ABCC7 p.Gln353Cys
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PMID: 10220340
[PubMed]
Guinamard R et al: "Arg352 is a major determinant of charge selectivity in the cystic fibrosis transmembrane conductance regulator chloride channel."
No.
Sentence
Comment
101
In cell-attached patches, with 140 mM NaCl in the pipet, the single-channel conductances (in picosiemens) were 6.0 ( 0.3 for the wild type (n ) 7), 5.3 ( 0.3 for R352C (n ) 11), 4.2 ( 0.1 for R352Q (n ) 10), 4.0 ( 0.2 for R352H (n ) 4), and 5.7 ( 0.2 for Q353C (n ) 8).
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ABCC7 p.Gln353Cys 10220340:101:255
status: NEW113 The single-channel conductances (in picosiemens) were 6.2 ( 0.5 for the wild type (n ) 6)2 (Figure 3B), 5.9 ( 0.3 for R352C (n ) 5), 4.2 ( 0.1 for R352Q (n ) 8), and 5.7 ( 0.3 for Q353C (n ) 8).
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ABCC7 p.Gln353Cys 10220340:113:180
status: NEW173 Mutation of the adjacent residue (Q353C) did not alter the reversal potential relative to that of the wild type [Erev ) -51.1 ( 1.7 mV (n)7)], and thus, the Cl- to Na+ permeability ratio was the same as that of the wild type (PCl/ PNa ) 36).
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ABCC7 p.Gln353Cys 10220340:173:34
status: NEW
PMID: 18056267
[PubMed]
Beck EJ et al: "Conformational changes in a pore-lining helix coupled to cystic fibrosis transmembrane conductance regulator channel gating."
No.
Sentence
Comment
100
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.
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ABCC7 p.Gln353Cys 18056267:100:336
status: NEW218 Finally, the MTSEA reactivity was restricted to only five of twenty-six residues in and flanking TM6 in our study, whereas in the earlier study, residues F337C, S341C, I344C, R347C, T351C, R352C, and Q353C were also shown to be accessible to MTS reagents.
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ABCC7 p.Gln353Cys 18056267:218:200
status: NEW
PMID: 19754156
[PubMed]
Alexander C et al: "Cystic fibrosis transmembrane conductance regulator: using differential reactivity toward channel-permeant and channel-impermeant thiol-reactive probes to test a molecular model for the pore."
No.
Sentence
Comment
52
We proposed that these spontaneous changes, that are not seen in either wt or Cys-less CFTR, reflect the coordination of trace Table 1: Percent Change in Oocyte Conductance in the Presence of Compounda MTSETþ MTSES- [Ag(CN)2]- [Au(CN)2]- G330C O O O O I331C -51.6 ( 6.3 -28.9 ( 2.1 -63.1 ( 8.8 O I332C O O O O L333C -58.5 ( 4.8 -47.5 ( 7.6 -83.1 ( 2.2 O R334C þ76.9 ( 11.3 -84.4 ( 1.5 -67.4 ( 7.4 -41.4 ( 3.1 K335C þ10.7 ( 2.4 -37.3 ( 1.5 -29.1 ( 6.4 -54.6 ( 4.7 I336C -54.4 ( 7.9 -75.0 ( 0.6 -81.2 ( 10.5 O F337C O O -89.6 ( 1.9 -90.1 ( 1.3 T338C -37.1 ( 3.3 -85.4 ( 2.5 -75.0 ( 5.2 -88.3 ( 1.6 T339C O O -24.5 ( 7.2 O I340C O O -93.8 ( 1.0 O S341C O O -49.3 ( 4.8 O F342C O O -84.7 ( 1.8 O C343 O O O O I344C O O -66.9 ( 9.3 -77.9 ( 2.1 V345C O O -49.1 ( 9.3 O L346C O O O O R347C O O O O M348C O O -47.9 ( 8.8 -50.1 ( 3.3 A349C O O -19.0 ( 2.0 O V350C O O O O T351C O O O O R352C O O -77.5 ( 1.3 O Q353C O O -72.6 ( 4.5 -76.7 ( 2.8 a Values are means ( SE of three or more oocytes.
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ABCC7 p.Gln353Cys 19754156:52:916
status: NEW281 Note the lack of consistent results reported for F337C, S341C, I344C, R347C, T351C, R352C, and Q353C (shaded).
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ABCC7 p.Gln353Cys 19754156:281:95
status: NEW
PMID: 20805575
[PubMed]
Bai Y et al: "Dual roles of the sixth transmembrane segment of the CFTR chloride channel in gating and permeation."
No.
Sentence
Comment
137
(E) MTSES reduced the current of cysless/Q353C channels.
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ABCC7 p.Gln353Cys 20805575:137:41
status: NEW138 (F) For cysless/Q353C, the treatment of MTSET had little influence on channel function, but prevented the current decrease in response to MTSES, as in E. behavior and conduction properties were recovered after the application of 10 mM DTT.
