ABCMdb

ABCC7 p.Glu1401Lys

ClinVar:	c.4202A>C , p.Glu1401Ala ? , not provided c.4201G>T , p.Glu1401* ? , not provided c.4202A>G , p.Glu1401Gly ? , not provided c.4201G>A , p.Glu1401Lys ? , not provided
CF databases:	c.4201G>A , p.Glu1401Lys (CFTR1) D , The new missense mutation in the exon 23 was detected by DGGE and identified by direct sequencing. The mutation was found in an isolated CBAVD patient, carrying [delta]F508 on the other allele. The mutation was not observed in 200 CFTR alleles from 100 healthy fertile males, 20 CF alleles from 10 CF patients and 90 chromosomes from 45 CBAVD patients. c.4202A>G , p.Glu1401Gly (CFTR1) D , The mutation was identified in a CBAVD patient who was also found to carry deltaF508. No segregation analysis was performed yet. No other CFTR mutation was found after extensive screening of the coding regions. Biochemical and phylogenic inspection of the amino-acid change suggest a deleterious effect of E1401G. c.4202A>C , p.Glu1401Ala (CFTR1) ? , This mutation was detected by multiplex DGGE and identified by direct sequencing. It was founf in a Czech CF pacient with lung disease. [delta]F508 was found on the other chromosome. The patient was referred by the department of Medical Genetics,niversity Hospital Hradec Kralove,Cz.
Predicted by SNAP2:	A: N (72%), C: D (66%), D: N (66%), F: D (85%), G: N (53%), H: D (80%), I: D (80%), K: D (63%), L: D (63%), M: D (75%), N: N (57%), P: D (66%), Q: N (53%), R: D (71%), S: N (61%), T: N (53%), V: N (61%), W: D (85%), Y: D (80%),
Predicted by PROVEAN:	A: N, C: N, D: N, F: D, G: N, H: N, I: N, K: N, L: N, M: N, N: N, P: N, Q: N, R: N, S: N, T: N, V: N, W: D, Y: N,

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[hide] Molecular characterization of the cystic fibrosis ... Genet Med. 2007 Mar;9(3):163-72. Grangeia A, Sa R, Carvalho F, Martin J, Girodon E, Silva J, Ferraz L, Barros A, Sousa M
Molecular characterization of the cystic fibrosis transmembrane conductance regulator gene in congenital absence of the vas deferens.
Genet Med. 2007 Mar;9(3):163-72., [PMID:17413420]

Abstract [show]
PURPOSE: Approximately 20% of patients with congenital absence of the vas deferens remain without two mutations identified. We applied a strategy of serial screening steps to 45 patients with congenital absence of the vas deferens and characterized cystic fibrosis transmembrane conductance regulator gene mutations in all cases. METHODS: DNA samples of 45 patients with congenital absence of the vas deferens were screened by successive different molecular genetics approaches. RESULTS: Initial screening for the 31 most frequent cystic fibrosis mutations, IVS8 poly(TG)m, poly(T)n, and M470V polymorphisms, identified 8 different mutations in 40 patients (88.9%). Extensive cystic fibrosis transmembrane conductance regulator gene analysis by denaturing gradient gel electrophoresis, denaturing high-performance liquid chromatography, and DNA sequencing detected 17 further mutations, of which three were novel. Cystic fibrosis transmembrane conductance regulator gene rearrangements were searched by semiquantitative fluorescent multiplex polymerase chain reaction, which detected a CFTRdele2,3 (21 kb) large deletion and confirmed two homozygous mutations. Overall, 42 patients (93.3%) had two mutations and 3 patients (6.7%) had one mutation detected. CONCLUSIONS: The present screening strategy allowed a higher mutation detection rate than previous studies, with at least one cystic fibrosis transmembrane conductance regulator gene mutation found in all patients with congenital absence of the vas deferens.

