ABCC8 p.Leu183Ala
| Predicted by SNAP2: | A: N (82%), C: N (93%), D: N (53%), E: N (53%), F: N (93%), G: N (61%), H: N (87%), I: N (93%), K: D (53%), M: N (97%), N: N (57%), P: N (53%), Q: N (72%), R: N (66%), S: N (72%), T: N (93%), V: N (97%), W: N (61%), Y: N (57%), |
| Predicted by PROVEAN: | A: N, C: N, D: D, E: D, F: N, G: D, H: D, I: N, K: D, M: N, N: D, P: D, Q: D, R: D, S: N, T: N, V: N, W: D, Y: N, |
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[hide] Structural model of ligand-G protein-coupled recep... J Biol Chem. 2011 Sep 9;286(36):31661-75. Epub 2011 Jun 17. Marquer C, Fruchart-Gaillard C, Letellier G, Marcon E, Mourier G, Zinn-Justin S, Menez A, Servent D, Gilquin B
Structural model of ligand-G protein-coupled receptor (GPCR) complex based on experimental double mutant cycle data: MT7 snake toxin bound to dimeric hM1 muscarinic receptor.
J Biol Chem. 2011 Sep 9;286(36):31661-75. Epub 2011 Jun 17., [PMID:21685390]
Abstract [show]
The snake toxin MT7 is a potent and specific allosteric modulator of the human M1 muscarinic receptor (hM1). We previously characterized by mutagenesis experiments the functional determinants of the MT7-hM1 receptor interaction (Fruchart-Gaillard, C., Mourier, G., Marquer, C., Stura, E., Birdsall, N. J., and Servent, D. (2008) Mol. Pharmacol. 74, 1554-1563) and more recently collected evidence indicating that MT7 may bind to a dimeric form of hM1 (Marquer, C., Fruchart-Gaillard, C., Mourier, G., Grandjean, O., Girard, E., le Maire, M., Brown, S., and Servent, D. (2010) Biol. Cell 102, 409-420). To structurally characterize the MT7-hM1 complex, we adopted a strategy combining double mutant cycle experiments and molecular modeling calculations. First, thirty-three ligand-receptor proximities were identified from the analysis of sixty-one double mutant binding affinities. Several toxin residues that are more than 25 A apart still contact the same residues on the receptor. As a consequence, attempts to satisfy all the restraints by docking the toxin onto a single receptor failed. The toxin was then positioned onto two receptors during five independent flexible docking simulations. The different possible ligand and receptor extracellular loop conformations were described by performing simulations in explicit solvent. All the docking calculations converged to the same conformation of the MT7-hM1 dimer complex, satisfying the experimental restraints and in which (i) the toxin interacts with the extracellular side of the receptor, (ii) the tips of MT7 loops II and III contact one hM1 protomer, whereas the tip of loop I binds to the other protomer, and (iii) the hM1 dimeric interface involves the transmembrane helices TM6 and TM7. These results structurally support the high affinity and selectivity of the MT7-hM1 interaction and highlight the atypical mode of interaction of this allosteric ligand on its G protein-coupled receptor target.
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No. Sentence Comment
156 hM1 mutant Region pKi NMS pKX MT7 KX(mut)/KX(WT) WT 10.22 Ϯ 0.04 10.47 Ϯ 0.04 H90A E1 10.20 Ϯ 0.09 10.66 Ϯ 0.20 0.6 W91A E1 9.23 ؎ 0.33 9.30 ؎ 0.06 14.9 W101A TM3 9.29 Ϯ 0.02 9.79 Ϯ 0.13 4.8 Y166A E2 9.83 Ϯ 0.03 10.33 Ϯ 0.32 1.4 L167F E2 9.93 Ϯ 0.02 10.48 Ϯ 0.11 1.0 E170A E2 10.20 Ϯ 0.04 10.20 Ϯ 0.07 1.9 E170K E2 10.16 Ϯ 0.02 9.31 ؎ 0.11 14.6 R171A E2 9.76 Ϯ 0.01 9.50 ؎ 0.29 9.4 L174A E2 10.21 Ϯ 0.08 10.51 Ϯ 0.01 0.9 L174P E2 10.15 Ϯ 0.01 9.37 ؎ 0.14 12.5 Q177E E2 10.08 Ϯ 0.04 10.58 Ϯ 0.17 0.8 Y179F E2 9.85 Ϯ 0.02 9.25 ؎ 0.18 16.5 L183A E2 10.00 Ϯ 0.11 10.44 Ϯ 0.20 1.1 Q185A E2 10.18 Ϯ 0.10 10.59 Ϯ 0.37 0.8 K392A E3 10.20 Ϯ 0.12 10.57 Ϯ 0.18 0.8 D393A E3 9.96 Ϯ 0.09 10.50 Ϯ 0.23 0.9 E397A E3 10.21 Ϯ 0.06 10.54 Ϯ 0.20 0.9 W400A TM7 10.12 Ϯ 0.04 9.43 ؎ 0.02 11.0 E401A TM7 9.76 Ϯ 0.02 10.20 Ϯ 0.10 1.9 TABLE 2 Affinity constants and variations in free energy of interaction of wild-type and modified MT7 toxins for wild-type and mutated hM1 receptors Data were inferred from binding experiments with ͓3 H͔NMS and calculated with the ternary complex equation.
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ABCC8 p.Leu183Ala 21685390:156:693
status: NEW