ABCC7 p.Glu826Lys
| ClinVar: |
c.2476G>A
,
p.Glu826Lys
?
, not provided
|
| CF databases: |
c.2476G>A
,
p.Glu826Lys
(CFTR1)
?
, The above sequence alteration was found by DGGE analysis of a sarcoidosis patient. It was sequenced and confirmed with a Restriction site Generating PCR assay. It was absent by DGGE analysis in 208 control chromosomes, and by sequencing 60 chromosomes with unknown CFTR mutations.
|
| Predicted by SNAP2: | A: N (61%), C: D (53%), D: D (53%), F: N (53%), G: D (63%), H: N (66%), I: N (57%), K: N (66%), L: N (57%), M: N (61%), N: N (53%), P: D (63%), Q: N (66%), R: D (59%), S: N (53%), T: N (57%), V: N (53%), W: D (66%), Y: N (53%), |
| Predicted by PROVEAN: | A: N, C: D, D: N, F: D, G: N, H: N, I: D, K: N, L: D, M: D, N: N, P: N, Q: N, R: N, S: N, T: N, V: N, W: D, Y: D, |
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[hide] Insight in eukaryotic ABC transporter function by ... FEBS Lett. 2006 Feb 13;580(4):1064-84. Epub 2006 Jan 19. Frelet A, Klein M
Insight in eukaryotic ABC transporter function by mutation analysis.
FEBS Lett. 2006 Feb 13;580(4):1064-84. Epub 2006 Jan 19., 2006-02-13 [PMID:16442101]
Abstract [show]
With regard to structure-function relations of ATP-binding cassette (ABC) transporters several intriguing questions are in the spotlight of active research: Why do functional ABC transporters possess two ATP binding and hydrolysis domains together with two ABC signatures and to what extent are the individual nucleotide-binding domains independent or interacting? Where is the substrate-binding site and how is ATP hydrolysis functionally coupled to the transport process itself? Although much progress has been made in the elucidation of the three-dimensional structures of ABC transporters in the last years by several crystallographic studies including novel models for the nucleotide hydrolysis and translocation catalysis, site-directed mutagenesis as well as the identification of natural mutations is still a major tool to evaluate effects of individual amino acids on the overall function of ABC transporters. Apart from alterations in characteristic sequence such as Walker A, Walker B and the ABC signature other parts of ABC proteins were subject to detailed mutagenesis studies including the substrate-binding site or the regulatory domain of CFTR. In this review, we will give a detailed overview of the mutation analysis reported for selected ABC transporters of the ABCB and ABCC subfamilies, namely HsCFTR/ABCC7, HsSUR/ABCC8,9, HsMRP1/ABCC1, HsMRP2/ABCC2, ScYCF1 and P-glycoprotein (Pgp)/MDR1/ABCB1 and their effects on the function of each protein.
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No. Sentence Comment
345 E822K and E826K reduced channel conductance and opening [160,165].
X
ABCC7 p.Glu826Lys 16442101:345:10
status: NEW[hide] A functional R domain from cystic fibrosis transme... Proc Natl Acad Sci U S A. 2000 May 9;97(10):5657-62. Ostedgaard LS, Baldursson O, Vermeer DW, Welsh MJ, Robertson AD
A functional R domain from cystic fibrosis transmembrane conductance regulator is predominantly unstructured in solution.
Proc Natl Acad Sci U S A. 2000 May 9;97(10):5657-62., 2000-05-09 [PMID:10792060]
Abstract [show]
Phosphorylation of the regulatory (R) domain initiates cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channel activity. To discover how the function of this domain is determined by its structure, we produced an R domain protein (R8) that spanned residues 708-831 of CFTR. Phosphorylated, but not unphosphorylated, R8 stimulated activity of CFTR channels lacking this domain, indicating that R8 is functional. Unexpectedly, this functional R8 was predominantly random coil, as revealed by CD and limited proteolysis. The CD spectra of both phosphorylated and nonphosphorylated R8 were similar in aqueous buffer. The folding agent trimethylamine N-oxide induced only a small increase in the helical content of nonphosphorylated R8 and even less change in the helical content of phosphorylated R8. These data, indicating that the R domain is predominantly random coil, may explain the seemingly complex way in which phosphorylation regulates CFTR channel activity.
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No. Sentence Comment
198 From this region, the mutations R792G, A800G, E822K, and E826K increase or decrease current, but have not been reported to alter channel properties (38, 39).
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ABCC7 p.Glu826Lys 10792060:198:57
status: NEW[hide] A short segment of the R domain of cystic fibrosis... J Biol Chem. 2002 Jun 21;277(25):23019-27. Epub 2002 Apr 11. Xie J, Adams LM, Zhao J, Gerken TA, Davis PB, Ma J
A short segment of the R domain of cystic fibrosis transmembrane conductance regulator contains channel stimulatory and inhibitory activities that are separable by sequence modification.
