ABCMdb

ABCC7 p.Thr1142Ala

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Publications

PMID: 16442101 [PubMed] Frelet A et al: "Insight in eukaryotic ABC transporter function by mutation analysis."

No. Sentence Comment

376 Gupta et al. [178] have examined the effect of T1134A, M1137A, N1138A, S1141A and T1142A and found that, in contrast to those in TM6, mutations in TM12 have little effect on channel permeation properties, suggesting that TM6 and TM12 make highly asymmetric contributions to the pore. Login to comment
377 N1138A and T1142A significantly blocked the channel by indirectly reducing interaction between ClÀ ions and glibenclamide within the pore [179]. Login to comment

PMID: 11380256 [PubMed] Gupta J et al: "Asymmetric structure of the cystic fibrosis transmembrane conductance regulator chloride channel pore suggested by mutagenesis of the twelfth transmembrane region."

No. Sentence Comment

79 Five alanine-substitution mutations in TM12 were constructed: T1134A, M1137A, N1138A, S1141A, and T1142A (Figure 1B). Login to comment
92 Block of N1138A (Figure 2B), as well as T1134A, M1137A, and T1142A (data not shown), appeared somewhat weaker than for wild-type, whereas block of S1141A actually appeared stronger than for wild-type (Figure 2B). Login to comment
95 Unitary Current Properties. Expression of wild-type, T1134A, M1137A, N1138A, and T1142A-CFTR in CHO cells led to the appearance of unitary PKAand ATP-dependent Cl-channel currents in excised membrane patches (Figure 3A). Login to comment
96 As described above, S1141A could not be expressed in CHO cells; however, unitary S1141A-CFTR Table 1: Relative Anion Permeabilities for Wild-Type and Mutant CFTRa wild-type T1134A M1137A N1138A S1141A T1142A Cl 1.00 ( 0.01 (10) 1.00 ( 0.06 (5) 1.00 ( 0.03 (6) 1.00 ( 0.02 (4) 1.00 ( 0.02 (5) 1.00 ( 0.04 (7) Br 1.37 ( 0.07 (8) 1.42 ( 0.03 (4) 1.61 ( 0.02 (5)* 1.32 ( 0.08 (7) 1.54 ( 0.05 (4) 1.44 ( 0.04 (5) I 0.83 ( 0.03 (6) 0.85 ( 0.04 (4) 0.88 ( 0.02 (3) 0.83 ( 0.03 (4) 0.78 ( 0.01 (3) 0.86 ( 0.03 (3) F 0.103 ( 0.007 (9) 0.077 ( 0.008 (3) 0.107 ( 0.014 (3) 0.089 ( 0.005 (4) 0.053 ( 0.003 (7)** 0.094 ( 0.007 (3) SCN 3.55 ( 0.26 (7) 3.70 ( 0.33 (4) 3.58 ( 0.14 (5) 3.53 ( 0.21 (6) 3.37 ( 0.35 (3) 3.64 ( 0.22 (5) NO3 1.58 ( 0.04 (10) 1.62 ( 0.05 (4) 1.60 ( 0.04 (3) 1.58 ( 0.05 (6) 1.57 ( 0.03 (3) 1.64 ( 0.07 (3) ClO4 0.25 ( 0.01 (8) 0.22 ( 0.00 (3) 0.29 ( 0.02 (3) 0.44 ( 0.05 (5)** 0.22 ( 0.01 (3) 0.30 ( 0.03 (5) formate 0.24 ( 0.01 (9) 0.26 ( 0.01 (3) 0.27 ( 0.03 (3) 0.27 ( 0.01 (4) 0.26 ( 0.02 (4) 0.25 ( 0.03 (3) acetate 0.091 ( 0.003 (10) 0.102 ( 0.021 (3) 0.103 ( 0.011 (3) 0.087 ( 0.022 (3) 0.066 ( 0.007 (4)* 0.086 ( 0.009 (3) a Relative permeabilities (PX/PCl) for different anions present in the intracellular solution under biionic conditions were calculated from macroscopic current reversal potentials according to eq 1 (see Experimental Procedures), as described in detail previously (16, 20, 36). Login to comment
109 Block by SCN- appeared somewhat weakened in N1138A (left), as well as in T1134A, M1137A and T1142A (data not shown, but see Figure 5B), and strengthened in S1141A (right). Login to comment
144 However, for N1138A, S1141A, and T1142A, different degrees of inhibition were observed, suggesting that SCN- has some other effect on these mutants which is both distinct from the observed reduction in unitary current amplitude and absent in wild-type CFTR. Since the single channel results report the isolated effects of SCN- on unitary current amplitude, which is presumably determined by the relative tightness of SCN- binding within the pore, we believe that the reduced apparent SCN- binding affinity suggested by the single channel results with T1134A, M1137A, and S1141A is the best reporter of altered pore function in these mutants. Login to comment
176 This effect did not appear to be a nonspecific result of mutagenesis within TM12, as SCN- block was unaltered in both N1138A and T1142A. Login to comment
185 A similar trend was observed in N1138A and T1142A (Figure 4B), although in these cases the reduction in mean PO was not statistically significant (0.05 < P < 0.15 in both cases, two-tailed t-test). Login to comment

