Archives of Toxicology

, Volume 85, Issue 4, pp 305–313 | Cite as

Characterization of toxins from the broad-banded water snake Helicops angulatus (Linnaeus, 1758): isolation of a cysteine-rich secretory protein, Helicopsin

  • Amalid Estrella
  • Elda E. Sánchez
  • Jacob A. Galán
  • W. Andy Tao
  • Belsy Guerrero
  • Luis F. Navarrete
  • Alexis Rodríguez-Acosta
Molecular Toxicology

Abstract

Helicops angulatus (broad-banded water snake) according to recent proposals is presently cited in the family Dipsadidae, subfamily Xenodontinae, forming the tribe Hydropsini along with the genera Hydrops and Pseudoeryx. The current work characterizes the proteolytic and neurotoxic activities of H. angulatus crude toxins from salivary excretion (SE) and describes the isolation and identification of a cysteine-rich secretory protein (CRISP) called helicopsin. The SE lethal dose (LD50) was 5.3 mg/kg; however, the SE did not contain hemorrhagic activity. Helicopsin was purified using activity-guided, Superose 12 10/300 GL molecular exclusion, Mono Q10 ion exchange, and Protein Pak 60 molecular exclusion. Sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) showed a highly purified band of approximately 20 kDa. The minimal lethal dose for helicopsin was 0.4 mg/kg. Liquid chromatography mass spectrometry (LC-MS/MS) analysis identified 2 unique peptides MEWYPEAAANAER and YTQIVWYK, representing a protein sequence (deleted homology) belonging to cysteine-rich secretory proteins, which are conserved in snake venoms (CRISPs). CRISPs are a large family of cysteine-rich secretory proteins found in various organisms and participate in diverse biological processes. Helicopsin exhibited robust neurotoxic activity as evidenced by immediate death (~8 min) due to respiratory paralysis in NIH mice. These observations for helicopsin purified from H. angulatus provide further evidence of the extensive distribution of highly potent neurotoxins in the Colubroidea superfamily of snakes than previously described.

Keywords

CRISP Colubroidea Dipsadidae salivary excretion Neurotoxin Helicops angulatus Broad-banded water snake 

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Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Amalid Estrella
    • 1
  • Elda E. Sánchez
    • 2
  • Jacob A. Galán
    • 3
  • W. Andy Tao
    • 3
  • Belsy Guerrero
    • 4
  • Luis F. Navarrete
    • 1
  • Alexis Rodríguez-Acosta
    • 1
  1. 1.Immunochemistry SectionTropical Medicine Institute of the Universidad Central de VenezuelaCaracasVenezuela
  2. 2.Department of Chemistry, College of Arts and SciencesTexas A&M University-KingsvilleKingsvilleUSA
  3. 3.Departments of Biochemistry, Chemistry, Medicinal Chemistry & Molecular PharmacologyPurdue UniversityWest LafayetteUSA
  4. 4.Pathophysiology Laboratory, Medicine Experimental CenterInstituto Venezolano de Investigaciones Científicas (IVIC)CaracasVenezuela

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