ABCC7 p.Lys329Cys
| Predicted by SNAP2: | A: N (53%), C: D (66%), D: N (57%), E: D (59%), F: D (66%), G: N (72%), H: N (82%), I: D (71%), L: D (53%), M: D (66%), N: N (82%), P: D (66%), Q: N (72%), R: N (66%), S: N (72%), T: N (72%), V: D (53%), W: D (85%), Y: D (59%), |
| Predicted by PROVEAN: | A: N, C: N, D: N, E: N, F: N, G: N, H: N, I: N, L: N, M: N, N: N, P: N, Q: N, R: N, S: N, T: N, V: N, W: D, Y: N, |
[switch to compact view]
Comments [show]
None has been submitted yet.
[hide] Conformational changes in a pore-lining helix coup... J Biol Chem. 2008 Feb 22;283(8):4957-66. Epub 2007 Dec 3. Beck EJ, Yang Y, Yaemsiri S, Raghuram V
Conformational changes in a pore-lining helix coupled to cystic fibrosis transmembrane conductance regulator channel gating.
J Biol Chem. 2008 Feb 22;283(8):4957-66. Epub 2007 Dec 3., 2008-02-22 [PMID:18056267]
Abstract [show]
Cystic fibrosis transmembrane conductance regulator (CFTR), the protein dysfunctional in cystic fibrosis, is unique among ATP-binding cassette transporters in that it functions as an ion channel. In CFTR, ATP binding opens the channel, and its subsequent hydrolysis causes channel closure. We studied the conformational changes in the pore-lining sixth transmembrane segment upon ATP binding by measuring state-dependent changes in accessibility of substituted cysteines to methanethiosulfonate reagents. Modification rates of three residues (resides 331, 333, and 335) near the extracellular side were 10-1000-fold slower in the open state than in the closed state. Introduction of a charged residue by chemical modification at two of these positions (resides 331 and 333) affected CFTR single-channel gating. In contrast, modifications of pore-lining residues 334 and 338 were not state-dependent. Our results suggest that ATP binding induces a modest conformational change in the sixth transmembrane segment, and this conformational change is coupled to the gating mechanism that regulates ion conduction. These results may establish a structural basis of gating involving the dynamic rearrangement of transmembrane domains necessary for vectorial transport of substrates in ATP-binding cassette transporters.
Comments [show]
None has been submitted yet.
No. Sentence Comment
94 Cd2ϩ had a small but significant potentiating effect on K329C channels, but MTSEA, which by itself is without any significant effect, was able to abolish the potentiating effect of Cd2ϩ (supplemental Fig. S2).
X
ABCC7 p.Lys329Cys 18056267:94:62
status: NEW100 The oocytes 750 500 250 0 µS 180012006000 s IBMX MTSEA Cd 2+ DTT 200 100 0 µS 180012006000 s IBMX DTT Cd 2+ MTSEA A B C -100 -80 -60 -40 -20 0 20 40 % Change in conductance Y325C A326C L327C I328C K329C G330C I331C I332C L333C R334C K335C I336C F337C T338C T339C I340C S341C F342C WT I344C V345C R347C M348C A349C V350C T351C Q353C * * * * * Cd 2+ 1mM MTSEA 1mM D FIGURE 1.
X
ABCC7 p.Lys329Cys 18056267:100:207
status: NEW213 We believe that K329C, whose whole cell conductance was stimulated by Cd2ϩ , is an example of one such residue that reacts with MTSEA, but the modification is without effect on channel function (supplemental Fig. S2).
X
ABCC7 p.Lys329Cys 18056267:213:16
status: NEW[hide] Identification of cystic fibrosis transmembrane co... Biophys J. 1996 Jun;70(6):2688-95. Cheung M, Akabas MH
Identification of cystic fibrosis transmembrane conductance regulator channel-lining residues in and flanking the M6 membrane-spanning segment.
Biophys J. 1996 Jun;70(6):2688-95., [PMID:8744306]
Abstract [show]
The cystic fibrosis transmembrane conductance regulator (CFTR) forms a chloride channel that is regulated by phosphorylation and ATP binding. Work by others suggested that some residues in the sixth transmembrane segment (M6) might be exposed in the channel and play a role in ion conduction and selectivity. To identify the residues in M6 that are exposed in the channel and the secondary structure of M6, we used the substituted cysteine accessibility method. We mutated to cysteine, one at a time, 24 consecutive residues in and flanking the M6 segment and expressed these mutants in Xenopus oocytes. We determined the accessibility of the engineered cysteines to charged, lipophobic, sulfhydryl-specific methanethiosulfonate (MTS) reagents applied extracellularly. The cysteines substituted for Ile331, Leu333, Arg334, Lys335, Phe337, Ser341, Ile344, Arg347, Thr351, Arg352, and Gln353 reacted with the MTS reagents, and we infer that they are exposed on the water-accessible surface of the protein. From the pattern of the exposed residues we infer that the secondary structure of the M6 segment includes both alpha-helical and extended regions. The diameter of the channel from the extracellular end to the level of Gln353 must be at least 6 A to allow the MTS reagents to reach these residues.
Comments [show]
None has been submitted yet.
No. Sentence Comment
91 Effects of MTS reagents on wild-type cysteines RESULTS in CFTR To identify the residues in and flanking the M6 membrane-spanning segment that are on the water-exposed surface of As reported previously (Akabas et al., 1994b), extracellular applications of the MTS reagents to Xenopus oocytes ex- L2j K329C L. _J *G330C 1331C 1332C L333C R334C K335C 1336C F337C T338C T339C 1340C S341C T342C C343,WT 1344C V345C L346C R347C M348C A349C V350C T351C R352C Q353C 0 2000 4000 6000 8000 0 25 50 PEAK CURRENTS (nA) TIME TO REACH PLATEAU (min) FIGURE 2 Peak CFTR-induced currents and time to reach the plateau current after stimulation with cAMP-activating reagents for 24 cysteine-substitution mutants and wild-type CFTR.
X
ABCC7 p.Lys329Cys 8744306:91:299
status: NEW90 Effects of MTS reagents on wild-type cysteines RESULTS in CFTR To identify the residues in and flanking the M6 membrane-spanning segment that are on the water-exposed surface of As reported previously (Akabas et al., 1994b), extracellular applications of the MTS reagents to Xenopus oocytes ex- L2j K329C L. _J *G330C 1331C 1332C L333C R334C K335C 1336C F337C T338C T339C 1340C S341C T342C C343,WT 1344C V345C L346C R347C M348C A349C V350C T351C R352C Q353C 0 2000 4000 6000 8000 0 25 50 PEAK CURRENTS (nA) TIME TO REACH PLATEAU (min) FIGURE 2 Peak CFTR-induced currents and time to reach the plateau current after stimulation with cAMP-activating reagents for 24 cysteine-substitution mutants and wild-type CFTR.
X
ABCC7 p.Lys329Cys 8744306:90:299
status: NEW