ABCC7 p.Tyr1219Cys
| Predicted by SNAP2: | A: D (85%), C: D (85%), D: D (95%), E: D (91%), F: D (80%), G: D (91%), H: D (85%), I: D (85%), K: D (91%), L: D (91%), M: D (91%), N: D (91%), P: D (95%), Q: D (91%), R: D (91%), S: D (91%), T: D (91%), V: D (85%), W: D (80%), |
| Predicted by PROVEAN: | A: D, C: D, D: D, E: D, F: D, G: 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, |
[switch to compact view]
Comments [show]
None has been submitted yet.
[hide] Differences between cystic fibrosis transmembrane ... J Biol Chem. 2000 Sep 22;275(38):29407-12. Berger AL, Welsh MJ
Differences between cystic fibrosis transmembrane conductance regulator and HisP in the interaction with the adenine ring of ATP.
J Biol Chem. 2000 Sep 22;275(38):29407-12., 2000-09-22 [PMID:10893239]
Abstract [show]
The cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channel is a member of the ATP-binding cassette transporter family. The most conserved features of this family are the nucleotide-binding domains. As in other members of this family, these domains bind and hydrolyze ATP; in CFTR this opens and closes the channel pore. The recent crystal structures of related bacterial transporters show that an aromatic residue interacts with the adenine ring of ATP to stabilize nucleotide binding. CFTR contains six aromatic residues that are candidates to coordinate the nucleotide base. We mutated each to cysteine and examined the functional consequences. None of the mutations disrupted channel function or the ability to discriminate between ATP, GTP, and CTP. We also applied [2-(triethylammonium)ethyl] methanethiosulfonate to covalently modify the introduced cysteines. The mutant channels CFTR-F429C, F430C, F433C, and F1232C showed no difference from wild-type CFTR, indicating that either the residues were not accessible to modification, or cysteine modification did not affect function. Although modification inactivated CFTR-Y1219C more rapidly than wild-type CFTR, and inactivation of CFTR-F446C was nucleotide-dependent; failure of these mutations to alter gating suggested that Tyr(1219) and Phe(446) were not important for nucleotide binding. The results suggest that ATP binding may not involve the coordination of the adenine ring by an aromatic residue analogous to that in some bacterial transporters. Taken together with earlier work, this study points to a model in which most of the binding energy for ATP is contributed by the phosphate groups.
Comments [show]
None has been submitted yet.
No. Sentence Comment
9 Although modification inactivated CFTR-Y1219C more rapidly than wild-type CFTR, and inactivation of CFTR-F446C was nucleotide-dependent; failure of these mutations to alter gating suggested that Tyr1219 and Phe446 were not important for nucleotide binding.
X
ABCC7 p.Tyr1219Cys 10893239:9:39
status: NEW77 When expressed in HeLa cells, CFTR-F429C, F430C, F433C, F446C, Y1219C, and F1232C all generated Cl-channel activity in excised, inside-out patches of membrane.
X
ABCC7 p.Tyr1219Cys 10893239:77:63
status: NEW98 CFTR-Y1219C was more rapidly inactivated than wild-type, and CFTR-F1232C showed a tendency for more rapid inactivation (0.05 Ͻ p Ͻ 0.10).
X
ABCC7 p.Tyr1219Cys 10893239:98:5
status: NEW116 In these experiments of modification without ATP, MTSET inhibited CFTR-Y1219C to a greater extent than wild-type CFTR and the other NBD mutants (Fig. 4, A and B).
X
ABCC7 p.Tyr1219Cys 10893239:116:71
status: NEW117 However, because MTSET also inactivated CFTR-Y1219C faster than the other channels in the presence of 1 mM ATP (Table II), we expected a greater inhibition during the timed application of MTSET in the absence of ATP.
X
ABCC7 p.Tyr1219Cys 10893239:117:45
status: NEW118 In the presence of ATP (Fig. 3), a 60-s treatment with MTSET reduced CFTR-Y1219C current to 47 Ϯ 7% (n ϭ 8) of the initial value.
X
ABCC7 p.Tyr1219Cys 10893239:118:74
status: NEW120 When we subtract the measured channel rundown of 37 Ϯ 16% for CFTR-Y1219C (because of the time for washing and the absence of ATP), we expected a current that was 30% of the basal value.
X
ABCC7 p.Tyr1219Cys 10893239:120:73
status: NEW126 Construct Po WT 0.29 Ϯ 0.04 F429C 0.21 Ϯ 0.03 F430C 0.30 Ϯ 0.03 F433C 0.21 Ϯ 0.03 F446C 0.34 Ϯ 0.06 Y1219C 0.26 Ϯ 0.07 F1232C 0.35 Ϯ 0.02 FIG. 2.
X
ABCC7 p.Tyr1219Cys 10893239:126:130
status: NEW142 CFTR variant k M -1 s-1 Wild-type 23.1 Ϯ 6.4 F429C 27.0 Ϯ 16.7 F430C 34.6 Ϯ 22.8 F433C 19.0 Ϯ 5.5 F446C 36.1 Ϯ 15.8 Y1219C 75.5 Ϯ 20.1* F1232C 68.5 Ϯ 19.6 not interfere with the ability of MTSET to inactivate CFTR-Y1219C.
X
ABCC7 p.Tyr1219Cys 10893239:142:146
status: NEWX
ABCC7 p.Tyr1219Cys 10893239:142:257
status: NEW143 To confirm more directly that the rate at which MTSET inactivated CFTR-Y1219C was independent of ATP, we examined the rate of inhibition in the presence of 25 M ATP.
X
ABCC7 p.Tyr1219Cys 10893239:143:71
status: NEW146 If ATP interferes with MTSET modification and inactivation of CFTR-Y1219C, we expect to see a faster inactivation rate constant.
X
ABCC7 p.Tyr1219Cys 10893239:146:67
status: NEW148 The rate of inactivation for CFTR-Y1219C in 25 M ATP (140 Ϯ 81 M -1 s-1 , n ϭ 4) was ϳ4 times the rate of inactivation for wild-type CFTR in 25 M ATP (38 Ϯ 3 M -1 s-1 , n ϭ 3), similar to the ratio of CFTR-Y1219C inactivation to wild-type CFTR inactivation in 1 mM ATP.
X
ABCC7 p.Tyr1219Cys 10893239:148:34
status: NEWX
ABCC7 p.Tyr1219Cys 10893239:148:252
status: NEW150 These results indicate that ATP did not alter MTSET modification of CFTR-Y1219C.
X
ABCC7 p.Tyr1219Cys 10893239:150:73
status: NEW168 Even though CFTR-Y1219C showed a FIG. 4.
X
ABCC7 p.Tyr1219Cys 10893239:168:17
status: NEW179 MTSET inactivation of wild-type CFTR and CFTR-Y1219C-CFTR in 25 M ATP.
X
ABCC7 p.Tyr1219Cys 10893239:179:46
status: NEW