ABCMdb

ABCC7 p.Gly551Ala

ClinVar:	c.1651G>A , p.Gly551Ser D , Pathogenic c.1652G>A , p.Gly551Asp D , Pathogenic
CF databases:	c.1652G>A , p.Gly551Asp D , CF-causing ; CFTR1: This mutation has been found in six Caucasian CF chromosomes out of 155 eamined for a frequency of 4 %. It has not been found on any Black CF chromosomes. This mutation appears to be associated with a particular ten site haplotype shown on the following pages. We have not detected this mutation on any normal Caucasian chromosomes with similar haplotypes or other haplotypes. c.1651G>A , p.Gly551Ser D , CF-causing ; CFTR1: This mutation can be detected using ASOs: normal 5' GAGTGGAGGTCAACG 3', mutant 5' GAGTGGAAGTCAACG 3' with a final wash at 42 degrees celsius in 40 mM NaHPO4, 1 mM EDTA, 0.5 % SDS for 15 minutes. Two patients were found to be homozygous for this mutation. Their parents are second cousins and each carries the G551S mutation. These patients are remarkable in that they have a mild disease without elevated Na+ levels. One patient had decreased lung function, Pseudomonas infections, chronic pancreatitis, clubbing, and is currently 49 years old. This mutation was not found in 363 non-[delta]F508 CF chromosomes, nor in over 700[delta]F508 chromosomes, nor in a small number of normal chromosomes.
Predicted by SNAP2:	A: D (95%), C: D (95%), D: D (71%), E: D (95%), F: 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: N (61%), T: D (95%), V: D (95%), W: D (95%), Y: D (95%),
Predicted by PROVEAN:	A: D, C: D, D: D, E: D, F: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: D, T: D, V: D, W: D, Y: D,

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Publications
[hide] Mutations at the signature sequence of CFTR create... J Gen Physiol. 2009 Jan;133(1):69-77. Wang X, Bompadre SG, Li M, Hwang TC
Mutations at the signature sequence of CFTR create a Cd(2+)-gated chloride channel.
J Gen Physiol. 2009 Jan;133(1):69-77., [PMID:19114635]

Abstract [show]
The canonical sequence LSGGQ, also known as the signature sequence, defines the adenosine triphosphate (ATP)-binding cassette transporter superfamily. Crystallographic studies reveal that the signature sequence, together with the Walker A and Walker B motifs, forms the ATP-binding pocket upon dimerization of the two nucleotide-binding domains (NBDs) in a head-to-tail configuration. The importance of the signature sequence is attested by the fact that a glycine to aspartate mutation (i.e., G551D) in cystic fibrosis transmembrane conductance regulator (CFTR) results in a severe phenotype of cystic fibrosis. We previously showed that the G551D mutation completely eliminates ATP-dependent gating of the CFTR chloride channel. Here, we report that micromolar [Cd(2+)] can dramatically increase the activity of G551D-CFTR in the absence of ATP. This effect of Cd(2+) is not seen in wild-type channels or in G551A. Pretreatment of G551D-CFTR with the cysteine modification reagent 2-aminoethyl methane thiosulfonate hydrobromide protects the channel from Cd(2+) activation, suggesting an involvement of endogenous cysteine residue(s) in mediating this effect of Cd(2+). The mutants G551C, L548C, and S549C, all in the signature sequence of CFTR's NBD1, show robust response to Cd(2+). On the other hand, negligible effects of Cd(2+) were seen with T547C, Q552C, and R553C, indicating that a specific region of the signature sequence is involved in transmitting the signal of Cd(2+) binding to the gate. Collectively, these results suggest that the effect of Cd(2+) is mediated by a metal bridge formation between yet to be identified cysteine residue(s) and the engineered aspartate or cysteine in the signature sequence. We propose that the signature sequence serves as a switch that transduces the signal of ligand binding to the channel gate.

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20 This effect of Cd2+ is not seen in wild-type channels or in G551A. Login to comment
96 Like the G551C mutant, G551A-CFTR remains responsive to ATP. Login to comment
97 However, the effect of Cd2+ on G551A-CFTR is negligibly small compared with that of G551C-CFTR (Fig. 5 A vs. Fig. 4 B). Login to comment
98 This difference between G551A and G551C was quantified in Fig. 5 B, where we compared the current generated by 1 mM ATP with the current generated by 10 μM Cd2+ for these two mutants. Login to comment
137 Figure 5. Comparison of Cd2+ and ATP-induced currents between G551A and G551C mutants. Login to comment
138 (A) Representative current trace of G551A-CFTR. Login to comment
139 Although G551A-CFTR remains ATP dependent, Cd2+ fails to increase the activity of the channels. Login to comment
140 (B) The ratio of currents induced by 10 μM Cd2+ and those with 1 mM ATP for G551C-CFTR and G551A-CFTR. Login to comment