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ABCC7 p.Gln353Cys 20805575:138:16
status: NEW186 Instead, we will focus on the four other positive hits (i.e., I344C, V345C, M348C, and Q353C).
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ABCC7 p.Gln353Cys 20805575:186:87
status: NEW187 Fig. 9 depicts a sample experiment with the cysless/Q353C construct.
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ABCC7 p.Gln353Cys 20805575:187:52
status: NEW220 This representative recording (among five patches) contains two cysless/ Q353C channels.
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ABCC7 p.Gln353Cys 20805575:220:73
status: NEW
No.
Sentence
Comment
148
Therefore, other sequences must account for the differ-end of the pore and R352C is located closer to the extracellular end of the pore than either T351C or Q353C (32).
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ABCC7 p.Gln353Cys 9922375:148:157
status: NEW
PMID: 9511930
[PubMed]
Akabas MH et al: "Probing the structural and functional domains of the CFTR chloride channel."
No.
Sentence
Comment
87
DIAMETER OF THE CHANNEL LUMEN MTSET+ can penetrate from the extracellular end to react with Q353C (Cheung and Akabas, 1996).
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ABCC7 p.Gln353Cys 9511930:87:92
status: NEW89 Therefore, the channel diameter from the extracellular end to the position of Q353C, flanking the cytoplasmic end of the M6 segment, must be at a minimum 6 A.
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ABCC7 p.Gln353Cys 9511930:89:78
status: NEW122 The major site of charge selectivity appears to be in the region of T351C and Q353C where the anion- to-cation selectivity rises to between 15 and 25 (Fig. 3B) (Cheung and Akabas, 1997).
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ABCC7 p.Gln353Cys 9511930:122:78
status: NEW127 Arg352, which is between T351C and Q353C, appears to be a majordeterminantof the anion selectivity in this region.
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ABCC7 p.Gln353Cys 9511930:127:35
status: NEW130 Moreover, based on our measurements of electrical distance, R352C is closer to the extracellular end of the channel than is T351C or Q353C (Fig. 3A).
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ABCC7 p.Gln353Cys 9511930:130:133
status: NEW131 Thus, ions passing from the extracellular end of the channel would first encounter Arg352, which we infer forms part of the charge-selectivity filter, before they could reach T351C or Q353C, thereby accounting for the greater anion selectivity observed at these residues.
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ABCC7 p.Gln353Cys 9511930:131:184
status: NEW159 The largest electrical distances that we measured, to T351C and Q353C, was only 0.6.
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ABCC7 p.Gln353Cys 9511930:159:64
status: NEW
PMID: 9089437
[PubMed]
Cheung M et al: "Locating the anion-selectivity filter of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel."
No.
Sentence
Comment
107
We did not measure the reaction rate constants for the most extracellular residue, I331C, because we thought that it was unlikely that the reaction rates would be voltage dependent given the absence of voltage dependence at the adjacent, more cytoplasmic residues. We also did not measure the reaction rate constants for the mutants I344C and R347C because, although MTSEAϩ reacted with these residues, MTSES- and MTSETϩ did not react with these k ψ( )( )ln k Ψ 0=( )( ) zFδ RT/( )-ln ψ= t a b l e i Second-order Rate Constants for the Reaction of the MTS Reagents with the Water-exposed Cysteine Mutants k ES (M-1s-1) k EA (M-1s-1) k ET (M-1s-1) mutant -25 mV -50 mV -75 mV -25 mV -50 mV -75 mV -25 mV -50 mV -75 mV L333C 71 Ϯ 3(3) 71 Ϯ 20(2) 71 Ϯ 23(3) 320 Ϯ 89(2) 320 Ϯ 128(2) 333 Ϯ 139(3) 952 Ϯ 136(2) 1,000 Ϯ 350(2) 1,053 Ϯ 443(2) R334C 48 Ϯ 14(2) 48 Ϯ 6(3) 44 Ϯ 8(4) 145 Ϯ 32(2) 163 Ϯ 7(2) 182 Ϯ 21(3) 444 Ϯ 49(2) 454 Ϯ 124(2) 588 Ϯ 95(3) K335C 36 Ϯ 20(3) 23 Ϯ 11(3) 27 Ϯ 16(3) 222 Ϯ 80(3) 121 Ϯ 51(4) 107 Ϯ 30(3) 217 Ϯ 111(3) 235 Ϯ 28(3) 217 Ϯ 95(4) F337C 91 Ϯ 17(2) 80 Ϯ 22(3) 71 Ϯ 20(4) 222 Ϯ 74(2) 222 Ϯ 86(3) 285 Ϯ 81(3) 740 Ϯ 246(3) 740 Ϯ 82(2) 714 Ϯ 51(2) S341C 56 Ϯ 18(3) 56 Ϯ 40(2) 43 Ϯ 12(3) 93 Ϯ 6(3) 110 Ϯ 22(3) 138 Ϯ 34(3) 690 Ϯ 356(3) 556 Ϯ 246(3) 800 Ϯ 224(4) T351C 100 Ϯ 25(5) 57 Ϯ 6(3) 26 Ϯ 9(6) 146 Ϯ 30(4) 195 Ϯ 42(4) 296 Ϯ 18(3) 308 Ϯ 47(10) 392 Ϯ 78(6) 769 Ϯ 89(5) R352C 42 Ϯ 4(3) 26 Ϯ 4(5) 21 Ϯ 6(4) 105 Ϯ 76(3) 137 Ϯ 46(3) 205 Ϯ 58(2) 417 Ϯ 138(4) 800 Ϯ 128(2) 952 Ϯ 408(2) Q353C 125 Ϯ 23(4) 51 Ϯ 12(4) 42 Ϯ 8(4) 83 Ϯ 24(4) 116 Ϯ 42(4) 160 Ϯ 92(3) 189 Ϯ 48(6) 220 Ϯ 48(3) 625 Ϯ 273(4) residues and therefore we could not determine the charge selectivity at these positions.2 The reaction rate constants that we have measured are between 10-and 500-fold slower than the rates of reaction with sulfhydryls in free solution (Table II) (Stauffer and Karlin, 1994).