PMID: 11889571 [PubMed] Gupta J et al: "Point mutations in the pore region directly or indirectly affect glibenclamide block of the CFTR chloride channel."

No. Sentence Comment

5 Interestingly, two mutations in the 12th transmembrane region (N1138A and T1142A) significantly strengthened block. Login to comment
63 While block of the TM12 mutants S1141A (Fig. 1) and T1134A and M1137A (data not shown) was indistinguishable from wild-type, block was significantly weakened in the TM6 mutants F337A and T338A, and significantly strengthened in the TM12 mutants N1138A and T1142A (Fig. 1). Login to comment
69 Mean fraction of control current remaining following addition of 60 µM glibenclamide (I/I0) is shown as a function of voltage for wild-type (q), T338A (s), N1138A (s), F337A (ss) and T1142A (xx). Login to comment
70 Mean of data from 5-10 patches, fitted by Eq. according to the mean parameters shown in Fig. 3 rent remaining following addition of glibenclamide (I/I0) was significantly reduced at all voltages in N1138A and T1142A (P<0.05), and significantly increased in F337A and T338A at negative membrane potentials (P<0.05). Login to comment
75 Consistent with the results shown in Fig. 2, Kd(0) was significantly increased in F337A and T338A, and significantly decreased in N1138A and T1142A (Fig. 3A). Login to comment
84 In contrast, reducing extracellular Cl-concentration from 154 mM to 10 mM had no significant effect on apparent glibenclamide affinity in the TM12 mutants N1138A (Fig. 5A) or T1142A (Figs. 4, 5A). Login to comment
85 As a result, the effect of these two mutations on Kd(0) was dependent on the extracellular Cl-concentration: in N1138A, Kd(0) was reduced to 35% of the wild-type value with 154 mM Cl- but only to 61% of wild-type with 10 mM Cl-, while in T1142A, Kd(0) was 40% of wild-type with 154 mM Cl- and 50% with 10 mM Cl-. Login to comment
98 A Example I-V relationships for wild-type (left), T338A (center) and T1142A (right), recorded with 10 mM extracellular Clas described in Materials and methods, before (Ctrl) and immediately following addition of 60 µM glibenclamide (+Glib) to the intracellular solution. B Mean fraction of control current remaining following addition of glibenclamide, with 154 mM (q) or 10 mM (qq) extracellular Cl-, for wild-type (left), T338A (center) and T1142A (right). Login to comment
115 In contrast to the effects of these mutations in TM6, two mutations in TM12 (N1138A and T1142A) caused a significant increase in the apparent affinity of glibenclamide block (Figs. 2, 3). Login to comment
120 Two kinds of interactions may underlie the ability of extracellular Cl- ions to weaken glibenclamide block of wild-type, but not N1138A or T1142A CFTR. Login to comment
122 In this case, N1138A and T1142A might alter the structure of a binding site for both Cl- and glibenclamide, reducing its affinity for Cl- ions and thereby indirectly increasing glibenclamide occupancy of the pore. Login to comment
132 However, it is interesting to note that both suggest that: (1) the mutations N1138A and T1142A can increase the apparent affinity of glibenclamide block without directly affecting the interaction between glibenclamide and the pore walls, and (2) these two mutations both interfere with Cl-binding within the pore. Login to comment
134 Furthermore, the suggestion that TM12 residues N1138 and T1142 contribute to Cl-binding within the pore conflicts with our previous finding that the mutations N1138A and T1142A have no effect on unitary Cl-conductance [10]. Login to comment

