Functional expression and characterization of an acidic actinoporin from sea anemone Sagartia rosea

Xiaoyu Jiang, Huiping Chen, Wenli Yang, Yun Liu, Wei Liu, Jianwen Wei, Hongbin Tu, Xiaojin Xie, Lei Wang, Anlong Xu

Research output: Contribution to journalArticle

23 Scopus citations

Abstract

Src I is the first reported acidic actinoporin from sea anemone Sagartia rosea with a pI value of 4.8 and comprises 13.9% α-helix, 65.1% β-sheet, and 18.2% random coil. For structure-function studies, Src I was expressed in Escherichia coli as a cleavable fusion protein. Recombinant Src I exhibited obviously hemolytic activity, but the fusion protein Trx-Src I almost lost its hemolytic activity, suggesting the importance of the N-terminal amphiphilic α-helix for its functional activity. The cytotoxic effects of Src I depending on the toxin concentration and incubation time were also observed on cultured cells. Among five cell lines: NIH/3T3, U251, NSCLC, BEL-7402, and BGC-823, NSCLC was the most sensitive cells with ID50 2.8μg/ml and BGC-823 was the least sensitive cells with ID50 7.4μg/ml. After incubated with lipid SUVs, such as SM-SUVs and SM/PC-SUVs, the hemolytic activity of Src I was inhibited to some extent. When incubated with calcein-entrapped lipid LUVs, such as SM-LUVs, SM/PC-LUVs, and SM/PG-LUVs, Src I induced release of entrapped calcein. According to the interaction with lipid vesicles, we proposed that it was the membrane matrix made up of phospholipids, not a particular phospholipid that facilitates Src I to react properly.

Original languageEnglish (US)
Pages (from-to)562-570
Number of pages9
JournalBiochemical and biophysical research communications
Volume312
Issue number3
DOIs
StatePublished - Dec 19 2003
Externally publishedYes

Keywords

  • Acidic actinoporin
  • Cytolytic and cytotoxic activity
  • Lipid vesicles
  • Sagartia rosea
  • Src I

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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