Minimal conformation of the alpha-conotoxin ImI for the alpha7 neuronal nicotinic acetylcholine receptor recognition: correlated CD, NMR and binding studies.
The alpha-ImI conotoxin, a selective potent inhibitor of the mammalian neuronal alpha7 nicotinic acetylcholine receptor (n-AchR), was shown by point mutation or by L-alanine scanning to display two regions essential for bioactivity: the active site Asp5-Pro6-Arg7 in the first loop and Trp10 in the second loop. The deletion of the Cys3,Cys12 disulfide bond in the alpha-ImI scaffold, e.g. peptide II, had no effect on its binding affinity. CD spectra, NMR studies and structure calculations were carried out on the wild type alpha-ImI, the weakest analog (R7A) and peptide II (equipotent to alpha-ImI) in order to point out the conformational differences between these compounds. Then, an attempt to correlate the conformational data and the affinity results was proposed. CD and NMR data were identical for the R7A analog and alpha-ImI, revealing the crucial functional role of the Arg7 side chain. On the other hand, the scaffold of the first loop in peptide II was shown by NMR to represent the minimal conformation for the optimal interaction of the toxin with the neuronal alpha7 n-AchR. Last, the beta-turn forming property of the 6th residue (Pro) in the active site of the alpha-ImI can be correlated with its affinity.
Lamthanh
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CEA, Département d'Ingénierie et d'Etudes des Protéines, Gif sur Yvette, France. lamthanh@dsvidf.cea.fr
Jegou-Matheron
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C
Servent
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Ménez
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Lancelin
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D000595:Amino Acid Sequence; D000818:Animals; D002942:Circular Dichroism; D020916:Conotoxins; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D008978:Mollusk Venoms / Q000378:metabolism*; D009154:Mutation; D009842:Oligopeptides / Q000737:chemistry* / Q000378:metabolism*; D017354:Point Mutation; D011487:Protein Conformation; D011978:Receptors, Nicotinic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D064569:alpha7 Nicotinic Acetylcholine Receptor
D020916:Conotoxins; D008978:Mollusk Venoms; D009842:Oligopeptides; D011978:Receptors, Nicotinic; D064569:alpha7 Nicotinic Acetylcholine Receptor; C087686:alpha-conotoxin ImI