PMID: 18597042 [PubMed] Mornon JP et al: "Atomic model of human cystic fibrosis transmembrane conductance regulator: membrane-spanning domains and coupling interfaces."

No. Sentence Comment

207 Finally, two mutations of TM12 residues that also project towards the pore (N1138A and T1142A, Fig. 3B) were reported to significantly strengthen the glibenclamide block and to abolish its dependence on the extracellular Cl-concentration [69], suggesting that these mutations may alter interactions between glibenclamide and Cl- ions within the pore. Login to comment

PMID: 22160394 [PubMed] Cui G et al: "Differential contribution of TM6 and TM12 to the pore of CFTR identified by three sulfonylurea-based blockers."

No. Sentence Comment

150 Surprisingly, nine mutations of TM12, including N1138A, M1140A, T1142A, V1147A, N1148A, S1149A, S1150A, I1151A, and D1152A, exhibited significantly altered block by Glyb; the pattern was not consistent with either α-helix or β-strand secondary structure along the full length of the region studied. Login to comment
163 Effects on time-dependent block by mutations R334A and K335A Fractional block by Glip200 μM V1153A D1152A I1151A S1150A S1149A N1148A V1147A A1146S W1145A Q1144A L1143A T1142A S1141A M1140A I1139A N1138A M1137A A1136S L1135A T1134A WT 0 0.2 0.4 0.6 0.8 * ** ** ** ** ** ** * V1153A D1152A I1151A S1150A S1149A N1148A V1147A A1146S W1145A Q1144A L1143A T1142A S1141A M1140A I1139A N1138A M1137A A1136S L1135A T1134A WT 0 0.2 0.4 0.6 0.8 1.0 * * * * * ** ** ** ** Fractional block by Glyb50 μM Fig. 4 Alanine-scanning in TM12 to identify amino acids that interact with Glyb and Glip. Login to comment
225 Since these two drugs differ only at the non-sulfonylurea end of the molecular structure, it seems reasonable to conclude that it is this end of Glyb ΔFractionalblockfrom -20mVto-100mV ΔFractionalblockfrom -20mVto-100mV 0.0 0.1 0.2 0.3 0.4 0.5 * * #Glyb 0.0 0.1 0.2 0.3 0.4 0.5 * * * * * ## Glyb Vm(mV) -100 0.2 0.4 0.6 0.8 WT-Glyb50 μM F337A WT-Glyb100 μM T1142A Fractionalblock by50μMGlyb b a -200-40-60-80 Fig. 8 Voltage-dependent block of WT-CFTR and some important mutants in TM6 and TM12. Login to comment
226 a Voltage-dependence of block of WT-CFTR, F337A-CFTR, and T1142A-CFTR by 50 μM Glyb, and WT-CFTR by 100 μM Glyb, at VM=-100 mV to -20 mV. Fractional block was calculated from the steady-state currents at each potential. Login to comment