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89 E1401K was found in a 37-year-old patient who carried a deltaF508 mutation in the other chromosome. It leads to the substitution of a glutamic acid by a lysine in (NBD) 2 at its carboxy-terminal side (Tables 1 and 2). Login to comment
93 DeltaF508 was the second most common mutation, representing 21 (23.3%) of total alleles, followed by R334W (6, Table 1 CFTR gene mutations and polymorphisms in patients with congenital absence of the vas deferens Mutation Location Nucleotide alteration Effect Method 1 CFTRdele2,3 Exons 2-3 Deletion of exons 2 and 3 Frameshift QFM-PCR 2 R117H Exon 4 G¡A at 482 AA substitution 31 mutation panel 3 P205S Exon 6a C¡T at 745 AA substitution DGGE/dHPLC 4 L206W Exon 6a T¡G at 749 AA substitution DGGE/dHPLC 5 R258G Exon 6b A¡G at 904 AA substitution DGGE/dHPLC 6 R334W Exon 7 C¡T at 1132 AA substitution 31 mutation panel 7 T5 allele Intron 8 Deletion of 2T at 1342-12 to -6 Aberrant splicing DGGE/DNA sequencing 8 P439S Exon 9 C¡T at 1447 AA substitution DGGE/dHPLC 9 D443Ya Exon 9 G¡T at 1459 AA substitution DGGE/dHPLC 10 I507del Exon 10 Deletion of 3 bp at 1648-1653 AA deletion 31 mutation panel 11 DeltaF508 Exon 10 Deletion of 3 bp at 1652-1655 AA deletion 31 mutation panel 12 G542X Exon 11 G¡T at 1756 Truncation 31 mutation panel 13 V562I Exon 12 G¡A at 1816 AA substitution DGGE/dHPLC 14 G576Aa Exon 12 G¡C at 1859 Aberrant splicing DGGE/dHPLC 15 D614G Exon 13 A¡G at 1973 AA substitution DGGE/dHPLC 16 R688Ca Exon 13 C¡T at 2134 AA substitution DGGE/dHPLC 17 V754M Exon 13 G¡A at 2392 AA substitution DGGE/dHPLC 18 E831X Exon 14a G¡T at 2623 Truncation DGGE/dHPLC 19 3272-26AϾG Intron 17a A¡G at 3272-26 Aberrant splicing DGGE/dHPLC 20 2789ϩ5G¡A Intron 14b G¡A at 2789ϩ5 Aberrant splicing 31 mutation panel 21 V1108L Exon 17b G¡C at 3454 AA substitution DGGE/dHPLC 22 L1227S Exon 19 T¡C at 3812 AA substitution DGGE/dHPLC 23 S1235R Exon 19 T¡G at 3837 AA substitution DGGE/dHPLC 24 P1290S Exon 20 C¡T at 4000 AA substitution DGGE/dHPLC 25 N1303K Exon 21 C¡G at 4041 AA substitution 31 mutation panel 26 E1401K Exon 23 G¡A at 4333 AA substitution DGGE/dHPLC Polymorphisms 1 TG repeats Intron 8 9-13 copies at 1342-12 to -35 Sequence variation DGGE/DNA sequencing 2 M470V Exon 10 A or G at 1540 Sequence variation DNA sequencing 3 125G/C Exon 1 G¡C at 125 Sequence variation DGGE/dHPLC 4 1001ϩ11T/C Intron 6b C¡4T at 1001ϩ11 Sequence variation DGGE/dHPLC 5 1716G/A Exon 10 G¡A at 1716 Sequence variation DGGE/dHPLC 6 1899-136T/G Intron 12 T¡G at 1899-136 Sequence variation DGGE/dHPLC 7 T854T Exon 14a T¡G at 2694 Sequence variation DGGE/dHPLC 8 3601-65C/A Intron 18 C¡A at 3601-65 Sequence variation DGGE/dHPLC 9 4521G/A Exon 24 G¡A at 4521 Sequence variation DGGE/dHPLC QFM-PCR, semiquantitative fluorescent multiplex polymerase chain reaction; bp, base pair; DGGE, denaturing gradient gel electrophoresis; dHPLC, denaturing high-performance liquid chromatography. Login to comment