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ABCC7 p.Gln353Cys 9089437:107:1917
status: NEW154 The distance from the extracellular end to T351C and Q353C is significantly greater than to the other residues (P Ͻ 0.05).
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ABCC7 p.Gln353Cys 9089437:154:53
status: NEW180 By measuring the relative rates of reaction of anionic and cationic MTS reagents with water-exposed cysteines in and flanking the M6 segment we have shown that a major determinant of anion selectivity occurs near the cytoplasmic end of the channel; access of the negatively charged MTSES- to T351C and Q353C is favored over the positively charged MTSETϩ (Fig. 5).
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ABCC7 p.Gln353Cys 9089437:180:302
status: NEW183 Consistent with this, the reaction rate constants for the reaction of MTSES- with T351C and Q353C are larger than the rates with other channel-lining residues (Table II, column 2).
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ABCC7 p.Gln353Cys 9089437:183:92
status: NEW185 The arginine that lies between T351C and Q353C, Arg352, appears to be a major determinant of the anion selectivity in this region; when cysteine is substituted for the arginine at position 352 the selectivity is similar to that observed in the rest of the channel (Fig. 5).
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ABCC7 p.Gln353Cys 9089437:185:41
status: NEW187 Based on our measurements of electrical distance, R352C is closer to the extracellular end of the channel than T351C and Q353C (Fig. 4, see below).
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ABCC7 p.Gln353Cys 9089437:187:121
status: NEW188 Thus, ions passing from the extracellular end of the channel would first encounter Arg352, which we infer forms part of the charge-selectivity filter, before they could reach T351C or Q353C; thereby accounting for the greater anion selectivity we observed at these residues.
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ABCC7 p.Gln353Cys 9089437:188:186
status: NEW191 The increase in the reaction rate constants for MTSES- with the mutants T351C and Q353C (Table II, column 2) is consistent with these residues being near an anion binding site which increases the dwell time of MTSES- in this region of the channel thereby effectively increasing the reaction rate constants.
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ABCC7 p.Gln353Cys 9089437:191:82
status: NEW200 The ability of the cationic MTS reagents to move past the anion-selectivity filter, i.e., to react with T351C and Q353C, is consistent with the lack of ideal anion selectivity that has been reported by others.
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ABCC7 p.Gln353Cys 9089437:200:114
status: NEW210 Note the marked increase in anion selectivity at the residues T351C and Q353C.
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ABCC7 p.Gln353Cys 9089437:210:72
status: NEW
PMID: 8744306
[PubMed]
Cheung M et al: "Identification of cystic fibrosis transmembrane conductance regulator channel-lining residues in and flanking the M6 membrane-spanning segment."
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.
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ABCC7 p.Gln353Cys 8744306:91:452
status: NEW109 Accessibility of substituted cysteines to MTSES- A 1-min application of 10 mM MTSES- significantly inhibited the CFIR-induced currents of 9 of the 24 cysteine-substituted mutants (Fig. 4 A), L333C, R334C, K335C, F337C, S341C, R347C, T351C, R352C, and Q353C.
X
ABCC7 p.Gln353Cys 8744306:109:126
status: NEWX
ABCC7 p.Gln353Cys 8744306:109:251
status: NEW110 An 8-min application of 10mM MTSES- to these mutants did not markedly increase the inhibitory effects, except for the mutant Q353C, in which the extent of inhibition increased (Fig. 4 B); hence the reactions for the other mutants were complete within 1 min.
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ABCC7 p.Gln353Cys 8744306:110:125
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.
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ABCC7 p.Gln353Cys 8744306:90:452
status: NEW108 Accessibility of substituted cysteines to MTSES- A 1-min application of 10 mM MTSES- significantly inhibited the CFIR-induced currents of 9 of the 24 cysteine-substituted mutants (Fig. 4 A), L333C, R334C, K335C, F337C, S341C, R347C, T351C, R352C, and Q353C.
X
ABCC7 p.Gln353Cys 8744306:108:251
status: NEW