Minimal conformation of the α-conotoxin ImI for the α7 neuronal nicotinic acetylcholine receptor recognition: correlated CD, NMR and binding studies

Abstract

The α-ImI conotoxin, a selective potent inhibitor of the mammalian neuronal α7 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 Asp 5-Pro 6-Arg 7 in the first loop and Trp 10 in the second loop. The deletion of the Cys 3, Cys 12 disulfide bond in the α-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 α-ImI, the weakest analog (R7A) and peptide II (equipotent to α-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 α-ImI, revealing the crucial functional role of the Arg 7 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 α7 n-AchR. Last, the β-turn forming property of the 6th residue (Pro) in the active site of the α-ImI can be correlated with its affinity.