[hide] Heterogeneity of reproductive tract abnormalities ... Fertil Steril. 1998 Oct;70(4):724-8. Jarvi K, McCallum S, Zielenski J, Durie P, Tullis E, Wilchanski M, Margolis M, Asch M, Ginzburg B, Martin S, Buckspan MB, Tsui LC
Heterogeneity of reproductive tract abnormalities in men with absence of the vas deferens: role of cystic fibrosis transmembrane conductance regulator gene mutations.
Fertil Steril. 1998 Oct;70(4):724-8., [PMID:9797105]

Abstract [show]
OBJECTIVE: To determine if the types of reproductive tract abnormalities linked to absence of the vas deferens varies with the cystic fibrosis transmembrane conductance regulator (CFTR) genotype. DESIGN: Prospective data gathering. SETTING: University infertility clinic. PATIENT(S): Forty-six infertile men with absence of the scrotal vas deferens and no signs of cystic fibrosis. INTERVENTION(S): All had blood taken for CFTR gene analysis, 33 had scrotal ultrasounds, and 25 had transrectal ultrasounds. MAIN OUTCOME MEASURE(S): The frequency of testicular, seminal vesicle, and ampullae of the vas deferens malformations was compared between subgroups of men with two, one, or no CFTR gene mutations. RESULT(S): None (0 of 21) of the men with at least one CFTR gene mutations had normal ampullae of the vas or seminal vesicles bilaterally. Two (50%) of 4 men with no CFTR gene mutations had normal ampullae of the vas deferens bilaterally, and 50% had normal bilateral seminal vesicles (statistically significantly different). There was no correlation between testicular malformations and CFTR genotype. CONCLUSION(S): This study indicates that the severity of the malformations in the testis is unrelated to the CFTR genotype, whereas the frequency and severity of wolffian duct malformations are related directly to the CFTR genotype.

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60 of men Single mutation 5T /Unknown 13 ⌬F508 /Unknown 8 R117H /Unknown 2 W1282X /Unknown 1 4016insT /Unknown 1 N1303K /Unknown 1 Total 26 Two mutations ⌬F508 /5T 4 ⌬F508 /R117H 2 ⌬F508 /R75Q 1 5T /2183AA3G 1 5T /N1303K 1 5T /G542X 1 R117H /G551A 1 R117H /2184insA 1 A455E /3849ϩ10KbC 1 3T Total 13 No mutations 7 Total no. Login to comment

[hide] CFTR: the nucleotide binding folds regulate the ac... J Gen Physiol. 1996 Jan;107(1):103-19. Wilkinson DJ, Mansoura MK, Watson PY, Smit LS, Collins FS, Dawson DC
CFTR: the nucleotide binding folds regulate the accessibility and stability of the activated state.
J Gen Physiol. 1996 Jan;107(1):103-19., [PMID:8741733]

Abstract [show]
The functional roles of the two nucleotide binding folds, NBF1 and NBF2, in the activation of the cystic fibrosis transmembrane conductance regulator (CFTR) were investigated by measuring the rates of activation and deactivation of CFTR Cl- conductance in Xenopus oocytes. Activation of wild-type CFTR in response to application of forskolin and 3-isobutyl-1-methylxanthine (IBMX) was described by a single exponential. Deactivation after washout of the cocktail consisted of two phases: an initial slow phase, described by a latency, and an exponential decline. Rate analysis of CFTR variants bearing analogous mutations in NBF1 and NBF2 permitted us to characterize amino acid substitutions according to their effects on the accessibility and stability of the active state. Access to the active state was very sensitive to substitutions for the invariant glycine (G551) in NBF1, where mutations to alanine (A), serine (S), or aspartic acid (D) reduced the apparent on rate by more than tenfold. The analogous substitutions in NBF2 (G1349) also reduced the on rate, by twofold to 10-fold, but substantially destabilized the active state as well, as judged by increased deactivation rates. In the putative ATP-binding pocket of either NBF, substitution of alanine, glutamine (Q), or arginine (R) for the invariant lysine (K464 or K1250) reduced the on rate similarly, by two- to fourfold. In contrast, these analogous substitutions produced opposite effects on the deactivation rate. NBF1 mutations destabilized the active state, whereas the analogous substitutions in NBF2 stabilized the active state such that activation was prolonged compared with that seen with wild-type CFTR. Substitution of asparagine (N) for a highly conserved aspartic acid (D572) in the ATP-binding pocket of NBF1 dramatically slowed the on rate and destabilized the active state. In contrast, the analogous substitution in NBF2 (D1370N) did not appreciably affect the on rate and markedly stabilized the active state. These results are consistent with a hypothesis for CFTR activation that invokes the binding and hydrolysis of ATP at NBF1 as a crucial step in activation, while at NBF2, ATP binding enhances access to the active state, but the rate of ATP hydrolysis controls the duration of the active state. The relatively slow time courses for activation and deactivation suggest that slow processes modulate ATP-dependent gating.

