Characterization of the Native Disulfide Isomers of the Novel χ-Conotoxin PnID: Implications for Further Increasing Conotoxin Diversity
Michael J. Espiritu,
Jonathan K. Taylor,
Christopher K. Sugai,
Parashar Thapa,
Nikolaus M. Loening,
Emma Gusman,
Zenaida G. Baoanan,
Michael H. Baumann,
Jon-Paul Bingham
Affiliations
Michael J. Espiritu
Department of Molecular Biosciences and Bioengineering, College of Tropical Agriculture and Human Resources, University of Hawai’i, Honolulu, HI 96822, USA
Jonathan K. Taylor
School of Pharmacy, Pacific University, 222 SE 8th Ave, Ste. 451, Hillsboro, OR 97123, USA
Christopher K. Sugai
Department of Molecular Biosciences and Bioengineering, College of Tropical Agriculture and Human Resources, University of Hawai’i, Honolulu, HI 96822, USA
Parashar Thapa
Department of Molecular Biosciences and Bioengineering, College of Tropical Agriculture and Human Resources, University of Hawai’i, Honolulu, HI 96822, USA
Nikolaus M. Loening
Department of Chemistry, Lewis & Clark College, 615 S Palatine Hill Road, Portland, OR 97219, USA
Emma Gusman
School of Pharmacy, Pacific University, 222 SE 8th Ave, Ste. 451, Hillsboro, OR 97123, USA
Zenaida G. Baoanan
Department of Molecular Biosciences and Bioengineering, College of Tropical Agriculture and Human Resources, University of Hawai’i, Honolulu, HI 96822, USA
Michael H. Baumann
Designer Drug Research Unit, Intramural Research Program, National Institute on Drug Abuse (NIDA), National Institutes of Health (NIH), 333 Cassell Drive Suite 4400, Baltimore, MD 21224, USA
Jon-Paul Bingham
Department of Molecular Biosciences and Bioengineering, College of Tropical Agriculture and Human Resources, University of Hawai’i, Honolulu, HI 96822, USA
χ-Conotoxins are known for their ability to selectively inhibit norepinephrine transporters, an ability that makes them potential leads for treating various neurological disorders, including neuropathic pain. PnID, a peptide isolated from the venom of Conus pennaceus, shares high sequence homology with previously characterized χ-conotoxins. Whereas previously reported χ-conotoxins seem to only have a single native disulfide bonding pattern, PnID has three native isomers due to the formation of different disulfide bond patterns during its maturation in the venom duct. In this study, the disulfide connectivity and three-dimensional structure of these disulfide isomers were explored using regioselective synthesis, chromatographic coelution, and solution-state nuclear magnetic resonance spectroscopy. Of the native isomers, only the isomer with a ribbon disulfide configuration showed pharmacological activity similar to other χ-conotoxins. This isomer inhibited the rat norepinephrine transporter (IC50 = 10 ± 2 µM) and has the most structural similarity to previously characterized χ-conotoxins. In contrast, the globular isoform of PnID showed more than ten times less activity against this transporter and the beaded isoform did not display any measurable biological activity. This study is the first report of the pharmacological and structural characterization of an χ-conotoxin from a species other than Conus marmoreus and is the first report of the existence of natively-formed conotoxin isomers.