We here describe a novel α-conopeptide, Eu1.6 from Conus eburneus, which exhibits strong anti-nociceptive activity by an unexpected mechanism of action. Unlike other α-conopeptides that largely target nicotinic acetylcholine receptors (nAChRs), Eu1.6 displayed only weak inhibitory activity at the α3β4 and α7 nAChR subtypes and TTX-resistant sodium channels, and no activity at TTX-sensitive sodium channels in rat dorsal root ganglion (DRG) neurons, or opiate receptors, VR1, KCNQ1, L- and T-type calcium channels expressed in HEK293 cells. However, Eu1.6 inhibited high voltage-activated N-type calcium channel currents in isolated mouse DRG neurons which was independent of GABAB receptor activation. In HEK293 cells expressing CaV2.2 channels alone, Eu1.6 reversibly inhibited depolarization-activated Ba2+ currents in a voltage- and state-dependent manner. Inhibition of CaV2.2 by Eu1.6 was concentration-dependent (IC50 ~1 nM). Significantly, systemic administration of Eu1.6 at doses of 2.5–5.0 μg/kg exhibited potent analgesic activities in rat partial sciatic nerve injury and chronic constriction injury pain models. Furthermore, Eu1.6 had no significant side-effect on spontaneous locomotor activity, cardiac and respiratory function, and drug dependence in mice. These findings suggest α-conopeptide Eu1.6 is a potent analgesic for the treatment of neuropathic and chronic pain and opens a novel option for future analgesic drug design.
The influenza fusion peptide located at the N-terminus of the hemagglutinin HA2 subunit initiates the fusing process of the viral membrane with the host cell endosomal membrane. It had been reported that the structure of a 20-residue H3 subtype fusion peptide (H3-HAfp20) was significantly different with that of a H1 subtype 23-residue one (H1-HAfp23). The sequential difference between the 12th and 15th residues of H1 and H3 subtypes could not fully explain the conformational variation. The first and last three amino acids of H3-HAfp23 involved in formation of hydrogen bonds may play an important role in fusion process. To confirm this hypothesis, we investigate the structures of H3-HAfp23 peptide and its mutants, G1S and G1V, in dodecylphosphatidyl choline micelles by using heteronuclear NMR technology. The results demonstrate that, similar to H1-HAfp23 but significantly different with H3-HAfp20, H3-HAfp23 also has tight helical hairpin structure with the N- and C-terminuses linked together because of the hydrogen bonds between Gly1 and the last three amino acids, Trp21―Tyr22―Gly23. Although the ‘hemifusion’ G1S and lethal G1V mutants have hairpin-like helical structures, the distances between the N- and C-terminuses are increased as shortage of the hydrogen bonds and the larger kink angle between the antiparallel helices. The paramagnetic ion titration experiments show that the terminuses are inserted into the dodecylphosphatidyl choline micelles used as solving media. These may imply that the tight helical hairpin structure, especially the closed conformation at terminus, plays an important role in fusion activity.
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