α‑Conotoxin [S9A]TxID Potently Discriminates between α3β4 and α6/α3β4 Nicotinic Acetylcholine Receptors

α3β4
nAChRs have been implicated in various pathophysiological conditions.
However, the expression profile of α3β4 nAChRs and α6/α3β4
nAChRs overlap in a variety of tissues. To distinguish between these
two subtypes, we redesigned peptide 1 (α-conotoxin
TxID), which inhibits α3β4 and α6/α3β4
nAChR subtypes. We systematically mutated 1 to evaluate
analogue selectivity for α3β4 vs α6/α3β4
nAChRs expressed in Xenopus laevis oocytes.
One analogue, peptide 7 ([S9A]­TxID), had 46-fold greater
potency for α3β4 versus α6/α3β4 nAChRs.
Peptide 7 had IC50s > 10 μM for other
nAChR subtypes. Molecular dynamics simulations suggested that Ser-9
of TxID was involved in a weak hydrogen bond with β4 Lys-81
in the α6β4 binding site but not in the α3β4
binding site. When Ser-9 was substituted by an Ala, this hydrogen
bond interaction was disrupted. These results provide further molecular
insights into the selectivity of 7 and provide a guide
for designing ligands that block α3β4 nAChRs.