Construction and screening of a combinatorial shiga toxin library

grantor: University of TorontoShiga-like toxin I (SLTI) is a bacterial toxin that is able to penetrate and kill mammalian cells. In this project, a library of SLTI mutants with altered receptor specificity was constructed using a synthetic oligonucleotide that varied 9 amino acids in the specificity-determining region of the B subunit. A high throughput screening procedure based on the cytotoxic activity of the toxin mutants was used as a direct approach to assay for toxins that killed SK-BR-3, a breast cancer cell line resistant to native SLTI. Seventeen out of the 1000 bacterial lysates screened were found to be cytotoxic towards SK-BR-3 cells. Analysis of the mutants revealed that receptor expression and cellular morphology of SK-BR-3 cells changed over time, underscoring the importance of the Shiga-like toxin I library as a potentially valuable source of therapeutic agents.M.Sc