101 The missense M470V polymorphism was evaluated in all 45 pa- tientswithCAVD(Table2).TheallelicfrequencyoftheM470variant Table 2 CFTR genotypes identified in patients with congenital absence of the vas deferens CFTR mutation genotypes [(TG)mTn] genotype M470V Patients N % DeltaF508 (TG)10T9 (TG)12T5 M V 11 24.4 DeltaF508 (TG)10T9 (TG)11T5 M M 1 2.2 DeltaF508 R117H (TG)10T9 (TG)10T7 M M 2 4.4 G542X (TG)10T9 (TG)12T5 M V 2a 4.4 DeltaF508 R334W (TG)10T9 (TG)11T7 M V 1 2.2 DeltaF508 D443Y-G576A-R668C (TG)10T9 (TG)10T7 M M 1 2.2 DeltaF508 D614G (TG)10T9 (TG)11T7 M V 1 2.2 DeltaF508 E831X (TG)10T9 (TG)11T7 M V 1 2.2 DeltaF508 L1227S (TG)10T9 (TG)11T7 M M 1 2.2 DeltaF508 E1401K (TG)10T9 (TG)11T7 M V 1 2.2 I507del D614G (TG)11T7 (TG)10T7 M V 1 2.2 N1303K L206W (TG)10T9 (TG)9T9 M M 1 2.2 R117H P205S (TG)11T7 (TG)10T7 M V 1 2.2 R117H R334W (TG)10T7 (TG)11T7 M V 1 2.2 R334W P439S (TG)11T7 (TG)11T7 M V 1 2.2 R334W R334Wb (TG)11T7 (TG)11T7 V V 1 2.2 R334W V562I (TG)11T7 (TG)11T5 V M 1 2.2 D443Y-G576A-R668C 3272-26A¡G (TG)10T7 (TG)10T7 M M 1 2.2 G576A-R668C V754Mb (TG)10T7 (TG)11T7 M M 1 2.2 S1235R S1235Rb (TG)13T5 (TG)13T5 M M 1 2.2 2789ϩ5G¡A S1235Rb (TG)10T7 (TG)13T5 M M 1 2.2 3272-26A¡G P1290S (TG)11T7 (TG)10T7 M V 1 2.2 P205S (TG)11T7 (TG)12T5 V V 1 2.2 G576A-R668C b (TG)10T7 (TG)11T5 M M 1 2.2 V1108L b (TG)11T7 (TG)11T5 V M 1 2.2 N1303K (TG)10T9 (TG)12T5 M V 1 2.2 3272-26A¡G b (TG)10T7 (TG)12T5 M V 1 2.2 CFTRdele2,3 b (TG)11T7 (TG)13T5 V M 1 2.2 b (TG)11T5 (TG)12T5 M V 1 2.2 b (TG)13T5 (TG)12T5 M V 1 2.2 DeltaF508 - (TG)10T9 (TG)11T7 M V 1a 2.2 L206W -b (TG)9T9 (TG)11T7 M V 1 2.2 R258G -b (TG)11T7 (TG)11T7 V V 1 2.2 a CUAVD. Login to comment
110 Large Table 3 Allelic frequencies of CFTR mutations in patients with congenital absence of the vas deferens CBAVD CUAVD Total Patients 42 3 45 Alleles 84 6 90 Mutations N % N % N % 1 T5 allele 26a 31 2 33.3 28 31.1 2 DeltaF508 20 23.8 1 16.7 21 23.3 3 R334W 6a 7.1 0 0 6 6.7 4 R117H 4 4.8 0 0 4 4.4 5 G576A 4b 4.8 0 0 4 4.4 6 R688C 4b 4.8 0 0 4 4.4 7 S1235R 3a 3.6 0 0 3 3.3 8 3272-26A¡G 3 3.6 0 0 3 3.3 9 P205S 2 2.4 0 0 2 2.2 10 L206W 2 2.4 0 0 2 2.2 11 D443Y 2b 2.4 0 0 2 2.2 13 D614G 2 2.4 0 0 2 2.2 14 N1303K 2 2.4 0 0 2 2.2 12 G542X 0 0 2 33.3 2 2.2 15 R258G 1 1.2 0 0 1 1.1 16 P439S 1 1.2 0 0 1 1.1 17 I507del 1 1.2 0 0 1 1.1 18 V562I 1 1.2 0 0 1 1.1 19 V754M 1 1.2 0 0 1 1.1 20 E831X 1 1.2 0 0 1 1.1 21 2789ϩ5G¡A 1 1.2 0 0 1 1.1 22 V1108L 1 1.2 0 0 1 1.1 23 L1227S 1 1.2 0 0 1 1.1 24 P1290S 1 1.2 0 0 1 1.1 25 E1401K 1 1.2 0 0 1 1.1 26 CFTRdele2,3 1 1.2 0 0 1 1.1 CBAVD, congenital bilateral absence of the vas deferens; CUAVD, congenital unilateral absence of the vas deferens. Login to comment
