Purification and Characterization of Alpha Toxin Clostridium perfringens Type A

Heshmati, Zahra; Fathi Najafi, Mohsen; Ronaghi, Gholam Hossein

doi:10.21608/eajbsc.2020.115949

Purification and Characterization of Alpha Toxin Clostridium perfringens Type A

Document Type : Original Article

Authors

¹ Department of Science, Mashhad Branch, Islamic Azad University, Mashhad, Iran

² Education and Extension Organization (AREEO), Agricultural Research, Razi Vaccine and Serum Research Institute, Mashhad Branch, Mashhad, Iran

10.21608/eajbsc.2020.115949

Abstract

Clostridium perfringens is a toxin-producing anaerobe with the ability to cause multiple diseases in humans and animals. One of the toxins produced by all strains of this bacterium is an alpha toxin (phospholipase C). This study aimed to purify and characterize alpha-toxin. C. perfringens type A was grown in an optimized medium, the culture supernatant was separated, and ammonium sulfate precipitated protein was subjected to ion exchange and gel filtration chromatography methods. The purification steps were monitored by SDS-PAGE and hemolysin activity for each sample and performed by the hemolysin assay method. The purified alpha-toxin appeared in a single band in 43 KDa. The enzymatic efficiency yield was about 88% with a specific activity of 69170 and 654 fold of purification. According to the results, alpha-toxin is Zn and Mg metal-enzyme dependent on the temperature and pH optimum of 40 °C and 6, respectively. The enzyme K_mand V_ma_xvalues of 3.8 mg/ml and 0.19 were calculated as the enzyme in the presence of lecithin. The partially purified alpha toxin with specific characters can be used for different applications such as antibody production and determination of the anti-toxin rate in animal serum.

Keywords