PMID: 11278813

Hammerle MM, Aleksandrov AA, Riordan JR
Disease-associated mutations in the extracytoplasmic loops of cystic fibrosis transmembrane conductance regulator do not impede biosynthetic processing but impair chloride channel stability.
J Biol Chem. 2001 May 4;276(18):14848-54. Epub 2001 Feb 6., 2001-05-04 [PubMed]
Sentences
No. Mutations Sentence Comment
14 ABCC7 p.Arg117His
X
ABCC7 p.Arg117His 11278813:14:57
status: NEW
view ABCC7 p.Arg117His details
However, one relatively common missense mutation in EL1, R117H, was shown earlier to reduce chloride conductance (2). Login to comment
75 ABCC7 p.Arg117His
X
ABCC7 p.Arg117His 11278813:75:355
status: NEW
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ABCC7 p.Arg117Leu
X
ABCC7 p.Arg117Leu 11278813:75:395
status: NEW
view ABCC7 p.Arg117Leu details
ABCC7 p.Arg117Cys
X
ABCC7 p.Arg117Cys 11278813:75:315
status: NEW
view ABCC7 p.Arg117Cys details
ABCC7 p.Glu217Gly
X
ABCC7 p.Glu217Gly 11278813:75:475
status: NEW
view ABCC7 p.Glu217Gly details
ABCC7 p.Ser108Phe
X
ABCC7 p.Ser108Phe 11278813:75:75
status: NEW
view ABCC7 p.Ser108Phe details
ABCC7 p.Pro111Leu
X
ABCC7 p.Pro111Leu 11278813:75:235
status: NEW
view ABCC7 p.Pro111Leu details
ABCC7 p.Tyr109Cys
X
ABCC7 p.Tyr109Cys 11278813:75:115
status: NEW
view ABCC7 p.Tyr109Cys details
ABCC7 p.Thr908Asn
X
ABCC7 p.Thr908Asn 11278813:75:515
status: NEW
view ABCC7 p.Thr908Asn details
ABCC7 p.Arg117Pro
X
ABCC7 p.Arg117Pro 11278813:75:435
status: NEW
view ABCC7 p.Arg117Pro details
ABCC7 p.Pro1013Leu
X
ABCC7 p.Pro1013Leu 11278813:75:555
status: NEW
view ABCC7 p.Pro1013Leu details
ABCC7 p.Pro111Ala
X
ABCC7 p.Pro111Ala 11278813:75:195
status: NEW
view ABCC7 p.Pro111Ala details
ABCC7 p.Glu116Lys
X
ABCC7 p.Glu116Lys 11278813:75:275
status: NEW
view ABCC7 p.Glu116Lys details
ABCC7 p.Asp110His
X
ABCC7 p.Asp110His 11278813:75:155
status: NEW
view ABCC7 p.Asp110His details
TABLE I Oligonucleotide primers used to generate mutations Mutation Primer S108F GGAAGAATCATAGCTTtCTATGACCCGGATAAC Y109C AGAATCATAGCTTCCTgTGACCCGGATAACAAG D110H ATCATAGCTTCCTATcACCCGGATAACAAGGAG P111A ATAGCTTCCTATGACgCGGATAACAAGGAGGAA P111L ATAGCTTCCTATGACCtGGATAACAAGGAGGAA E116K CCGGATAACAAGGAGaAACGCTCTATCGCGATT R117C GATAACAAGGAGGAAtGCTCTATCGCGATTTAT R117H GATAACAAGGAGGAACaCTCTATCGCGATTTAT R117L GATAACAAGGAGGAACtCTCTATCGCGATTTAT R117P GATAACAAGGAGGAACcCTCTATCGCGATTTAT E217G ATGGGGCTAATCTGGGgGTTGTTACAGGCGTCT T908N TATGCAGTGATTATCAaCAGCACCAGTTCGTAT P1013L GTCGCAGTTTTACAACtCTACATCTTTGTTGCA FIG. 2. Login to comment
80 ABCC7 p.Gly1127Glu
X
ABCC7 p.Gly1127Glu 11278813:80:35
status: NEW
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tion reported in a patient in EL6, G1127E, was not expressed. Login to comment
86 ABCC7 p.Glu116Lys
X
ABCC7 p.Glu116Lys 11278813:86:24
status: NEW
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However, in the case of E116K the surface signal is disproportionally weaker relative to the immunoblot indicating that it could be somewhat impeded in transport to the cell surface. Login to comment
91 ABCC7 p.Ser108Phe
X
ABCC7 p.Ser108Phe 11278813:91:15
status: NEW
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ABCC7 p.Thr908Asn
X
ABCC7 p.Thr908Asn 11278813:91:73
status: NEW
view ABCC7 p.Thr908Asn details
Exceptions are S108F, which matures as efficiently as the wild type, and T908N, which matures even more efficiently. Login to comment
99 ABCC7 p.Glu217Gly
X
ABCC7 p.Glu217Gly 11278813:99:13
status: NEW
