Modification of α-conotoxin frameworks through cyclization via an oligopeptide linker has previously been shown as an effective strategy for improving in vivo stability. We have extended this strategy by investigating cyclic analogs of α-conotoxin AuIB, a selective α(3)β(4) nicotinic acetylcholine receptor (nAChR) antagonist, to examine a range of oligopeptide linker lengths on the oxidative formation of disulfide bonds, activity at nAChRs, and stability to degradation by chymotrypsin. Upon nondirected random oxidation, the ribbon isomer formed preferentially with the globular isomer occurring as a minor by-product. Therefore, a regioselective disulfide bond forming strategy was used to prepare the cAuIB-2 globular isomer in high yield and purity. The cAuIB-2 globular isomer exhibited a threefold decrease in activity for the α(3)β(4) nAChR compared to wild-type-AuIB, although it was selective for α(3)β(4) over α(7) and α(4)β(2) subtypes. On the other hand, the cAuIB-2 ribbon isomer was shown to be inactive at all three nAChR subtypes. Nonetheless, all of the cyclic analogs were found to be significantly more stable to degradation by chymotrypsin than wild-type AuIB. As such, the cAuIB-2 globular isomer could constitute a useful probe for studying the role of the α(3)β(4) nAChR in a range of in vivo experimental paradigms.