J Biol Chem. 2002 Jun 21;277(25):23019-27. Epub 2002 Apr 11., 2002-06-21 [PMID:11950844]
Abstract [show]
The regulatory (R) domain of the cystic fibrosis transmembrane conductance regulator (CFTR) contains consensus phosphorylation sites for cAMP-dependent protein kinase (PKA) that are the basis for physiological regulation of the CFTR chloride channel. A short peptide segment in the R domain with a net negative charge of B9 (amino acids 817-838, NEG2) and predicted helical tendency is shown to play a critical role in CFTR chloride channel function. Deletion of NEG2 from CFTR completely eliminates the PKA dependence of channel activity. Exogenous NEG2 peptide interacts with CFTR to exert both stimulatory and inhibitory effects on the channel function. The NEG2 peptide with sequence scrambled to remove helical tendencies also inhibits channel function, but does not stimulate. Similar results are found for a NEG2 peptide whose helical structure is disrupted by a proline residue. When six of the negatively charged carboxylic acid residues are replaced by their cognate amides, reducing net negative charge to B3, but increasing helical propensity as assessed by circular dichroism, the peptide stimulates CFTR channel function, but does not inhibit. We speculate that the NEG2 region interacts with other cytosolic domains of CFTR to control opening and closing transitions of the chloride channel.
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No. Sentence Comment
232 Three mutations are reported in the NEG2 region (E822K, E826K, and D836Y), two of which were obtained from patients with cystic fibrosis (E822K and D836Y).
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ABCC7 p.Glu826Lys 11950844:232:56
status: NEW233 Single channel studies of E822K and E826K indicate that both mutations result in reduced Po compared with wt-CFTR (34).
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ABCC7 p.Glu826Lys 11950844:233:36
status: NEW[hide] The phenotypic consequences of CFTR mutations. Ann Hum Genet. 2003 Sep;67(Pt 5):471-85. Rowntree RK, Harris A
The phenotypic consequences of CFTR mutations.
Ann Hum Genet. 2003 Sep;67(Pt 5):471-85., [PMID:12940920]
Abstract [show]
Cystic fibrosis is a common autosomal recessive disorder that primarily affects the epithelial cells in the intestine, respiratory system, pancreas, gall bladder and sweat glands. Over one thousand mutations have currently been identified in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene that are associated with CF disease. There have been many studies on the correlation of the CFTR genotype and CF disease phenotype; however, this relationship is still not well understood. A connection between CFTR genotype and disease manifested in the pancreas has been well described, but pulmonary disease appears to be highly variable even between individuals with the same genotype. This review describes the current classification of CFTR mutation classes and resulting CF disease phenotypes. Complex disease alleles and modifier genes are discussed along with alternative disorders, such as disseminated bronchiectasis and pancreatitis, which are also thought to result from CFTR mutations.
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No. Sentence Comment
79 Furthermore, mutants T665S and E826K showed no difference from the wild-type channel conductance.
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ABCC7 p.Glu826Lys 12940920:79:31
status: NEW[hide] Gender-sensitive association of CFTR gene mutation... Mol Hum Reprod. 2005 Aug;11(8):607-14. Epub 2005 Aug 26. Morea A, Cameran M, Rebuffi AG, Marzenta D, Marangon O, Picci L, Zacchello F, Scarpa M
Gender-sensitive association of CFTR gene mutations and 5T allele emerging from a large survey on infertility.
Mol Hum Reprod. 2005 Aug;11(8):607-14. Epub 2005 Aug 26., [PMID:16126774]
Abstract [show]
Human infertility in relation to mutations affecting the cystic fibrosis transmembrane regulator (CFTR) gene has been investigated by different authors. The role of additional variants, such as the possible forms of the thymidine allele (5T, 7T and 9T) of the acceptor splice site of intron 8, has in some instances been considered. However, a large-scale analysis of the CFTR gene and number of thymidine residues, alone and in combination, in the two sexes had not yet been addressed. This was the aim of this study. Two groups were compared, a control group of 20,532 subjects being screened for perspective reproduction, and the patient group represented by 1854 idiopathically infertile cases. Analyses involved PCR-based CFTR mutations assessment, reverse dot-blot IVS8-T polymorphism analyses, denaturing gradient gel electrophoresis (DGGE) and DNA sequencing. The expected 5T increase in infertile men was predominantly owing to the 5/9 genotypic class. The intrinsic rate of 5T fluctuated only slightly among groups, but some gender-related differences arose when comparing their association. Infertile men showed a significantly enriched 5T + CFTR mutation co-presence, distributed in the 5/9 and 5/7 classes. In contrast, females, from both the control and the infertile groups, showed a trend towards a pronounced reduction of such association. The statistical significance of the difference between expected and observed double occurrence of 5T + CFTR traits in women suggests, in line with other reports in the literature, a possible survival-hampering effect. Moreover, regardless of the 5T status, CFTR mutations appear not to be involved in female infertility. These results underline the importance of (i) assessing large sample populations and (ii) considering separately the two genders, whose genotypically opposite correlations with these phenomena may otherwise tend to mask each other.
