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Characterization, purification and phylogenetic analysis of a cytolysin from the sea anemone Heteractis magnifica of the Indian Ocean

2010; BioMed Central; Volume: 16; Issue: 2 Linguagem: Inglês

10.1590/s1678-91992010000200006

1678-4936

S Karthikayalu, V. Rama, R. Kirubagaran, R. Venkatesan,

Erythrocyte Function and Pathophysiology

It is well established that sea anemones comprise a rich source of cytolytic toxins. The present study reports the isolation and characterization of a cytolysin obtained from the sea anemone Heteractis magnifica collected in the Andaman Islands of the Indian Ocean. The crude extract was screened for hemolytic activity by a blood agar plate method and a 6-mm zone of clearance was observed after incubation. The hemolytic property of the crude extract, tested by the microtiter plate method, revealed positive results at concentrations as low as 120 ng/mL. Furthermore, it was favored by alkaline pH and was stable up to 60°C. On the other hand, the hemolytic effect was abolished by the addition of human serum. Purification steps involved ammonium sulfate precipitation and subsequent desalting by dialysis, followed by anion- and cation-exchange chromatographies. The purified fractions displayed the presence of a 19-kDa cytolysin when analyzed by SDS-PAGE. The conserved region of the cytolysin (with 303 bp) was amplified by RT-PCR and was sequenced. The sequence showed maximum homology (97%) with the already reported cytolysins from other sea anemone species.