ABCMdb

ABCC7 p.Ala1067Cys

ClinVar:	c.3200C>T , p.Ala1067Val ? , not provided c.3199G>C , p.Ala1067Pro ? , not provided c.3200C>G , p.Ala1067Gly ? , not provided c.3200C>A , p.Ala1067Asp ? , not provided c.3199G>A , p.Ala1067Thr ? , Conflicting interpretations of pathogenicity, not provided
CF databases:	c.3200C>A , p.Ala1067Asp (CFTR1) D , This substitution involves a residue conserved among species, located in an intracellular loop, and affects the charge of the CFTR protein. It was found at the homozygous state in a patient originating from India, and having a classical severe form of CF. A1067D creates a MaeIII restriction site. c.3199G>C , p.Ala1067Pro (CFTR1) D , The mutation was detected by DHPLC analysis and characterized by direct seqencing c.3199G>A , p.Ala1067Thr (CFTR1) ? , This child is 2 years old, carries the [delta]F508 mutation on the other chromosome, and is at this time a mild form of the disease. c.3200C>G , p.Ala1067Gly (CFTR1) ? , This change has been detected by DGGE analysis and direct sequencing in one Spanish allele c.3200C>T , p.Ala1067Val (CFTR1) ? , Ala to Val at 1067
Predicted by SNAP2:	C: D (91%), D: D (95%), E: D (95%), F: D (95%), G: D (95%), H: D (95%), I: D (95%), K: D (95%), L: D (95%), M: D (95%), N: D (95%), P: D (95%), Q: D (95%), R: D (95%), S: D (91%), T: N (87%), V: D (71%), W: D (95%), Y: D (95%),
Predicted by PROVEAN:	C: D, D: D, E: D, F: D, G: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: N, T: D, V: D, W: D, Y: D,

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Publications
[hide] A small-molecule modulator interacts directly with... Mol Pharmacol. 2009 Jun;75(6):1430-8. Epub 2009 Apr 1. Wellhauser L, Kim Chiaw P, Pasyk S, Li C, Ramjeesingh M, Bear CE
A small-molecule modulator interacts directly with deltaPhe508-CFTR to modify its ATPase activity and conformational stability.
Mol Pharmacol. 2009 Jun;75(6):1430-8. Epub 2009 Apr 1., [PMID:19339490]

Abstract [show]
The deletion of Phe-508 (DeltaPhe508) constitutes the most prevalent of a number of mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) that cause cystic fibrosis (CF). This mutation leads to CFTR misfolding and retention in the endoplasmic reticulum, as well as impaired channel activity. The biosynthetic defect can be partially overcome by small-molecule "correctors"; once at the cell surface, small-molecule "potentiators" enhance the channel activity of DeltaPhe508-CFTR. Certain compounds, such as VRT-532, exhibit both corrector and potentiator functions. In the current studies, we confirmed that the inherent chloride channel activity of DeltaPhe508-CFTR (after biosynthetic rescue) is potentiated in studies of intact cells and membrane vesicles. It is noteworthy that we showed that the ATPase activity of the purified and reconstituted mutant protein is directly modulated by binding of VRT-532 [4-methyl-2-(5-phenyl-1H-pyrazol-3-yl)-phenol] ATP turnover by reconstituted DeltaPhe508-CFTR is decreased by VRT-532 treatment, an effect that may account for the increase in channel open time induced by this compound. To determine whether the modification of DeltaPhe508-CFTR function caused by direct VRT-532 binding is associated with structural changes, we evaluated the effect of VRT-532 binding on the protease susceptibility of the major mutant. We found that binding of VRT-532 to DeltaPhe508-CFTR led to a minor but significant decrease in the trypsin susceptibility of the full-length mutant protein and a fragment encompassing the second half of the protein. These findings suggest that direct binding of this small molecule induces and/or stabilizes a structure that promotes the channel open state and may underlie its efficacy as a corrector of DeltaPhe508-CFTR.

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208 It is noteworthy that Loo et al. (2008) and Serohijos et al. (2008) showed that deletion of Phe508 impairs the chemical cross-linking that normally occurs between a non-native cysteine residue incorporated into NBD1, in the proximity of Phe508 (V510C), and a non-native cysteine introduced into the fourth intracellular loop (A1067C) Fig. 5. Login to comment

[hide] Correctors enhance maturation of DeltaF508 CFTR by... Biochemistry. 2009 Oct 20;48(41):9882-90. Loo TW, Bartlett MC, Clarke DM
Correctors enhance maturation of DeltaF508 CFTR by promoting interactions between the two halves of the molecule.
Biochemistry. 2009 Oct 20;48(41):9882-90., 2009-10-20 [PMID:19761259]

