ABCMdb

ABCC7 p.Gln220Asn

ClinVar:	c.658C>T , p.Gln220* D , Pathogenic c.659A>G , p.Gln220Arg ? , not provided
CF databases:	c.658C>T , p.Gln220* D , CF-causing c.659A>G , p.Gln220Arg (CFTR1) ? , Found in a patient with CBAVD.
Predicted by SNAP2:	A: N (57%), C: D (59%), D: N (66%), E: N (82%), F: D (75%), G: N (61%), H: N (53%), I: D (71%), K: D (53%), L: D (59%), M: D (71%), N: N (61%), P: D (63%), R: D (63%), S: N (57%), T: N (66%), V: N (57%), W: D (80%), Y: D (71%),
Predicted by PROVEAN:	A: N, C: N, D: N, E: N, F: N, G: N, H: N, I: D, K: N, L: D, M: N, N: N, P: N, R: N, S: N, T: N, V: N, W: N, Y: N,

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Publications
[hide] Role of the extracellular loop in the folding of a... Biochemistry. 2007 Jun 19;46(24):7099-106. Epub 2007 May 22. Wehbi H, Rath A, Glibowicka M, Deber CM
Role of the extracellular loop in the folding of a CFTR transmembrane helical hairpin.
Biochemistry. 2007 Jun 19;46(24):7099-106. Epub 2007 May 22., 2007-06-19 [PMID:17516627]

Abstract [show]
The folding of membrane-spanning domains into their native functional forms depends on interactions between transmembrane (TM) helices joined by covalent loops. However, the importance of these covalent linker regions in mediating the strength of helix-helix associations has not been systematically addressed. Here we examine the potential structural impact of cystic fibrosis-phenotypic mutations in the extracellular loop 2 (ECL2) on interactions between the TM3 and TM4 helices of the cystic fibrosis transmembrane conductance regulator (CFTR) in constructs containing CFTR residues 194-241. When the effects of replacements in ECL2 (including the CF-phenotypic mutants E217G and Q220R) were evaluated in a library of wild-type and mutant TM3-ECL2-TM4 hairpin constructs, we found that SDS-PAGE gel migration rates differed over a range of nearly 40% +/- the wild-type position and that decreased migration rates correlate with increasing hairpin alpha-helical content as measured by circular dichroism spectra in sodium dodecyl sulfate micelles. The decreased mobility of TM3/4 constructs by introduction of non-native residues is interpreted in terms of an elongation or "opening" of the helical hairpin and concomitant destabilization of membrane-based helix-helix interactions. Our results support a role for short loop regions in dictating the stability of membrane protein folds and highlight the interplay between membrane-embedded helix-helix interactions and loop conformation in influencing the structure of membrane proteins.

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No. Sentence Comment
66 We assessed the role of Q220 in TM3/4 folding by constructing two point mutants that could have variable effects on side chain electrostatic interactions: Q220G and Q220N. Login to comment
67 However, neither removal of the polar side chain in the Q220G mutant nor truncation of side chain length by a methylene group in the Q220N mutant effected a significant change in hairpin mobility (Figure 3, Table 1), suggesting that the WT Q220 was not likely to be participating in ECL2 side chain-side chain interactions. Login to comment
83 The Q220E replacement migrates faster than TM3/4 WT, the Q220G and Q220N mutants migrate at equivalent rates to TM3/4 WT, while the Q220W, Q220K, and Q220R substitutions migrate more slowly. Login to comment
87 While Q220G- and Q220N-substituted hairpins migrate at the same rate as WT TM3/4, we undertook to examine this position systematically to determine the molecular determinants of migration rate variation. Login to comment
97 When the changes in TM3/4 WT hairpin migration were compared to changes in overall hairpin helicity, a strong correlation (R ) 0.79) was observed (Figure 5), leading us to propose that increases in non-native R-helix structure within ECL2 might Table 1: Migration Behavior on SDS-PAGE Gels of Single and Double Mutants in the Loop Region of CFTR TM3/4 Constructs % change in apparent MW on SDS-PAGE mutant vs TM3/4 WT in WT loop mutantsa vs TM3/4 V232D in V232D loop mutantsa Pb E217G 6.8 ( 0.7 E217S 11.1 ( 3.4 5.4 ( 1.4 0.056 Q220R 15.2 ( 1.1 Q220G 0.3 ( 0.4 Q220N 2.1 ( 1.3 0.5 ( 0.3 0.108 Q220K 14.1 ( 1.0 Q220W 13.1 ( 1.3 11.5 ( 0.9 0.157 Q220E -11.1 ( 1.1 -4.0 ( 0.3 <0.001 S222G 12.0 ( 2.1 1.1 ( 0.6 0.001 S222E -0.3 ( 2.4 1.3 ( 0.5 0.512 E217G/S222G 12.4 ( 1.9 E217S/S222E 26.1 ( 4.5 averagec 10.4 ( 7.3 4.0 ( 4.2 0.067 a Values are the percentage difference vs TM3/4 WT or TM3/4 V232D migration of SDS-PAGE gels. Login to comment
117 The Q220N and Q220W replacements also affected folding less in the TM3/4 V232D background than in the TM3/4 WT hairpin (Table 1). Login to comment