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For example, E217G in EL2 reduces the efflux rate greatly. Login to comment
100 ABCC7 p.Pro1013Leu
X
ABCC7 p.Pro1013Leu 11278813:100:19
status: NEW
view ABCC7 p.Pro1013Leu details
On the other hand, P1013L in EL5 exhibits rates as high as wild type. Single Channel Properties of EL-Fig. 4 contains representative recordings of single channels in planar lipid bilayers with which membrane vesicles isolated from cells expressing each of the variants and the wild type were fused. Login to comment
104 ABCC7 p.Ser108Phe
X
ABCC7 p.Ser108Phe 11278813:104:51
status: NEW
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The behavior of the most N-terminal mutant in EL1, S108F, indicates that substitution of the small hydroxyl amino acid with the larger aromatic phenylalanine results in a channel with no stable open state. Login to comment
117 ABCC7 p.Ser108Phe
X
ABCC7 p.Ser108Phe 11278813:117:32
status: NEW
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ABCC7 p.Tyr109Cys
X
ABCC7 p.Tyr109Cys 11278813:117:50
status: NEW
view ABCC7 p.Tyr109Cys details
ABCC7 p.Asp110His
X
ABCC7 p.Asp110His 11278813:117:67
status: NEW
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A, squares, wild type; circles, S108F; triangles, Y109C; diamonds, D110H; crosses, wild type without stimulation. Login to comment
118 ABCC7 p.Pro111Leu
X
ABCC7 p.Pro111Leu 11278813:118:28
status: NEW
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ABCC7 p.Pro111Ala
X
ABCC7 p.Pro111Ala 11278813:118:12
status: NEW
view ABCC7 p.Pro111Ala details
ABCC7 p.Glu116Lys
X
ABCC7 p.Glu116Lys 11278813:118:46
status: NEW
view ABCC7 p.Glu116Lys details
B, squares, P111A; circles, P111L; triangles, E116K. Login to comment
119 ABCC7 p.Arg117His
X
ABCC7 p.Arg117His 11278813:119:28
status: NEW
view ABCC7 p.Arg117His details
ABCC7 p.Arg117Leu
X
ABCC7 p.Arg117Leu 11278813:119:46
status: NEW
view ABCC7 p.Arg117Leu details
ABCC7 p.Arg117Cys
X
ABCC7 p.Arg117Cys 11278813:119:12
status: NEW
view ABCC7 p.Arg117Cys details
ABCC7 p.Arg117Pro
X
ABCC7 p.Arg117Pro 11278813:119:63
status: NEW
view ABCC7 p.Arg117Pro details
C, squares, R117C; circles, R117H; triangles, R117L; diamonds, R117P. Login to comment
120 ABCC7 p.Glu217Gly
X
ABCC7 p.Glu217Gly 11278813:120:12
status: NEW
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ABCC7 p.Thr908Asn
X
ABCC7 p.Thr908Asn 11278813:120:28
status: NEW
view ABCC7 p.Thr908Asn details
ABCC7 p.Pro1013Leu
X
ABCC7 p.Pro1013Leu 11278813:120:46
status: NEW
view ABCC7 p.Pro1013Leu details
D, squares, E217G; circles, T908N; triangles, P1013L. Login to comment
122 ABCC7 p.Ser108Phe
X
ABCC7 p.Ser108Phe 11278813:122:188
status: NEW
view ABCC7 p.Ser108Phe details
ABCC7 p.Tyr109Cys
X
ABCC7 p.Tyr109Cys 11278813:122:95
status: NEW
view ABCC7 p.Tyr109Cys details
The substitution of the aromatic tyrosine in the adjacent position by the small thiol residue (Y109C) also results in a very unstable open state, but the tracing is different from that of S108F. Login to comment
126 ABCC7 p.Asp110His
X
ABCC7 p.Asp110His 11278813:126:67
status: NEW
view ABCC7 p.Asp110His details
Replacing the aspartate residue in the next position by histidine (D110H) resulted in a much more normal looking channel with a major mean conductance of about 8.5 pS. Login to comment
130 ABCC7 p.Pro111Leu
X
ABCC7 p.Pro111Leu 11278813:130:0
status: NEW
view ABCC7 p.Pro111Leu details
P111L appears very much like wild type with a mean conductance of 10.3 pS. Login to comment