140 Three novel missense mutations (E1401K, P439S, and V1108L), not detected in the general population or in patients with CF, were here first described in patients with CAVD. Login to comment
141 Although the pathogenicity of these mutations is still being assessed by expression and functional in vitro studies, the combination of the amino acid substitutions with other mutations (deltaF508/E1401K, R334W/P439S, V1108L/T5) might be responsible for the CBAVD phenotype. Login to comment
142 In fact, they occur in highly conserved regions of the CFTR protein, which share 100% amino acid sequence homology between species48 and affect the NBD1, NBD2, and transmembrane regions of the protein, which are known to regulate chloride conductance and permeability.49-51 P439S was previously reported in a child with CF with pancreatic insufficiency and mild lung disease, in association with the P439S/R688C genotype.52 The E1401K mutation occurs at a position in which other mutations, E1401X and E1401A, have been described in patients with CF with pancreatic insufficiency.8 Some difficulties in defining CF or CAVD-causing mutations were observed with some missense mutations.6,27 G576A and R668C have been found independently, in pairs, or combined with the D443Y mutation on the same chromosome in patients withaCF-relatedsyndrome.Inaccordancewithpreviousstudies, we expected that G576A and R668C were located in cis in two patients and combined with D443Y in the same chromosome in two patients.6,9,12 Although initially described as polymorphisms,27 they were later considered mild mutations associated with the CBAVD phenotype when combined in trans with the severedeltaF508mutation.53 However,ourpresentresultssuggest they might also cause the CAVD phenotype when associated with other mild CFTR mutations, because three of four patients carry- ingthesecomplexallelesharboredamildorverymildmutationin the other chromosome (D443Y-G576A-R668C/3272-26A¡G, Table 5 Comparative analysis of CFTR mutation allelic frequencies (%) in patients with congenital absence of the vas deferens Countries Patients T5 allele DeltaF508 R334W R117H References Argentina 36 NA 20.8 NA 5.6 43 Austria 22 NA 13.6 NA 9.1 44 Italy 12 8.3 29.2 NA 4.2 39 The Netherlands 21 9.5 19.0 NA 21.4 38 Germany 106 12.3 26.4 0.5 11.3 30 Greece 14 14.3 14.3 NA NA 32 France 800 16.3 21.8 NA 4.4 6 United States 92 17.9 21.2 NA 2.2 41 Canada 74 18.2 16.9 1.4 6.1 5 Turkey 51 19.6 2.9 NA NA 35 Brazil 17 20.6 11.7 NA 2.9 34 Spain 134 20.9 16.0 0.4 3.0 33 Iran 113 25.7 12.4 0.9 3.5 37 Egypt 16 43.7 6.2 NA NA 40 Taiwan 27 44.4 NA NA NA 42 Portugal 45 31.1 23.3 6.7 4.4 13, 36, PS NA, not available; PS, present study. Login to comment

