R R352E/E1104R mV -100 -50 50 100 -0.8 -0.4 0.4 0.8 pA 100 ms 0.2 nA Fig. 8 The double mutant R352E/D993R-CFTR recovers WT-like channel behavior. Login to comment
233 (A) Single channel i-V relationships are shown for full conductance states of WT-, R352E-, R352E/E873R-, R352E/ E1104R- and R352E/D993R-CFTR. Login to comment
258 Second, we identified the interaction partner as D993 by use of double mutants; R352E/E873R-CFTR and R352E/E1104R-CFTR exhibited permeation properties similar to those of R352E-CFTR, while R352E/D993R-CFTR behaved more like WT-CFTR. Login to comment

[hide] Evolutionary and functional divergence between the... Proc Natl Acad Sci U S A. 2008 Dec 2;105(48):18865-70. Epub 2008 Nov 19. Jordan IK, Kota KC, Cui G, Thompson CH, McCarty NA
Evolutionary and functional divergence between the cystic fibrosis transmembrane conductance regulator and related ATP-binding cassette transporters.
Proc Natl Acad Sci U S A. 2008 Dec 2;105(48):18865-70. Epub 2008 Nov 19., 2008-12-02 [PMID:19020075]

Abstract [show]
The cystic fibrosis transmembrane conductance regulator (CFTR) is a member of the ATP-binding cassette (ABC) transporter superfamily, an ancient family of proteins found in all phyla. In nearly all cases, ABC proteins are transporters that couple the hydrolysis of ATP to the transmembrane movement of substrate via an alternating access mechanism. In contrast, CFTR is best known for its activity as an ATP-dependent chloride channel. We asked why CFTR, which shares the domain architecture of ABC proteins that function as transporters, exhibits functional divergence. We compared CFTR protein sequences to those of other ABC transporters, which identified the ABCC4 proteins as the closest mammalian paralogs, and used statistical analysis of the CFTR-ABCC4 multiple sequence alignment to identify the specific domains and residues most likely to be involved in the evolutionary transition from transporter to channel activity. Among the residues identified as being involved in CFTR functional divergence, by virtue of being both CFTR-specific and conserved among all CFTR orthologs, was R352 in the sixth transmembrane helix (TM6). Patch-clamp experiments show that R352 interacts with D993 in TM9 to stabilize the open-channel state; D993 is absolutely conserved between CFTRs and ABCC4s. These data suggest that CFTR channel activity evolved, at least in part, by converting the conformational changes associated with binding and hydrolysis of ATP, as are found in true ABC Transporters, into an open permeation pathway by means of intraprotein interactions that stabilize the open state. This analysis sets the stage for understanding the evolutionary and functional relationships that make CFTR a unique ABC transporter protein.

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Sentences [show]
No. Sentence Comment
95 Isolated bursts of channel activity from oocytes expressing WT-CFTR, R352E-CFTR, R352E/E1104R-CFTR, and R352E/D993R-CFTR. Login to comment
99 Channels bearing the R352E mutation, or the double mutant R352E/E1104R, exhibited frequent transitions to subconductance levels. Login to comment