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No. Sentence Comment
76 This test involved nine subjects from the infertile group, revealing the occurrence of the following rare mutations: E217G, T1054A, W356X, D443Y and 3667insTC in males and L997F and R297Q in females and 29 subjects from the control, in which we found: A1009T, D110Y, E826K, G1069R, G1130A, G194V, I556V, L320F, M348K, M82V, P1290T, R117C, R352W, R74W, S42F, S660T, S911R, S912L, T1086A, T582S, V920L and Y89C.
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ABCC7 p.Glu826Lys 16126774:76:270
status: NEW144 *Genotypes include subjects (whose number is indicated within parentheses) that carry the following CFTR mutations 2789+5G/A, E826K, ∆F508, D443Y, 3849+10Kb C/T 1717-8G/A and 3667insTCAA.
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ABCC7 p.Glu826Lys 16126774:144:126
status: NEW[hide] Contribution of the CFTR gene, the pancreatic secr... Clin Genet. 2007 May;71(5):451-7. Tzetis M, Kaliakatsos M, Fotoulaki M, Papatheodorou A, Doudounakis S, Tsezou A, Makrythanasis P, Kanavakis E, Nousia-Arvanitakis S
Contribution of the CFTR gene, the pancreatic secretory trypsin inhibitor gene (SPINK1) and the cationic trypsinogen gene (PRSS1) to the etiology of recurrent pancreatitis.
Clin Genet. 2007 May;71(5):451-7., [PMID:17489851]
Abstract [show]
Acute recurrent/chronic pancreatitis (CP) is a complex multigenic disease. This is a case-control study consisting of 25 Greek patients with CP and a control population of 236 healthy Greek subjects. The whole coding area and neighboring intronic regions of the three genes were screened. Seventeen of 25 patients (68%) had mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene: nine compound heterozygotes with either mild or severe mutations and eight heterozygotes. Four patients (16%) carried CFTR-modulating haplotypes V470-TG11-T5 and V470-TG12-T7. All were negative for PRSS1 gene mutations, while variants c.486C/T and c.738C/T were found in nine patients each, three homozygotes for the minor alleles. Two carried SPINK1 gene mutation p.N34S, one being transheterozygote with CFTR mutation p.F1052V. The promoter variant -253T>C was found in four individuals (one homozygous for the minor allele), all four being transheterozygotes with mutations in the CFTR gene as well. Finally two carried c.272C/T in the 3' untranslated region, one being a p.N34S carrier as well. In total, 80% (20/25) of patients had a molecular defect in one or both of the CFTR and SPINK1 genes, suggesting that mutations/variants in the CFTR plus or minus mutations in the SPINK1, but not the PRSS1 gene, may confer a high risk for recurrent pancreatitis.
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No. Sentence Comment
93 a Additional mutations found in the controls: p.R1162L (1.66%), p.D565G (0.47%), p.A120T (0.47%) and 0.24% each for p.R297Q, p.L997F, p.E826K, p.I807M, p.S495Y and p.C491S.
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ABCC7 p.Glu826Lys 17489851:93:136
status: NEW[hide] Characterization of 19 disease-associated missense... Hum Mol Genet. 1998 Oct;7(11):1761-9. Vankeerberghen A, Wei L, Jaspers M, Cassiman JJ, Nilius B, Cuppens H
Characterization of 19 disease-associated missense mutations in the regulatory domain of the cystic fibrosis transmembrane conductance regulator.
Hum Mol Genet. 1998 Oct;7(11):1761-9., [PMID:9736778]
Abstract [show]
In order to gain a better insight into the structure and function of the regulatory domain (RD) of the cystic fibrosis transmembrane conductance regulator (CFTR) protein, 19 RD missense mutations that had been identified in patients were functionally characterized. Nine of these (I601F, L610S, A613T, D614G, I618T, L619S, H620P, G628R and L633P) resulted in aberrant processing. No or a very small number of functional CFTR proteins will therefore appear at the cell membrane in cells expressing these mutants. These mutations were clustered in the N-terminal part of the RD, suggesting that this subdomain has a folding pattern that is very sensitive to amino acid changes. Mutations that caused no aberrant processing were further characterized at the electrophysiological level. First, they were studied at the whole cell level in Xenopus laevis oocytes. Mutants that induced a whole cell current that was significantly different from wild-type CFTR were subsequently analysed at the single channel level in COS1 cells transiently expressing the different mutant and wild-type proteins. Three mutant chloride channels, G622D, R792G and E822K CFTR, were characterized by significantly lower intrinsic chloride channel activities compared with wild-type CFTR. Two mutations, H620Q and A800G, resulted in increased intrinsic chloride transport activities. Finally, T665S and E826K CFTR had single channel properties not significantly different from wild-type CFTR.