134 ABCC7 p.Pro111Ala
X
ABCC7 p.Pro111Ala 11278813:134:72
status: NEW
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Substitution of the proline with the smaller aliphatic alanine residue (P111A) produced a channel with much more rapid kinetics and a reduced conductance of 9.2 pS even at 21 °C (Fig. 4). Login to comment
135 ABCC7 p.Glu116Lys
X
ABCC7 p.Glu116Lys 11278813:135:94
status: NEW
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The mutation five residues further in the C-terminal direction resulted in a charge reversal (E116K). Login to comment
136 ABCC7 p.Ser108Phe
X
ABCC7 p.Ser108Phe 11278813:136:49
status: NEW
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This mutant displays properties quite similar to S108F; channel openings occur but cannot be maintained. Login to comment
138 ABCC7 p.Arg117His
X
ABCC7 p.Arg117His 11278813:138:4
status: NEW
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The R117H mutant was reported previously to exhibit a small reduction in both current amplitude and open time (2). Login to comment
142 ABCC7 p.Arg117Leu
X
ABCC7 p.Arg117Leu 11278813:142:15
status: NEW
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ABCC7 p.Arg117Cys
X
ABCC7 p.Arg117Cys 11278813:142:5
status: NEW
view ABCC7 p.Arg117Cys details
ABCC7 p.Ser108Phe
X
ABCC7 p.Ser108Phe 11278813:142:56
status: NEW
view ABCC7 p.Ser108Phe details
ABCC7 p.Glu116Lys
X
ABCC7 p.Glu116Lys 11278813:142:66
status: NEW
view ABCC7 p.Glu116Lys details
Both R117C and R117L had very unstable open states like S108F and E116K with the cysteine substitution able to maintain openings FIG. 4. Login to comment
148 ABCC7 p.Arg117Pro
X
ABCC7 p.Arg117Pro 11278813:148:0
status: NEW
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R117P also displayed only transient openings. Login to comment
149 ABCC7 p.Arg117His
X
ABCC7 p.Arg117His 11278813:149:23
status: NEW
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Thus the more frequent R117H mutation seems to compromise open-channel structure less than the other three substitutions at this position. Login to comment
152 ABCC7 p.Glu217Gly
X
ABCC7 p.Glu217Gly 11278813:152:42
status: NEW
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Strikingly, the single missense mutation (E217G) in the short EL2 also results in channels with only transient rather than stable openings, perhaps implying that this loop is involved also in stabilizing open pore structure. Login to comment
153 ABCC7 p.Thr908Asn
X
ABCC7 p.Thr908Asn 11278813:153:4
status: NEW
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The T908N mutation in EL4, the second of the two larger extracytoplasmic loops of CFTR, has a somewhat similar effect on the single channel behavior as those in EL1 that cause a very unstable open state. Login to comment
156 ABCC7 p.Thr908Asn
X
ABCC7 p.Thr908Asn 11278813:156:15
status: NEW
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As seen in the T908N trace, there are in fact relatively long openings with several brief closings within to generate burstlike behavior. Login to comment
162 ABCC7 p.Pro1013Leu
X
ABCC7 p.Pro1013Leu 11278813:162:46
status: NEW
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The final disease-associated mutant analyzed, P1013L, in the fifth exterior loop exhibited robust wild-type-like behavior, and consistent with its 36 Cl efflux ability, it has a Po of 0.5, higher than the wild type. Login to comment
171 ABCC7 p.Gly551Asp
X
ABCC7 p.Gly551Asp 11278813:171:50
