Intraspecific variations in Conus purpurascens injected venom using LC/MALDI-TOF-MS and LC-ESI-TripleTOF-MS
- 404 Downloads
The venom of cone snails is composed of highly modified peptides (conopeptides) that target a variety of ion channels and receptors. The venom of these marine gastropods represents a largely untapped resource of bioactive compounds of potential pharmaceutical value. Here, we use a combination of bioanalytical techniques to uncover the extent of venom expression variability in Conus purpurascens, a fish-hunting cone snail species. The injected venom of nine specimens of C. purpurascens was separated by reversed-phase high-performance liquid chromatography (RP-HPLC), and fractions were analyzed using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS) in parallel with liquid chromatography-electrospray ionization (LC-ESI)-TripleTOF-MS to compare standard analytical protocols used in preparative bioassay-guided fractionations with a deeper peptidomic analysis. Here, we show that C. purpurascens exhibits pronounced intraspecific venom variability. RP-HPLC fractionation followed by MALDI-TOF-MS analysis of the injected venom of these nine specimens identified 463 distinct masses, with none common to all specimens. Using LC-ESI-TripleTOF-MS, the injected venom of these nine specimens yielded a total of 5517 unique masses. We also compare the injected venom of two specimens with their corresponding dissected venom. We found 2566 and 1990 unique masses for the dissected venom compared to 941 and 1959 masses in their corresponding injected venom. Of these, 742 and 1004 masses overlapped between the dissected and injected venom, respectively. The results indicate that larger conopeptide libraries can be assessed by studying multiple individuals of a given cone snail species. This expanded library of conopeptides enhances the opportunities for discovery of molecular modulators with direct relevance to human therapeutics.
KeywordsConotoxin Conopeptide Cone snail Liquid chromatography Mass spectrometry Peptidomics
We thank C. Moller, N. Vanderweit, and H. Cano for their help with isolating injected venom from C. purpurascens. This work was partially funded by the Florida Sea Grant Program (Grant R/LR-MB-28).
- 13.Abdel-Rahman MA, Abdel-Nabi IM, El-Naggar MS, Abbas OA, Strong PN (2011) Intraspecific variation in the venom of the vermivorous cone snail Conus vexillum. Comp Biochem Physiol Part C: Toxicol Pharmacol 154:318–325Google Scholar
- 17.Dutertre S, Jin AH, Vetter I, Hamilton B, Sunagar K, Lavergne V, Dutertre V, Fry BG, Antunes A, Venter DJ, Alewood PF, Lewis RJ (2014) Evolution of separate predation- and defence-evoked venoms in carnivorous cone snails. Nat Commun 5:3521Google Scholar
- 32.Dowell C, Olivera BM, Garrett JE, Staheli ST, Watkins M, Kuryatov A, Yoshikami D, Lindstrom JM, McIntosh JM (2003) α-Conotoxin PIA is selective for α6 subunit-containing nicotinic acetylcholine receptors. J Neurosci 23:8445–8452Google Scholar
- 34.Shon K, Olivera BM, Watkins M, Jacobsen R, Gray WR, Floresca CZ, Cruz LJ, Hillyard DR, Brink A, Terlau H, Yoshikami D (1998) μ-Conotoxin PIIIA, a new peptide for discriminating among tetrodotoxin-sensitive Na channel subtypes. J Neurosci 18:4473–4481Google Scholar
- 43.Sunagar K, Undheim EA, Scheib H, Gren EC, Cochran C, Person CE, Koludarov I, Kelln W, Hayes WK, King GF, Antunes A, Fry BG (2014) Intraspecific venom variation in the medically significant Southern Pacific Rattlesnake (Crotalus oreganus helleri): biodiscovery, clinical and evolutionary implications. J Proteomics 99:68–83CrossRefGoogle Scholar
- 44.Jorge RJ, Monteiro HS, Goncalves-Machado L, Guarnieri MC, Ximenes RM, Borges-Nojosa DM, Luna KP, Zingali RB, Correa-Netto C, Gutierrez JM, Sanz L, Calvete JJ, Pla D (2015) Venomics and antivenomics of Bothrops erythromelas from five geographic populations within the Caatinga ecoregion of northeastern Brazil. J Proteomics 114:93–114CrossRefGoogle Scholar
- 47.Andrews GL, Simons BL, Young JB, Hawkridge AM, Muddiman DC (2011) Performance characteristics of a new hybrid quadrupole time-of-flight tandem mass spectrometer (TripleTOF 5600). Anal Chem 83:5442–5446Google Scholar