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No. Sentence Comment
9 Finally, T665S and E826K CFTR had single channel properties not significantly different from wild-type CFTR.
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ABCC7 p.Glu826Lys 9736778:9:19
status: NEW68 Primers used for mutagenesis Primer Sequence I601F (a1933t) 5'-CTA ACA AAA CTA GGT TTT TGG TCA CTT C-3' L610S (t1961c) 5'-CTA AAA TGG AAC ATT CAA AGA AAG CTG-3' A613T (g1969a) 5'-CAT TTA AAG AAA ACT GAC AAA ATA TTA-3' D614G (a1973g) 5'-CAT TTA AAG AAA GCT GGC AAA ATA TTA A-3' I618T (t1985c) 5'-GAC AAA ATA TTA ACT TTG CAT GAA GG-3' L619S (t1988c) 5'-GAC AAA ATA TTA ATT TCG CAT GAA GGT-3' H620P (a1991c) 5'-CAA AAT ATT AAT TTT GCC TGA AGG TAG C-3' H620Q (t1992g) 5'-AAT ATT AAT TTT GCA GGA AGG TAG CAG-3' G622D (g1997a) 5'-TTG CAT GAA GAT AGC AGC TAT TTT TAT G-3' G628R (g2014c) 5'-GCA GCT ATT TTT ATC GGA CAT TTT C-3' L633P (t2030c) 5'-CAT TTT CAG AAC CCC AAA ATC TAC AGC-3' D648V (a2075t) 5'-CTC ATG GGA TGT GTT TCT TTC GAC C-3' T665S (a2125t) 5'-CAA TCC TAA CTG AGT CCT TAC ACC G-3' F693L (t2209c) 5'-CAG ACT GGA GAG CTT GGG GAA AAA AG-3' R766M (g2429t) 5'-GCA CGA AGG ATG CAG TCT GTC CTG-3' R792G (c2506g) 5'-CAG CAT CCA CAG GAA AAG TGT CAC TG-3' A800G (c2531g) 5'-CTG GCC CCT CAG GGA AAC TTG ACT G-3' I807M (a2553g) 5'-CTG AAC TGG ATA TGT ATT CAA GAA GG-3' E822K (g2596a) 5'-GGC TTG GAA ATA AGT AAA GAA ATT AAC G-3' E826K (g2608a) 5'-GAA GAA ATT AAC AAA GAA GAC TTA AAG-3' Selection primer BstBI 5'-CTC TGG GGT CCG GAA TGA CCG AC-3' Two primers were used for each mutagenesis reaction.
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ABCC7 p.Glu826Lys 9736778:68:1122
status: NEW77 Mutations detected in patients (I601F, L610S, A613T, D614G, I618T, L619S, H620P, H620Q, D622G, G628R, L633P, T665S, F693L, K698R, V754M, R766M, R792G, A800G, I807M, E822K and E826K) are indicated in bold and underlined, the PKA phosphorylation sites by an arrow and the two acidic domains are boxed.
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ABCC7 p.Glu826Lys 9736778:77:175
status: NEW83 Four mutations (T665S, R792G, E822K and E826K) caused a significant reduction in the cAMP-induced chloride current.
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ABCC7 p.Glu826Lys 9736778:83:40
status: NEW87 Maturation pattern of RD mutations and their associated phenotype found in patients with the indicated genotype (when the mutation is associated with CF, only the pancreas status is given) Mutation A-form B-form C-form Clinical data Genotype Phenotype Reference I601F + + - I601F/G542X PS M. Schwarz, personal communication L610S + + - Unknown Unknown A613T + + - Unknown Unknown D614G + + - D614G/unknown PI 14 I618T + + - I618T/dF508 PS G.R. Cutting, personal communication L619S + + - L619S/unknown PI B. Tümmler, personal communication H620P + + - H620P/R1158X PS M. Schwarz, personal communication H620Q + + + H620Q/dF508 PI T. Dörk, personal communication G622D + + + G622D/unknown Oligospermia J. Zielenski, personal communication G628R + + - Unknown Unknown L633P + + - L633P/3659delC M. Schwarz, personal communication D648V + + + D648V/3849+10kb C/T PI C. Ferec, personal communication T665S + + + Unknown Unknown F693L + + + F693L/W1282X Healthy C. Ferec; CF Genetic Analysis Consortium R766M + + + R766M/R792G CBAVD D. Glavac, personal communication R792G + + + R766M/R792G CBAVD D. Glavac, personal communication A800G + + + A800G/unknown CBAVD 34 I807M + + + I807M/unknown CBAVD Our observation E822K + + + E822K/unknown PI 35 E826K + + + E826K/unknown Thoracic sarcoidosis C. Bombieri, personal communication +, the protein matures up to that form; -, the protein does not reach the respective maturation step.