status: NEW
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ABCC7 p.Arg117His
X
ABCC7 p.Arg117His 11278813:171:180
status: NEW
view ABCC7 p.Arg117His details
ABCC7 p.Arg117Leu
X
ABCC7 p.Arg117Leu 11278813:171:192
status: NEW
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ABCC7 p.Arg117Cys
X
ABCC7 p.Arg117Cys 11278813:171:186
status: NEW
view ABCC7 p.Arg117Cys details
ABCC7 p.Glu217Gly
X
ABCC7 p.Glu217Gly 11278813:171:204
status: NEW
view ABCC7 p.Glu217Gly details
ABCC7 p.Ser108Phe
X
ABCC7 p.Ser108Phe 11278813:171:144
status: NEW
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ABCC7 p.Pro111Leu
X
ABCC7 p.Pro111Leu 11278813:171:162
status: NEW
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ABCC7 p.Tyr109Cys
X
ABCC7 p.Tyr109Cys 11278813:171:150
status: NEW
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ABCC7 p.Thr908Asn
X
ABCC7 p.Thr908Asn 11278813:171:210
status: NEW
view ABCC7 p.Thr908Asn details
ABCC7 p.Arg117Pro
X
ABCC7 p.Arg117Pro 11278813:171:198
status: NEW
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ABCC7 p.Pro1013Leu
X
ABCC7 p.Pro1013Leu 11278813:171:216
status: NEW
view ABCC7 p.Pro1013Leu details
ABCC7 p.Pro111Ala
X
ABCC7 p.Pro111Ala 11278813:171:168
status: NEW
view ABCC7 p.Pro111Ala details
ABCC7 p.Glu116Lys
X
ABCC7 p.Glu116Lys 11278813:171:174
status: NEW
view ABCC7 p.Glu116Lys details
ABCC7 p.Asp110His
X
ABCC7 p.Asp110His 11278813:171:156
status: NEW
view ABCC7 p.Asp110His details
For example a nucleotide binding domain mutation, G551D, precludes virtually all TABLE II Relative charge transport capacity of mutants Mutants S108F Y109C D110H P111L P111A E116K R117H R117C R117L R117P E217G T908N P1013L Imutant/Iwt 100% 11 15 27 173 105 12 80 27 5 11 10 48 170 FIG. 5. Login to comment
176 ABCC7 p.Arg117His
X
ABCC7 p.Arg117His 11278813:176:88
status: NEW
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Of missense mutations identified in codons for residues in extracytoplasmic loops, only R117H, which occurs relatively frequently in patients, has been studied in any detail (2). Login to comment
183 ABCC7 p.Glu116Lys
X
ABCC7 p.Glu116Lys 11278813:183:30
status: NEW
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The charge-reversal mutation, E116K, at the immediately preceding position had a remarkably similar effect to that of the most perturbing substitutions of Arg-117. Login to comment
186 ABCC7 p.Pro111Leu
X
ABCC7 p.Pro111Leu 11278813:186:4
status: NEW
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The P111L variant in contrast to all the other EL1 mutants differed from wild type only at elevated temperatures. Login to comment
188 ABCC7 p.Glu217Gly
X
ABCC7 p.Glu217Gly 11278813:188:4
status: NEW
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The E217G mutant in EL2 had a similar effect. Login to comment
190 ABCC7 p.Thr908Asn
X
ABCC7 p.Thr908Asn 11278813:190:4
status: NEW
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The T908N mutation in EL4, which is novel in that it results in the introduction of an additional consensus site for N-glycosylation that is used (17), also alters channel gating. Login to comment
191 ABCC7 p.Pro1013Leu
X
ABCC7 p.Pro1013Leu 11278813:191:18
status: NEW
view ABCC7 p.Pro1013Leu details
Thus although the P1013L change in EL5 did not seem to compromise channel activity except at higher temperatures, all of the others analyzed seriously detracted from the ability of CFTR to sustain a stable open pore. Login to comment