Full Length Research Paper

Cloning and expression of C-terminal of Clostridium perfringens type A enterotoxin and its biological activity

Atieyeh Taherian Fard¹, Fariha Hasan¹, Mojgan Bandehpour², Nariman Mosaffa², Fatemeh Mashhadi Abbas³, Abdul Hameed¹, Aamer Ali Shah¹ and Bahram Kazemi^2*

¹Department of Microbiology, Quaid - i - Azam University, Islamabad, Pakistan.

²Cellular and Molecular Biology Research Center, Shahid Beheshti University, M. C., Tehran, Iran.

³Department of Dentistry, Shahid Beheshti University, M. C, Tehran, Iran.

*Corresponding author. E-mail: bahram_14@yahoo.com. Tel: +982122439957.

Fax: +98 21 22439956.

Accepted 5 July, 2010

Abstract

Pathogenic clostridia produce exocellular toxins that resemble lipoteichoic acid and are described as super antigens. These toxins stimulate T-cell receptor-carrying lymphocytes in peripheral blood and have been used to study immunodeficiency diseases and cancers. The CPE C-terminal region from one of the local type A strain was cloned in the pET32a vector its expression induced with IPTG. The expressed protein was purified by Ni-NTA affinity chromatography and tested for biological activity with Vero cells assay. This region of Clostridium perfringens enterotoxin (CPE) has a predominant ligand-binding activity. In the present study, the biological activity of the C-terminal region of local purified CPE came under study with Vero cell assay, guinea pig skin test and mouse test to evaluate for future use as a therapeutic purpose. The result of this study showed that, the study’s local purified C-CPE had cytotoxic activity in Vero cells even at the minimum dilution of 0.625 ng after a 4-h incubation period. It caused transient increase in capillary permeability in guinea pigs. C-CPE did not have systemic effect on Balb/c mice. The use of the C-CPE peptide may provide a novel way to target drugs to Claudine-expressing cells.

Key words: Cloning, gene expression, Clostridium perfringens enterotoxin (CPE), vero cells, nigrosin, guinea pig skin test, mouse test, Claudine.