New Actinoporins from Sea Anemone Heteractis crispa: Cloning and Functional Expression

---

E. S. Tkacheva, E. V. Leychenko, M. M. Monastyrnaya^*, M. P. Issaeva, E. A. Zelepuga, S. D. Anastuk, P. S. Dmitrenok, and E. P. Kozlovskaya

Pacific Institute of Bioorganic Chemistry, Far Eastern Branch of the Russian Academy of Sciences, pr. 100 let Vladivostoku 159, 690022 Vladivostok, Russia; fax: (4232) 31-4050; E-mail: rita1950@mail.ru

^* To whom correspondence should be addressed.

Received May 13, 2011; Revision received June 7, 2011

---

A new actinoporin Hct-S4 (molecular mass 19,414 ± 10 Da) belonging to the sphingomyelin-inhibited α-pore forming toxin (α-PFT) family was isolated from the tropical sea anemone Heteractis crispa (also called Radianthus macrodactylus) and purified by methods of protein chemistry. The N-terminal nucleotide sequence (encoding 20 amino acid residues) of actinoporin Hct-S4 was determined. Genes encoding 18 new isoforms of H. crispa actinoporins were cloned and sequenced. These genes form a multigene Hct-S family characterized by presence of N-terminal serine in the mature proteins. Highly conserved residues comprising the aromatic phosphorylcholine-binding site and significant structure–function changes in the N-terminal segment (10-27 amino acid residues) of actinoporins were established. Two expressed recombinant actinoporins (rHct-S5 and rHct-S6) were one order less hemolytically active than native actinoporins.

---

KEY WORDS: sea anemone, actinoporin, multigene Hct-S family, recombinant actinoporin, hemolytic activity, structure–function analysis

DOI: 10.1134/S0006297911100063

---