[hide] Molecular and functional characterization of CBAVD... Cell Physiol Biochem. 2008;22(1-4):79-92. Epub 2008 Jul 25. Grangeia A, Barro-Soria R, Carvalho F, Damas AM, Mauricio AC, Kunzelmann K, Barros A, Sousa M
Molecular and functional characterization of CBAVD-causing mutations located in CFTR nucleotide-binding domains.
Cell Physiol Biochem. 2008;22(1-4):79-92. Epub 2008 Jul 25., [PMID:18769034]

Abstract [show]
BACKGROUND: About 98% of male affected with cystic fibrosis (CF [MIM 219700]) are infertile due to bilateral absence of vas deferens (CBAVD [MIM 277180]), which makes up 1-2 % of all cases with male infertility. A previous screening of the entire coding region of the cystic fibrosis transmembrane conductance regulator gene (CFTR [MIM 602421]) in CBAVD patients identified three novel mutations: P439S is located in the first nucleotide binding domain (NBD1) of CFTR, whereas P1290S and E1401K are located in NBD2. METHODS: We analysed the effects of these novel mutations on CFTR processing and chloride (Cl(-)) channel activity. RESULTS: Although maturation patterns were not affected, total amounts of mature P439S-CFTR and P1290S-CFTR were reduced. Confocal microscopy showed correct membrane localisation of E1401K-CFTR, whereas P439S-CFTR and P1290S-CFTR mutants were located mainly in the cytoplasm. Iodide influx assay and whole-cell patch clamp demonstrated significantly reduced cAMP-dependent anion conductances for all three mutants. CONCLUSION: Dysfunction of CFTR is caused by either defective CFTR trafficking (P439S and P1290S) or/and Cl- channel function (P1290S and E1401K). Thus reduced Cl- conductance caused by the three CFTR mutations affects normal development of vas deferens and leads to CBAVD, but the remaining function is sufficient to prevent other typical CF symptoms.