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150 The values listed in Table I show that NBF mutations generally reduced the value of (ko,, + ko~), in some cases by more TABLE I Summary ofActivation and DeactivationDatafor Wild-typeCFFR and Mutants of theInvariant Glycinein NBFI ((;,551)orNBF2 (G1349) CFTR Activation Deactivation klon KA (k,,n+ k,,n) (10 ~miu-1 k,,n kos latency *k,,t~ (raM) n (10-~min l) raM-l) (10-3min 1) (10 ~rain-j) n (min) (10-s min-I) wt 0.65 + 0.08 26 664 _+51 118 _+9 588 +-45 76 + 6 20 6.0 _+0.3 88 -+6 16 G551A 3.0 -+0.5*r 6 104 _+5"r 13 _+0.6*r 65 + 3*z 39 -+2* 5 7.7 +_0.5: 70 -+13: 4 G551S 4.7 +-0.5* 5 82 _+6*r 8 -+0.6*: 42 -+3*: 40 -+3*r 10 3.9 +_0.3*** 88 +-6: 6 G551D 9.3 -+0.01" 6 57 _+9*r 4 -+0.6*: 20 -+3*: 37 -+6"r 5 1.8 _+0.2"~ 84 -+10~ 6 G1349A 1.1 + 0.07*: 5 210 _+24"~ 35 -+4*: 172 -+20*: 38 +-4* 4 1.7 _+0.3"~ 184 + 20*: 5 G1349S 3.5 +-0.3* 4 199 _+46*: 23 -+5*: 117 -+27*r 82 -+19+ 6 2.3 _+0.5*+ 144 -+15": 6 G1349D 9.3 + 0.01" 8 114 _+16*++ 8 -+1": 40 +-6*r 74 -+11~ 5 0.6 -+0.1*++ 286 -+37*: 4 Valuesweredetermined as describedin Methods.The symbols(*) and (~) indicatesignificantdifferencesfrom wild-typeCFFRand the analogousmu- tant, respectively(P< 0.05). Login to comment
170 Substitutions for the invariant glycine in either NBF produced similar increases in KA, with the exception of the mutation that CFFR Activation: Roles of NBF1 and NBF2 was conservative with respect to polarity and size, alanine for glycine, which in NBF1 (G551A) produced a nearly fivefold increase in KA but in NBF2 (G1349A) produced less than a twofold increase. Login to comment
176 In NBF1, substitution to alanine (G551A), the most conservative change possible, reduced the relaxation rate by more than sixfold, and the less conservative substitutions to serine (G551S) and aspartic acid (G551D) progressively reduced the relaxation rate. Login to comment
178 Substitutions to serine (G1349S) and aspartic acid (G1349D) produced progressive reductions such that the relaxation rate for the least conservative mutation, G1349D, was about twice that for the comparable mutation in NBF1. Login to comment
221 The most conservative substitution (G551A) did not appreciably alter the latency. Login to comment
223 The progressive destabilization of the active state suggested by the decreased latency seen with the G551S and G551D substitutions was not evident, however, in the subsequent phase of exponential decline. Login to comment
254 sensus B aspartic acid in NBF1 (D572N) produced a profound reduction in the rate of approach to steady state activation (Fig. 4 C); the value of (kon + kof0 was comparable with that seen with the G551A mutation (cf. Tables I and II). Login to comment
256 In contrast, the analogous substitution in NBF2 (D1370N) produced only a modest decrease in (ko, + kor0, evident in Fig. 4 C. The values of the derived parameters (k'o,, ko~) for this slightly hypersensitive mutant, however, suggest that the decrease in KAwas a reflection of a fourfold decrease in ko~, whereas the apparent kon was not significantly different from that of wild-type CFTR. Login to comment
355 The most conservative substitution for the glycine (G551A) dramatically attenuated the on rate but tended to stabilize the active state. Login to comment
152 The values listed in Table I show that NBF mutations generally reduced the value of (ko,, + ko~), in some cases by more TABLE I Summary ofActivation and DeactivationDatafor Wild-typeCFFR and Mutants of theInvariant Glycinein NBFI ((;,551)orNBF2 (G1349) CFTR Activation Deactivation klon KA (k,,n+ k,,n) (10 ~miu-1 k,,n kos latency *k,,t~ (raM) n (10-~min l) raM-l) (10-3min 1) (10 ~rain-j) n (min) (10-s min-I) wt 0.65 + 0.08 26 664 _+51 118 _+9 588 +-45 76 + 6 20 6.0 _+0.3 88 -+6 16 G551A 3.0 -+0.5*r 6 104 _+5"r 13 _+0.6*r 65 + 3*z 39 -+2* 5 7.7 +_0.5: 70 -+13: 4 G551S 4.7 +-0.5* 5 82 _+6*r 8 -+0.6*: 42 -+3*: 40 -+3*r 10 3.9 +_0.3*** 88 +-6: 6 G551D 9.3 -+0.01" 6 57 _+9*r 4 -+0.6*: 20 -+3*: 37 -+6"r 5 1.8 _+0.2"~ 84 -+10~ 6 G1349A 1.1 + 0.07*: 5 210 _+24"~ 35 -+4*: 172 -+20*: 38 +-4* 4 1.7 _+0.3"~ 184 + 20*: 5 G1349S 3.5 +-0.3* 4 199 _+46*: 23 -+5*: 117 -+27*r 82 -+19+ 6 2.3 _+0.5*+ 144 -+15": 6 G1349D 9.3 + 0.01" 8 114 _+16* + + 8 -+1": 40 +-6*r 74 -+11~ 5 0.6 -+0.1* + + 286 -+37*: 4 Valuesweredetermined as describedin Methods.The symbols(*) and (~) indicatesignificantdifferencesfrom wild-typeCFFRand the analogousmu- tant, respectively(P< 0.05). Login to comment
172 Substitutions for the invariant glycine in either NBF produced similar increases in KA, with the exception of the mutation that CFFR Activation: Roles of NBF1 and NBF2 was conservative with respect to polarity and size, alanine for glycine, which in NBF1 (G551A) produced a nearly fivefold increase in KA but in NBF2 (G1349A) produced less than a twofold increase. Login to comment
357 The most conservative substitution for the glycine (G551A) dramatically attenuated the on rate but tended to stabilize the active state. Login to comment

