ABCMdb

ABCC7 p.His1348Gly

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Publications

PMID: 20421370 [PubMed] Tsai MF et al: "Stable ATP binding mediated by a partial NBD dimer of the CFTR chloride channel."

No. Sentence Comment

150 Of particular note is the H1348G mutation, which increased the time constant of the second phase (Fig. 5 B), suggesting that this mutation actually prolongs the ATP dwell time in NBD1. Login to comment

PMID: 20861014 [PubMed] Tsai MF et al: "Optimization of the degenerated interfacial ATP binding site improves the function of disease-related mutant cystic fibrosis transmembrane conductance regulator (CFTR) channels."

No. Sentence Comment

4 Here, we report a strategy to optimize site 1 for ATP binding by converting two amino acid residues to ABC consensus (i.e. H1348G) or more commonly seen residues in other ABC proteins (i.e. W401Y,W401F). Login to comment
37 Our data suggested that the former is likely the case as the effects of ATP on W401F/G551D channels can be further enhanced by the H1348G mutation in the NBD2 tail subdomain. Login to comment
54 For the W401F/H1348G/G551D channel, the steady-state open probability (Po) in the presence of ATP was estimated by stationary noise analysis of macroscopic currents using the equation ␴2 /i ϭ ͑1 - Po͒I (Eq. 1) where ␴2 is the variance, i is the unitary current amplitude, and I is the amplitude of the steady-state currents. Login to comment
62 This can be readily done for WT and W401F/ H1348G/⌬F508 channels (as seen in Fig. 5). Login to comment
131 We thus converted the His-1348 residue to Gly (H1348G) in the G551D background. Login to comment
133 Moreover, the H1348G mutation further improved the function of W401F/G551D channels so that the application of ATP and PATP increased the basal activity by ϳ25- and ϳ75-fold, respectively (Fig. 3, C and D, and supplemental Fig. S5), further supporting the notion that optimizing ATP binding in site 1 enhances the function of G551D channels. Login to comment
139 A, the S1347G mutation diminished the response of W401F/G551D channels to ATP or PATP. B, incorporating the H1348G mutation into G551D-CFTR conferred responsiveness to ATP. Login to comment
140 C, H1348G enhanced the response of W401F/G551D-CFTR to ATP. Login to comment
145 Single-channel kinetics of G551D channels with W401Y,W401F or H1348G mutations. Login to comment
156 W401F and H1348G Mutations Improve the Function of WT and ⌬F508 Channels-To this point, we have demonstrated that optimizing the interactions of ATP with site 1 components, NBD1 head (W401Y and W401F) and NBD2 tail (H1348G), ameliorates the gating defects of G551D channels, which hold a non-functional site 2. Login to comment
160 Indeed, when W401F and H1348G mutations were engineered into WT channels (Fig. 5A), the mean open time of WT-CFTR was more than quadrupled (Fig. 5C) with the already high Po (ϳ0.4) nearly doubled (ϳ0.78, Fig. 5D). Login to comment
162 In either case, W401F/H1348G mutations did not significantly alter the opening rate (Fig. 5E). Login to comment
165 First, ATP-site 1 interactions of the CFTR channel can be strengthened by introducing mutations in both the head domain of NBD1 (i.e. W401Y,W401F) and the tail domain of NBD2 (i.e. H1348G). Login to comment
180 Effects of W401F/H1348G mutations on WT and ⌬F508 channels. Login to comment
181 A, 30-s single-channel recordings of WT and W401F/H1348G channels exposed to 2.75 mM ATP. Login to comment
182 B, current recordings of ⌬F508 and ⌬F508/W401F/ H1348G channels in the presence of 2.75 mM ATP. Login to comment
191 It is this correlation between the chemical nature of mutations and the stability of the lock-open state that grants us the confidence that W401Y,W401F and H1348G mutations, which prolonged the lock-open duration of WT-CFTR, indeed tighten ATP binding in site 1. Login to comment
199 Unlike WT and ⌬F508 channels, whose Po in the presence or absence of W401F/H1348G mutations (Fig. 5) can be measured with reasonable accuracy, the G551D-containing channels exhibit a Po too low to be derived from single-channel analysis. Login to comment
204 For instance, that the W401F/H1348G/G551D channel has an IATP/IBasal of ϳ25 indicates that the Po for this mutant is ϳ0.1. Login to comment
205 To verify this value, we performed stationary noise analysis for the W401F/ H1348G/G551D mutant, and the resulting Po of 0.09 Ϯ 0.01 (supplemental Fig. S7) did provide some reassurance. Login to comment
209 We have observed that W401F/H1348G mutations in site 1 prolonged the mean open time of WT-CFTR (Fig. 5C). Login to comment
211 Thus, it appears that a stronger ATP binding in site 1, due to the presence of W401F/H1348G mutations, can allosterically tighten the connection between two NBDs around site 2, thereby slowing down channel closure. Login to comment

PMID: 21486785 [PubMed] Jih KY et al: "The most common cystic fibrosis-associated mutation destabilizes the dimeric state of the nucleotide-binding domains of CFTR."

No. Sentence Comment

105 Tight binding of nucleotides in NBD1 prolongs the channel locked-open time In a previous report (Tsai et al. 2010a), we demonstrated that the locked-open time of WT-CFTR induced by PPi is prolonged by replacing ATP with the high affinity ATP analogue N6 -phenylethyl-ATP (P-ATP), or by introducing 'gain-of-function` mutations to the ATP-binding site 1 (mutations which increase the Po of CFTR, such as W401F and H1348G) as the locked-open state reflects an NBD dimer with ATP-binding site 1 occupied by ATP and ATP-binding site 2 by PPi (Tsai et al. 2009). Login to comment
106 In Fig. 3, we show that the gain-of-function mutations W401F and H1348G (Fig. 3A) and P-ATP (Fig. 3B) also prolong the locked-open time of F508-CFTR channels. Login to comment
107 Compared to F508-CFTR, the double mutant W401F/ F508-CFTR ( F508/DM)prolongedthelocked-opentimeby~2-fold, and the triple mutant W401F/H1348G/ F508-CFTR ( F508/TM) by ~4-fold. Login to comment
127 C, summary of PPi locked-open times for each construct ( F508/DM: W401F/ F508-CFTR, F508/TM: W401F/H1348G/ F508-CFTR). Login to comment