Resurfacing receptor binding domain of Colicin N to enhance its cytotoxic effect on human lung cancer cells.

Colicin N (ColN) is a bacteriocin secreted by Escherichia coli ( E. coli ) to kill other Gram-negative bacteria by forcefully generating ion channels in the inner membrane. In addition to its bactericidal activity, ColN have been reported to selectively induce apoptosis in human lung cancer cells via the suppression of integrin modulated survival pathway. However, ColN showed mild toxicity against human lung cancer cells which could be improved for further applications. The protein resurfacing strategy was chosen to engineer ColN by extensive mutagenesis at solvent--exposed residues on ColN. The highly accessible Asp and Glu on wild-type ColN (ColN ^WT ) were replaced by Lys to create polycationic ColN (ColN ⁺¹² ). Previous studies have shown that increase of positive charges on proteins leads to the enhancement of mammalian cell penetration as well as increased interaction with negatively charged surface of cancer cells. Those solvent--exposed residues of ColN were identified by Rosetta and AvNAPSA (Average number of Neighboring Atoms Per Side-chain Atom) approaches. The findings revealed that the structural features and stability of ColN ⁺¹² determined by circular dichroism were similar to ColN ^WT . Furthermore, the toxicity of ColN ⁺¹² was cancer -selective. Human lung cancer cells, H460 and H23, were sensitive to ColN but human dermal papilla cells were not. ColN ⁺¹² also showed more potent toxicity than ColN ^WT in cancer cells. This confirmed that polycationic resurfacing method has enabled us to improve the anticancer activity of ColN towards human lung cancer cells.
Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(© 2021 The Author(s).)