Key residues in the nicotinic acetylcholine receptor ?2 subunit contribute to ?-conotoxin LvIA binding.

ABSTRACT: ?-Conotoxin LvIA (?-CTx LvIA) is a small peptide from the venom of the carnivorous marine gastropod Conus lividus and is the most selective inhibitor of ?3?2 nicotinic acetylcholine receptors (nAChRs) known to date. It can distinguish the ?3?2 nAChR subtype from the ?6?2* (* indicates the other subunit) and ?3?4 nAChR subtypes. In this study, we performed mutational studies to assess the influence of residues of the ?2 subunit versus those of the ?4 subunit on the binding of ?-CTx LvIA. Although two ?2 mutations, ?3?2[F119Q] and ?3?2[T59K], strongly enhanced the affinity of LvIA, the ?2 mutation ?3?2[V111I] substantially reduced the binding of LvIA. Increased activity of LvIA was also observed when the ?2-T59L mutant was combined with the ?3 subunit. There were no significant difference in inhibition of ?3?2[T59I], ?3?2[Q34A], and ?3?2[K79A] nAChRs when compared with wild-type ?3?2 nAChR. ?-CTx LvIA displayed slower off-rate kinetics at ?3?2[F119Q] and ?3?2[T59K] than at the wild-type receptor, with the latter mutant having the most pronounced effect. Taken together, these data provide evidence that the ?2 subunit contributes to ?-CTx LvIA binding and selectivity. The results demonstrate that Val(111) is critical and facilitates LvIA binding; this position has not previously been identified as important to binding of other 4/7 framework ?-conotoxins. Thr(59) and Phe(119) of the ?2 subunit appear to interfere with LvIA binding, and their replacement by the corresponding residues of the ?4 subunit leads to increased affinity.

SUBMITTER: Zhangsun D 

PROVIDER: S-EPMC4392282 | biostudies-literature | 2015 Apr

REPOSITORIES: biostudies-literature