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ABCC7 p.Glu826Lys 9736778:87:1251
status: NEWX
ABCC7 p.Glu826Lys 9736778:87:1263
status: NEW123 Mutations that did not affect maturation (H620Q, G622D, D648V, T665S, F693L, R766M, R792G, A800G, I807M, E822K and E826K) were subsequently analysedat theelectrophysiologi- cal level.
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ABCC7 p.Glu826Lys 9736778:123:115
status: NEW131 The remaining mutations (D648V, T665S, F693L, R766M, I807M and E826K) caused no significant alterations in intrinsic chloride channel activity.
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ABCC7 p.Glu826Lys 9736778:131:63
status: NEW[hide] Complete mutational screening of the CFTR gene in ... Hum Genet. 1998 Dec;103(6):718-22. Bombieri C, Benetazzo M, Saccomani A, Belpinati F, Gile LS, Luisetti M, Pignatti PF
Complete mutational screening of the CFTR gene in 120 patients with pulmonary disease.
Hum Genet. 1998 Dec;103(6):718-22., [PMID:9921909]
Abstract [show]
In order to determine the possible role of the cystic fibrosis transmembrane regulator (CFTR) gene in pulmonary diseases not due to cystic fibrosis, a complete screening of the CFTR gene was performed in 120 Italian patients with disseminated bronchiectasis of unknown cause (DBE), chronic bronchitis (CB), pulmonary emphysema (E), lung cancer (LC), sarcoidosis (S) and other forms of pulmonary disease. The 27 exons of the CFTR gene and their intronic flanking regions were analyzed by denaturing gradient gel electrophoresis and automatic sequencing. Mutations were detected in 11/23 DBE (P = 0.009), 7/25 E, 5/27 CB, 5/26 LC, 5/8 S (P = 0.013), 1/4 tuberculosis, and 1/5 pneumonia patients, and in 5/33 controls. Moreover, the IVS8-5T allele was detected in 6/25 E patients (P = 0.038). Four new mutations were identified: D651N, 2377C/T, E826K, and P1072L. These results confirm the involvement of the CFTR gene in disseminated bronchiectasis of unknown origin, and suggest a possible role for CFTR gene mutations in sarcoidosis, and for the 5T allele in pulmonary emphysema.
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No. Sentence Comment
4 Four new mutations were identified: D651N, 2377C/T, E826K, and P1072L.
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ABCC7 p.Glu826Lys 9921909:4:52
status: NEW63 Three novel mutations were first identified in this study: D651N, E826K, and P1072L.
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ABCC7 p.Glu826Lys 9921909:63:66
status: NEW77 E826K, a G to A substitution, was found at nucleotide 2608 in exon 13.
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ABCC7 p.Glu826Lys 9921909:77:0
status: NEW88 of cases CFTR gene PolyTb status tested mutationa DBE 23 1 G576A-R668C/L997F 7/9 1 ∆F508/L997F 9/9 1 ∆F508/- 7/9 1 R1066C/- 5/7 1 3667ins4/- 5/7 1 R75Q/- 7/7 1 M1137V/- 7/7 1 -/- 5/5 3 -/- 5/7 10 -/- 7/7 2 -/- 7/9 CB 27 1 P111L/- 7/7 1 R117H/- 7/7 1 E585X/- 7/7 1 P1072L/- 7/7 1 -/- 5/7 15 -/- 7/7 6 -/- 7/9 1 -/- 9/9 E 25 1 R668C/- 7/7 6 -/- 5/7 16 -/- 7/7 6 -/- 7/9 S 8 1 E826K/- 7/7 1 ∆F508/- 7/9 1 4382delA/- 7/7 1 L997F/- 7/9 1 V754M/- 7/9 3 -/- 7/7 LC 26 1 I148T/- 5/7 1 D1270N-R74W 5/7 1 D651N/- 7/7 1 Y301C/- 7/7 1 -/- 5/7 16 -/- 7/7 5 -/- 7/9 TB 4 1 -/- 5/7 1 -/- 7/7 2 -/- 7/9 Pneumonia 5 4 -/- 7/7 1 -/- 5/7 Pnx 2 2 -/- 7/7 Controls 68 1 L997F/- 7/9 1 R31C/- 7/7 1 I506V/- 5/7 1 -/- 5/7 1 -/- 5/9 23 -/- 7/7 4 -/- 7/9 1 -/- 9/9 2 ?
