Abstract:
Bacteria, exposed to stress from the environment or competition, release toxins called bacteriocins. Bacteriocins are classified by their mode of cytotoxicity such as pore former, endonuclease, etc. Colicins are the member of bacteriocins produced by Escherichia coli (E.coli). Among several types of colicins cytotoxicity, Pore-forming colicins has been reported in both bacteria and various type of cancer cells. Colicin N is also a pore-forming colicin and the anticancer activity of colicin N has not been extensively reported. Here we examined the cytotoxic effects of colicin N and its domain on cancer cells. The expression and purification of recombinant full-length and truncated colicin N were performed in E. coli and an affinity chromatography. The physicochemical properties of purified proteins were then assessed by SDS-PAGE, western blot, Circular Dichroism and mass spectrometry. Then, HCT-116, HT-29, MCF-7, MDA-MB-231 and A549 cells were treated with full-length Colicin N for anticancer test by MTT assay. Furthermore, a series of truncated colicin N with a deletion of one or two domains were tested on HCT-116 cells. We show that, full-length and truncated colicin N were produced successfully and showed the expected physical properties such as molecular weight and secondary structures. Regarding the cytotoxicity against cancer cells, we found that HCT-116 cells (colon cancer cells) was the most sensitive to full length colicin N. Likewise, HT-29, MDA-MB-231 and A549 cells were also sensitive to full-length colicin N. In contrast to these cancer cells, the cell viability of MCF-7 was promoted in the presence of full-length colicin N. The effect of full-length colicin N on cancer cells seemed to be dependent on types of cancer cells. The truncated colicin N did not cause cytotoxicity to cancer cells hence the toxic domain of colicin N is not sufficient for anticancer activity. Furthermore, colicin N mutant with increased number of positive charges on its surface was constructed. The preliminary results demonstrated that this mutant was more cytotoxic than wild type colicin N. This could be due to the difference in charges or conformational changes of colicin N mutant. The results from this study can improve the basic knowledge about colicin N related cytotoxic activity on cancer cells and suggestions that colicin N may be considered for its promising application of therapeutic and natural antitumor drugs.
