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J. Venom. Anim. Toxins incl. Trop. Dis.
V.20, 2014.
Original paper - ISSN 1678-9199.
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J. Venom. Anim. Toxins incl. Trop. Dis. V.20, 2014. Original paper - ISSN 1678-9199. |
Identification of two novel cytolysins from the hydrozoan Olindias sambaquiensis (Cnidaria)
1Sao Paulo Experimental Coast Campus , Sao Paulo State University (UNESP -Univ Estadual Paulista), Sao Vicente , Sao Paulo State , Brazil .
2Botucatu Medical School , Sao Paulo State University (UNESP - Univ Estadual Paulista), Botucatu , Sao Paulo State , Brazil .
3Department of Biochemistry , Institute of Biology , State University of Campinas (UNICAMP), Campinas , Sao Paulo State , Brazil .
4Center of Biological and Health Sciences , Mackenzie Presbyterian University , Sao Paulo , Sao Paulo State , Brazil .
5UNESP , Campus do Litoral Paulista , Unidade Sao Vicente , Praga Infante D. Henrique, s/n, Sao Vicente , SP CEP11330-900, Brasil .
ABSTRACT
Background
Although the hydrozoan Olindias sambaquiensis is the most common jellyfish associated with human envenomation in southeastern and southern Brazil, information about the composition of its venom is rare. Thus, the present study aimed to analyze pharmacological aspects of O. sambaquiensis venom as well as clinical manifestations observed in affected patients. Crude protein extracts were prepared from the tentacles of animals; peptides and proteins were sequenced and submitted to circular dichroism spectroscopy. Creatine kinase, cytotoxicity and hemolytic activity were evaluated by specific methods.
Results
We identified two novel cytolysins denominated oshem 1 and oshem 2 from the tentacles of this jellyfish. The cytolysins presented the amino acid sequences NEGKAKCGNTAGSKLTFKSADECTKTGQK (oshem 1) and NNSKAKCGDLAGWSKLTFKSADECTKTGQKS (oshem 2) with respective molecular masses of 3.013 kDa and 3.375 kDa. Circular dichroism revealed that oshem 1 has random coils and small α-helix conformation as main secondary structure whereas oshem 2 presents mainly random coils as its main secondary structure probably due to the presence of W (13) in oshem 2. The hemolysis levels induced by oshem 1 and oshem 2 using a peptide concentration of 0.2 mg/mL were, respectively, 51.7 ± 6.5% and 32.9 ± 8.7% (n = 12 and p ≤ 0.05). Oshem 1 and oshem 2 showed significant myonecrotic activity, evaluated by respective CK level measurements of 1890.4 ± 89 and 1212.5 ± 103 (n = 4 and p ≤ 0.05). In addition, myonecrosis was also evaluated by cell survival, which was measured at 72.4 ± 8.6% and 83.5 ± 6.7% (n = 12 and p ≤ 0.05), respectively. The structural analysis showed that both oshem 1 and oshem 2 should be classified as a small basic hemolytic peptide.
Conclusion
The amino acid sequences of two peptides were highly similar while the primary amino acid sequence analysis revealed W (22th) as the most important mutation. Finally oshem 1 and oshem 2 are the first cytolytic peptides isolated from the Olindias sambaquiensis and should probably represent a novel class of cytolytic peptides.
Key words: Olindias sambaquiensis; Hemolytic; Myonecrosis; Cytotoxicity; Cytolysin; Cnidaria venom
Ethics committee approval
The present study was approved by the Committee for Ethics in Animal Experimentation on the Sao Paulo Experimental Coast Campus of UNESP. The animal utilization was approved by the Committee for Ethics in Animal Experimentation of the Institute of Biology, UNICAMP, certificate number 1320-1. The utilized Swiss mice were supplied by CEMIB, the Multidisciplinary Center for Biological Research of UNICAMP.
Received: August 21, 2013; Accepted: January 30, 2014; Revised: March 25, 2014