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ABCC7 p.Glu826Lys 9921909:88:388
status: NEW105 Two deletions (∆F508 and 4382delA, a frameshift deletion generating a stop codon 15 amino acids downstream) and three missense mutations (V754M, E826K, L997F) were detected.
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ABCC7 p.Glu826Lys 9921909:105:152
status: NEW[hide] Phosphorylation site independent single R-domain m... FEBS Lett. 1998 Nov 13;439(1-2):121-6. Wei L, Vankeerberghen A, Cuppens H, Droogmans G, Cassiman JJ, Nilius B
Phosphorylation site independent single R-domain mutations affect CFTR channel activity.
FEBS Lett. 1998 Nov 13;439(1-2):121-6., [PMID:9849891]
Abstract [show]
We investigated CFTR channel activity of mature R-domain mutants showing single alterations at sites other than the predicted phosphorylation sites. All mutations were found in cystic fibrosis (CF) patients (H620Q, E822K and E826K). The macroscopic CFTR chloride conductance induced by phosphorylation was significantly enhanced in Xenopus oocytes injected with mRNA of H620Q but reduced in the E822K and E826K mutants compared to wild type CFTR. The anion permeability sequence for all three mutants was the same as that of wild type CFTR. Cell attached single channel studies in COS cells revealed that both open channel probability and/or the number of functional channels were either higher (H620Q) or lower (E822K and E826K) than in wild type CFTR. Single channel conductances were unchanged in all mutants. Our results suggest that additional sites in the R-domain other than phosphorylation sites influence gating of CFTR channels.
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No. Sentence Comment
1 All mutations were found in cystic fibrosis (CF) patients (H620Q, E822K and E826K).
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ABCC7 p.Glu826Lys 9849891:1:76
status: NEW2 The macroscopic CFTR chloride conductance induced by phosphorylation was significantly enhanced in Xenopus oocytes injected with mRNA of H620Q but reduced in the E822K and E826K mutants compared to wild type CFTR.
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ABCC7 p.Glu826Lys 9849891:2:172
status: NEW4 Cell attached single channel studies in COS cells revealed that both open channel probability and/or the number of functional channels were either higher (H620Q) or lower (E822K and E826K) than in wild type CFTR.
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ABCC7 p.Glu826Lys 9849891:4:182
status: NEW25 Three di¡erent mutations, t1992g (= H620Q), g2596a (= E822K) and g2608a (= E826K), were introduced using the Transformer Site-Directed Mutagenesis kit (Clontech).
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ABCC7 p.Glu826Lys 9849891:25:79
status: NEW76 COS cells were transfected with wt CFTR, E822K CFTR (top), H620Q CFTR (bottom) and E826K CFTR (bottom) and selected for 2 weeks with G418.
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ABCC7 p.Glu826Lys 9849891:76:83
status: NEW88 In this study, the maturation pattern of three mutant R-domain proteins (H620Q-CFTR, E822K-CFTR and E826K-CFTR) has been characterized.
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ABCC7 p.Glu826Lys 9849891:88:100
status: NEW92 Whole cells currents of mutants in Xenopus oocytes Whole cell membrane currents were recorded from Xenopus oocytes injected with RNA transcribed from either wild type or mutant (H620Q, E822K, E826K) constructs.
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ABCC7 p.Glu826Lys 9849891:92:192
status: NEW100 The two R-domain proteins E822K and E826K, in which a negatively charged glutamic acid was exchanged for a positively charged lysine, showed a signi'cantly smaller phos-cock activated conductance.
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ABCC7 p.Glu826Lys 9849891:100:36
status: NEW101 Oocytes expressing the mutant R-domain protein (H620Q), in which a predominantly positively charged histidine was substituted by a less charged glutamine (at pH 7.2), showed a much larger conductance activated by application of phos-cock.
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ABCC7 p.Glu826Lys 9849891:101:30
status: NEW103 The conductance for E822K and E826K was 3.55 þ 0.44 WS (n = 6) and 4.24 þ 0.37 WS (n = 6), as compared to 7.57 þ 0.65 WS (n = 14) in the wild type.
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ABCC7 p.Glu826Lys 9849891:103:30
status: NEW134 We found that the open probability of the E822K and E826K mutants was signi'cantly lower than that of wild type CFTR, whereas that of the H620Q mutant was strongly enhanced compared to wild type (not shown).
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ABCC7 p.Glu826Lys 9849891:134:52
status: NEWX
ABCC7 p.Glu826Lys 9849891:134:192
status: NEW136 The average number of activated channels is 2.6 þ 0.306 (n = 10) for wild-type CFTR, 3.51 þ 0.428 (n = 6) for H620Q, 1.0 þ 0.000 (n = 4) for E822K and 1.66 þ 0.211 (n = 6) for E826K.