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2 A previous screening of the entire coding region of the cystic fibrosis transmembrane conductance regulator gene (CFTR [MIM 602421]) in CBAVD patients identified three novel mutations: P439S is located in the first nucleotide binding domain (NBD1) of CFTR, whereas P1290S and E1401K are located in NBD2. Login to comment
5 Confocal microscopy showed correct membrane localisation of E1401K-CFTR, whereas P439S-CFTR and P1290S-CFTR mutants were located mainly in the cytoplasm. Login to comment
7 Conclusion: Dysfunction of CFTR is caused by either defective CFTR trafficking (P439S and P1290S) or/and Cl-channel function (P1290S and E1401K). Login to comment
59 Plasmids, cell culture, transient transfection CFTR mutations (P439S, P1290S and E1401K) were generated (QuikChange site-directed mutagenesis kit, Stratagene, La Jolla, CA) in the eukaryotic expression vector pCMVCFTRNot6.2, according to the manufacturer`s instructions. Login to comment
112 Multiple alignments of CFTR amino acid sequences from different species and localization of NBD1, NBD2 and three novel CFTR mutations (P439S, P1290S, E1401K). Login to comment
123 E1401K (NBD2) was found in a 37 year old patient who carried the severe F508del mutation on the other chromosome. Login to comment
126 Biosynthesis of CFTR mutants Maturation patterns of wild-type (wt) CFTR and mutant CFTR (F508del-CFTR, P439S-CFTR, P1290S-CFTR and E1401K-CFTR) were compared by Western blotting. Login to comment
137 The mutants P439S-CFTR and P1290S-CFTR were mainly expressed in the cytoplasm with little membrane expression, while E1401K-CFTR was clearly present in the cell membrane, similar to wtCFTR (Fig. 3A-B). Login to comment
142 wtCFTR and E1401K-CFTR were found in the plasma membrane, whereas F508del-CFTR is trapped in the cytoplasm. Login to comment
159 Intracellular cAMP was enhanced by stimulation of the cells with 2 µM FSK and 100 µM IBMX, which largely increased the rate of I- influx in HEK293 cells expressing wtCFTR, while non-transfected cells or F508del-CFTR expressing cellsdidnotrespondtostimulation(Fig.4A-B).Expression of all three mutants P439S-CFTR, P1290S-CFTR and E1401K-CFTR allowed for cAMP-induced increase in I- influx, although the rate of I- influx was significantly lower than that for wtCFTR (Fig. 4A-B). Login to comment
160 Reduced I- influx seems to be in accordance with the reduced levels of protein expression for two mutants P439S-CFTR, P1290S-CFTR.Although E1401K did not interfere with Fig. 5. Login to comment
165 GFP = green fluorescence protein, wt = wtCFTR, F508del = F508del-CFTR, P439S = P439S-CFTR, P1290S = P1290S-CFTR, E1401K = E1401K-CFTR. Login to comment
178 Similar to the results from I- uptake studies, CFTR mutants P439S-CFTR, P1290S-CFTR and E1401K-CFTR showed a reduced but significant current increase and activation of whole cell conductance. Login to comment
188 Highlights of the possible location of E1401K and P1290S mutations refer to the SAV1866 model. Login to comment
198 While P439S and E1401K are close toATP binding sites and therefore likely to affect NBD-dimerization andATP binding, respectively, P1290S is located near the interface NBD/MSD, and may interfere with side-chain contacts within the membrane spanning subunit. Login to comment
200 In the present study, we report biochemical and functional data on three CFTR missense mutations located in NBD1 (P439S) and in NBD2 (P1290S, E1401K). Login to comment
214 E1401K was identified in compound heterozygosity with the severe F508del mutation. Login to comment
222 In contrast, E1401K-CFTR was correctly localized in the cell membrane, similar to wtCFTR. Login to comment
224 Thus E1401K probably reduces the activity of CFTR Cl- chan- nels while P439S-CFTR and P1290S-CFTR reduce the number of CFTR channels in the membrane. Login to comment
225 Further studies using single channel patch clamp will assess if E1401K reduces whole cell conductance by affecting channel open probability or single channel conductance, and if P439S and P1290S also affect CFTR channel pore/gating mechanism. Login to comment
237 Thus a charge alteration as found in E1401K (glutamic acid → lysine) is likely to disturb ATP binding and therefore reduce CFTR Cl-channel activity, as demonstrated here. Login to comment
240 Therefore, this suggests that a positive charge introduced by E1401K will reduceATPbinding. Login to comment
247 Polyphen analysis indicated that while E1401K (PSIC 1.609) is "possibly damaging", P439S (PSIC 2.396) and P1290S (PSIC 2.108) are "probably damaging" (deleterious) to protein function. Login to comment
250 Although, data obtained with the available bioinformatic tools must be interpreted with caution, these results confirmed experimental data, suggesting that P439S, P1290S and E1401K interfere with protein function. Login to comment
251 In summary, the combination of reduced CFTR whole-cell conductance and normal protein expression detected for E1401K indicates that this mutation causes a defect in CFTR channel activity probably by reducing ATP binding, as suggested by structural analysis. Login to comment
252 Thus, E1401K might belong to class III (defective CFTR regulation). Login to comment
259 This observation might explain the lower whole-cell Cl- currents determined for P1290S-CFTR when compared with P439S-CFTR and E1401K-CFTR. Login to comment
264 Although P439S, P1290S and E1401K might not lead to a typical CF phenotpye, the knowledge of the mechanism by which they affect CFTR, improve the ability to interpret and predict the clinical phenotype in patients carrying the studied mutations and may give important insights about the molecular consequences of similar sequence alterations that have been, or remain to be characterized. Login to comment