[hide] Functional roles of the nucleotide-binding folds i... Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):9963-7. Smit LS, Wilkinson DJ, Mansoura MK, Collins FS, Dawson DC
Functional roles of the nucleotide-binding folds in the activation of the cystic fibrosis transmembrane conductance regulator.
Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):9963-7., [PMID:7694298]

Abstract [show]
The cystic fibrosis transmembrane conductance regulator (CFTR), a member of the traffic ATPase superfamily, possesses two putative nucleotide-binding folds (NBFs). The NBFs are sufficiently similar that sequence alignment of highly conserved regions can be used to identify analogous residues in the two domains. To determine whether this structural homology is paralleled in function, we compared the activation of chloride conductance by forskolin and 3-isobutyl-1-methylxanthine in Xenopus oocytes expressing CFTRs bearing mutations in NBF1 or NBF2. Mutation of a conserved glycine in the putative linker domain in either NBF produced virtually identical changes in the sensitivity of chloride conductance to activating conditions, and mutation of this site in both NBFs produced additive effects, suggesting that in the two NBFs this region plays a similar and critical role in the activation process. In contrast, amino acid substitutions in the Walker A and B motifs, thought to form an integral part of the nucleotide-binding pockets, produced strikingly different effects in NBF1 and NBF2. Substitutions for the conserved lysine (Walker A) or aspartate (Walker B) in NBF1 resulted in a marked decrease in sensitivity to activation, whereas the same changes in NBF2 produced an increase in sensitivity. These results are consistent with a model for the activation of CFTR in which both NBF1 and NBF2 are required for normal function but in which either the nature or the exact consequences of nucleotide binding differ for the two domains.

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60 For this reason, expression levels for easily activated constructs (wild type, G551A, G1349A, K1250Q, and D1370N) were adjusted by reducing the amount of injected RNA so that the maximum Cl- conductance was similar to that attained by less sensitive constructs. Login to comment
68 G551D, associated with severe CF (35, 36), and G1349D, also a CF mutation (37), both exhibited a dramatic reduction in sensitivity (K1l2 = 2.5 0 0 wt (12) 100 E .E CO) NBF1 A A G551A c O G551S V v G551 D NBF2 (8) A-A G1349A (9) * * G1349S (6) '-V G1349D (4) (6) (8) 0.2 0.5 1 IBMX, mM FIG. 2. Login to comment