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ABCC7 p.Glu826Lys 9849891:136:196
status: NEW137 The di¡erences between wild type CFTR and E822K and E826K are signi'- cant (P 6 0.05).
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ABCC7 p.Glu826Lys 9849891:137:57
status: NEW141 All the mutations studied here are located outside the phosphorylation sites, which may suggest that other regions in the R-domain, especially the highly conserved regions where E822K and E826K were l,ocated are important for the regulation of the CFTR Cl3 channel.
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ABCC7 p.Glu826Lys 9849891:141:188
status: NEW145 A comparison of the three R-domain mutants leads to a remarkable conclusion: both mutations, E822K and E826K, in which negatively charged glutamic acids were replaced by positively charged lysine had a signi'cantly reduced CFTR channel activity, whereas the H620Q mutation, in which positively charged histidine was replaced by the more neutral amino acid glutamine, had a much higher channel activity.
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ABCC7 p.Glu826Lys 9849891:145:103
status: NEW74 COS cells were transfected with wt CFTR, E822K CFTR (top), H620Q CFTR (bottom) and E826K CFTR (bottom) and selected for 2 weeks with G418.
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ABCC7 p.Glu826Lys 9849891:74:83
status: NEW86 In this study, the maturation pattern of three mutant R-domain proteins (H620Q-CFTR, E822K-CFTR and E826K-CFTR) has been characterized.
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ABCC7 p.Glu826Lys 9849891:86:100
status: NEW90 Whole cells currents of mutants in Xenopus oocytes Whole cell membrane currents were recorded from Xenopus oocytes injected with RNA transcribed from either wild type or mutant (H620Q, E822K, E826K) constructs.
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ABCC7 p.Glu826Lys 9849891:90:192
status: NEW98 The two R-domain proteins E822K and E826K, in which a negatively charged glutamic acid was exchanged for a positively charged lysine, showed a signi'cantly smaller phos-cock activated conductance.
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ABCC7 p.Glu826Lys 9849891:98:36
status: NEW132 We found that the open probability of the E822K and E826K mutants was signi'cantly lower than that of wild type CFTR, whereas that of the H620Q mutant was strongly enhanced compared to wild type (not shown).
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ABCC7 p.Glu826Lys 9849891:132:52
status: NEW135 The di&#a1;erences between wild type CFTR and E822K and E826K are signi'- cant (P 6 0.05).
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ABCC7 p.Glu826Lys 9849891:135:56
status: NEW139 All the mutations studied here are located outside the phosphorylation sites, which may suggest that other regions in the R-domain, especially the highly conserved regions where E822K and E826K were l,ocated are important for the regulation of the CFTR Cl3 channel.
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ABCC7 p.Glu826Lys 9849891:139:188
status: NEW143 A comparison of the three R-domain mutants leads to a remarkable conclusion: both mutations, E822K and E826K, in which negatively charged glutamic acids were replaced by positively charged lysine had a signi'cantly reduced CFTR channel activity, whereas the H620Q mutation, in which positively charged histidine was replaced by the more neutral amino acid glutamine, had a much higher channel activity.
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ABCC7 p.Glu826Lys 9849891:143:103
status: NEW[hide] Definition of a "functional R domain" of the cysti... Mol Genet Metab. 2000 Sep-Oct;71(1-2):245-9. Chen JM, Scotet V, Ferec C
Definition of a "functional R domain" of the cystic fibrosis transmembrane conductance regulator.
Mol Genet Metab. 2000 Sep-Oct;71(1-2):245-9., [PMID:11001817]
Abstract [show]
The R domain of the cystic fibrosis transmembrane conductance regulator (CFTR) was originally defined as 241 amino acids, encoded by exon 13. Such exon/intron boundaries provide a convenient way to define the R domain, but do not necessarily reflect the corresponding functional domain within CFTR. A two-domain model was later proposed based on a comparison of the R-domain sequences from 10 species. While RD1, the N-terminal third of the R domain is highly conserved, RD2, the large central region of the R domain has less rigid structural requirements. Although this two-domain model was given strong support by recent functional analysis data, the simple observation that two of the four main phosphorylation sites are excluded from RD2 clearly indicates that RD2 still does not satisfy the requirements of a "functional R domain." Nevertheless, knowledge of the CFTR structure and function accumulated over the past decade and reevaluated in the context of a comprehensive sequence comparison of 15 CFTR homologues made it possible to define such a "functional R domain," i.e., amino acids C647 to D836. This definition is validated primarily because it contains all of the important potential consensus phosphorylation sequences. In addition, it includes the highly charged motif from E822 to D836. Finally, it includes all of the deletions/insertions in this region. This definition also aids in understanding the effects of missense mutations occurring within this domain.