[hide] Do common in silico tools predict the clinical con... Clin Genet. 2010 May;77(5):464-73. Epub 2009 Jan 6. Dorfman R, Nalpathamkalam T, Taylor C, Gonska T, Keenan K, Yuan XW, Corey M, Tsui LC, Zielenski J, Durie P
Do common in silico tools predict the clinical consequences of amino-acid substitutions in the CFTR gene?
Clin Genet. 2010 May;77(5):464-73. Epub 2009 Jan 6., [PMID:20059485]

Abstract [show]
Computational methods are used to predict the molecular consequences of amino-acid substitutions on the basis of evolutionary conservation or protein structure, but their utility in clinical diagnosis or prediction of disease outcome has not been well validated. We evaluated three popular computer programs, namely, PANTHER, SIFT and PolyPhen, by comparing the predicted clinical outcomes for a group of known CFTR missense mutations against the diagnosis of cystic fibrosis (CF) and clinical manifestations in cohorts of subjects with CF-disease and CFTR-related disorders carrying these mutations. Owing to poor specificity, none of tools reliably distinguished between individual mutations that confer CF disease from mutations found in subjects with a CFTR-related disorder or no disease. Prediction scores for CFTR mutations derived from PANTHER showed a significant overall statistical correlation with the spectrum of disease severity associated with mutations in the CFTR gene. In contrast, PolyPhen- and SIFT-derived scores only showed significant differences between CF-causing and non-CF variants. Current computational methods are not recommended for establishing or excluding a CF diagnosis, notably as a newborn screening strategy or in patients with equivocal test results.

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64 Mutations in the CFTR gene grouped by clinical category Cystic fibrosis CFTR-related disease No disease T338I D614G L320V V920L L90S M470V H199R S1251N I203M G550R P111A I148T Q1291H R560K L1388Q L183I R170H I1027T S549R D443Y P499A L1414S T908N R668C S549N A455E E1401K Q151K G27E I1234L Y563N R347P C866R S1118C P1290S R75Q A559T V520F P841R M469V E1401G P67L G85E S50Y E1409K R933G G458V G178R Y1032C R248T I980K G85V V392G L973P L137H T351S R334W I444S V938G R792G R560T R555G L1339F D1305E P574H V1240G T1053I D58G G551D L1335P I918M F994C S945L L558S F1337V R810G D1152H G1247R P574S R766M D579G W1098R H949R F200I R352Q L1077P K1351E M244K L206W M1101K D1154G L375F N1303K R1066C E528D D110Y R347H R1070Q A800G P1021S S549K A1364V V392A damaging` (is supposed to affect protein function or structure) and 'probably damaging` (high confidence of affecting protein function or structure). Login to comment

[hide] The H-loop in the second nucleotide-binding domain... Cell Physiol Biochem. 2010;25(2-3):169-80. Epub 2010 Jan 12. Kloch M, Milewski M, Nurowska E, Dworakowska B, Cutting GR, Dolowy K
The H-loop in the second nucleotide-binding domain of the cystic fibrosis transmembrane conductance regulator is required for efficient chloride channel closing.
Cell Physiol Biochem. 2010;25(2-3):169-80. Epub 2010 Jan 12., [PMID:20110677]

Abstract [show]
The cystic fibrosis transmembrane conductance regulator (CFTR) is an ATP-binding cassette (ABC) transporter that functions as a cAMP-activated chloride channel. The recent model of CFTR gating predicts that the ATP binding to both nucleotide-binding domains (NBD1 and NBD2) of CFTR is required for the opening of the channel, while the ATP hydrolysis at NBD2 induces subsequent channel closing. In most ABC proteins, efficient hydrolysis of ATP requires the presence of the invariant histidine residue within the H-loop located in the C-terminal part of the NBD. However, the contribution of the corresponding region (H-loop) of NBD2 to the CFTR channel gating has not been examined so far. Here we report that the alanine substitution of the conserved dipeptide HR motif (HR-->AA) in the H-loop of NBD2 leads to prolonged open states of CFTR channel, indicating that the H-loop is required for efficient channel closing. On the other hand, the HR-->AA substitution lead to the substantial decrease of CFTR-mediated current density (pA/pF) in transfected HEK 293 cells, as recorded in the whole-cell patch-clamp analysis. These results suggest that the H-loop of NBD2, apart from being required for CFTR channel closing, may be involved in regulating CFTR trafficking to the cell surface.

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207 Similar decrease in whole-cell currents has been very recently reported for another H-loop mutation E1401K, associated with CBAVD [66].The above results may seem surprising in light of our finding that the HR→AA CFTR mutant shows increased open probability in single-channel analysis (Fig. 3A). Login to comment