Abstract [show]
Deletion of Phe508 in cystic fibrosis transmembrane conductance regulator (DeltaF508 CFTR) causes cystic fibrosis. CFTR consists of two homologous halves with each containing a nucleotide-binding domain (NBD) and a transmembrane domain (TMD). DeltaF508 CFTR appears to be trapped in an incompletely folded state. Small molecules (correctors) promote folding of DeltaF508 CFTR with relatively low efficiency. Understanding the mechanism of repair may lead to the development of more effective correctors. Here we tested the effect of correctors and the DeltaF508 mutation on interactions between the halves of CFTR when expressed as separate polypeptides. Glycosylation of C-half CFTR was defective when expressed alone as a mixture of core and unglycosylated proteins was detected. Coexpression of C-half CFTR with either wild-type N-half or DeltaF508/N-half CFTR, however, increased the amount of core-glycosylated protein, but only coexpression with wild-type N-half promoted maturation of C-half CFTR (Endo H resistant). This suggested that the DeltaF508 mutation inhibited some interactions between N-half and C-half CFTRs. Interaction of A52-tagged wild-type N-half or DeltaF508/N-half CFTR with histidine-tagged C-half CFTR was then followed by nickel-chelate chromatography. Coexpression of A52-tagged wild-type N-half or DeltaF508/N-half CFTR with histidine-tagged C-half CFTR resulted in the wild-type N-half CFTR but not DeltaF508/N-half CFTR protein being retained on the column. Coexpression of DeltaF508/N-half and C-half CFTR in the presence correctors VX-325 and corr-4a, however, restored interactions between the two halves. An interaction that was restored was that between NBD1 and TMD2 as the correctors restored cross-linking of mutant DeltaF508/NBD1(V510C)/TMD2(A1067C). Therefore, correctors promote proper interactions between the two halves of CFTR.

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11 Coexpression of ΔF508/N-half and C-half CFTR in the presence correctors VX-325 and corr-4a, however, restored interactions between the two halves. An interaction that was restored was that between NBD1 and TMD2 as the correctors restored cross-linking of mutant ΔF508/NBD1- (V510C)/TMD2(A1067C). Login to comment
78 For example, mature CFTR shows cross-linking between cysteines in TM segment 6 (N-half) and TM segment 12 (C-half) or between cysteines in NBD1 [N-half (V510C)] and ICL4 [C-half (A1067C)], but the immature form of ΔF508 does not (17, 25). Login to comment
101 The black dot indicates the predicted location of an introduced cysteine (A1067C) in TMD2 that will directly cross-link to a cysteine introduced into NBD1 (V510C). Login to comment
174 To test if correctors VX-325 and corr-4a promoted NBD1-TMD2 interactions in ΔF508 CFTR, mutant ΔF508/V510C(NBD1)/A1067C- (TMD2) was expressed in the presence or absence of the correctors. Login to comment
182 Expression of ΔF508/V510C(NBD1)/A1067C- (TMD2) in the presence of correctors, however, yielded mature CFTR (180 kDa protein) (Figure 5, right). Login to comment

[hide] Benzbromarone stabilizes DeltaF508 CFTR at the cel... Biochemistry. 2011 May 31;50(21):4393-5. Epub 2011 May 3. Loo TW, Bartlett MC, Clarke DM
Benzbromarone stabilizes DeltaF508 CFTR at the cell surface.
Biochemistry. 2011 May 31;50(21):4393-5. Epub 2011 May 3., 2011-05-31 [PMID:21520952]

Abstract [show]
Deletion of Phe508 from the first nucleotide-binding domain of the CFTR chloride channel causes cystic fibrosis because it inhibits protein folding. Indirect approaches such as incubation at low temperatures can partially rescue DeltaF508 CFTR, but the protein is unstable at the cell surface. Here, we show that direct binding of benzbromarone to the transmembrane domains promoted maturation and stabilized DeltaF508 CFTR because its half-life at the cell surface was ~10-fold longer than that for low-temperature rescue. Therefore, a search for small molecules that can rescue and stabilize DeltaF508 CFTR could lead to the development of an effective therapy for cystic fibrosis.

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50 (C) Effect of benzbromarone on cross-linking (X-link) between cysteines in TMD1 and TMD2 (M348C/T1142C) or NBD1 and TMD2 (V510C/A1067C).7 (D) Immunoblot of cells expressing CFTR TMD1þ2 in the absence (À) or presence (þ) of 0.05 mM benzbromarone. Login to comment

[hide] Repair of CFTR folding defects with correctors tha... Methods Mol Biol. 2011;741:23-37. Loo TW, Clarke DM
Repair of CFTR folding defects with correctors that function as pharmacological chaperones.
Methods Mol Biol. 2011;741:23-37., [PMID:21594776]

Abstract [show]
The major cause of cystic fibrosis is the presence of processing mutations in CFTR (such as deletion of Phe-508 (F508del-CFTR)) that disrupt folding of the protein and trafficking to the cell surface. Processing mutations appear to inhibit folding of CFTR so that it accumulates in the endoplasmic reticulum as a partially folded protein. Expressing the proteins in the presence of small molecules called correctors can repair CFTR folding defects. Some correctors appear to function as pharmacological chaperones that specifically bind to the CFTR processing mutants and induce them to complete the folding process. In this chapter, we describe techniques to examine the effects of correctors on folding of CFTR processing mutants.

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No. Sentence Comment
252 The following protocols describe disulfide cross-linking between cysteines introduced into NBD1 (V510C) and the fourth intracellular loop in TMD2 (A1067C) of a F508del-CFTR processing mutant expressed in the presence or the absence of correctors. Login to comment
263 The next day, add 0.17 mg of (V510C)/(A1067C)/ F508del-CFTR cDNA to sterile water to give a total volume of 7.65 ml. Login to comment