Comments [show]
None has been submitted yet.
No. Sentence Comment
47 Conversely, E822K and E826K both change a stringently or well-conserved, negatively charged residue to a positively charged one and therefore would be speculated to produce some functional consequences.
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ABCC7 p.Glu826Lys 11001817:47:22
status: NEW[hide] Single-cell high resolution melting analysis: A no... J Cyst Fibros. 2015 Oct 19. pii: S1569-1993(15)00217-9. doi: 10.1016/j.jcf.2015.09.009. Destouni A, Poulou M, Kakourou G, Vrettou C, Tzetis M, Traeger-Synodinos J, Kitsiou-Tzeli S
Single-cell high resolution melting analysis: A novel, generic, pre-implantation genetic diagnosis (PGD) method applied to cystic fibrosis (HRMA CF-PGD).
J Cyst Fibros. 2015 Oct 19. pii: S1569-1993(15)00217-9. doi: 10.1016/j.jcf.2015.09.009., [PMID:26493493]
Abstract [show]
BACKGROUND: Institutions offering CF-PGD face the challenge of developing and optimizing single cell genotyping protocols that should cover for the extremely heterogeneous CF mutation spectrum. Here we report the development and successful clinical application of a generic CF-PGD protocol to facilitate direct detection of any CFTR nucleotide variation(s) by HRMA and simultaneous confirmation of diagnosis through haplotype analysis. METHODS: A multiplex PCR was optimized supporting co-amplification of any CFTR exon-region, along with 6 closely linked STRs. Single cell genotypes were established through HRM analysis following melting of the 2nd round PCR products and were confirmed by STR haplotype analysis of the 1st PCR products. The protocol was validated pre-clinically, by testing 208 single lymphocytes, isolated from whole blood samples from 4 validation family trios. Fifteen PGD cycles were performed and 103 embryos were biopsied. RESULTS: In 15 clinical PGD cycles, genotypes were achieved in 88/93 (94.6%) embryo biopsy samples, of which 57/88 (64.8%) were deemed genetically suitable for embryo transfer. Amplification failed at all loci for 10/103 blastomeres biopsied from poor quality embryos. Six clinical pregnancies were achieved (2 twin, 4 singletons). PGD genotypes were confirmed following conventional amniocentesis or chorionic villus sampling in all achieved pregnancies. CONCLUSIONS: The single cell HRMA CF-PGD protocol described herein is a flexible, generic, low cost and robust genotyping method, which facilitates the analysis of any CFTR genotype combination. Single-cell HRMA can be beneficial to other clinical settings, for example the detection of single nucleotide variants in single cells derived from clinical tumor samples.
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None has been submitted yet.
No. Sentence Comment
101 PGD case no. Genotype combination (HGVS CFTR reference sequences NM000492.3 and NG016465.1) LSC exons (legacy nomenclature) (Montgomery et al., 2007) No. of blastomeres received for diagnosis No. of blastomeres amplified No. of blastomeres genotyped No. of unaffected blastomeres Pregnancy Confirmation of PGD result by PNDa 1 p.Arg334Gln and c.489+3ANG 7.2 and 4.2 5 4 4 2 No N/A 2 p.Phe508del and p.Phe508del 10 11 10 10 6 Yes Yes 3 p.Phe508del and c.489+1GNT 10 and 4.2 8 7 7 4 Yes Yes 4 p.Phe508del and p.Leu732X 10 and 13.3 10 8 8 5 Yes Yes 5 p.Phe508del and p.Asp1152His 10 and 18 4 3 3 3 Yes Yes 6 p.Phe508del and c.2051_2052delAAinsG 10 and 13.2 5 4 4 3 Yes Yes 7 p.Phe508del and p.Gly1069Arg 10 and 17bA1 9 9 9 4 Yes No (BP) 8 p.Phe508del and p.Gly542X 10 and 11 3 3 3 2 Yes Yes 9 p.Phe508del and c.3140-26ANG 10 and 17bA1 8 8 7 3 No N/A 10 p.Gly542X and p.Gly542X 11 2 2 2 1 No transfer 11 p.Glu826Lys and p.Phe508del 13.4 and 10 6 6 4 3 Yes Miscarried 12 p.D1312G and c.489+1GNT 21 and 4.2 11 9 9 8 Yes Miscarried 13 p.Glu279Asp and p.Phe508del 6b and 10 7 7 5 3 No N/A 14 p.Phe508del and p.Gly1069Arg 10 and 17bA1 9 8 8 5 Yes No (BP) 15 p.Glu822X and p.Gly1069Arg 13.4 and 17bA1 5 5 5 5 Yesb Miscarried Total 103 93 88 57 N/A: non-applicable, BP: biochemical pregnancy.
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ABCC7 p.Glu826Lys 26493493:101:902
status: NEW