Characterization of the cytotoxic effect of a chimericrestriction enzyme, H1o-FokI

摘要

Our primary goal was to create an efficient cytotoxic agent. To do this, we created a gene that expresses a chimeric hybrid of the linker histone, H10 and the nuclease domain of the type IIs restriction enzyme, FokI. The linkage of the FokI nuclease domain to a high affinity but low DNA-sequence-specificity binding protein is unique. It is highly cytotoxic. We demonstrate, by transiently transfecting 3T3 mouse fibroblasts, that 63% of the cells taking up the chimeric gene are killed. The chimeric protein is localized to the nucleus. An extract of the protein produced in E. coli degrades DNA, indicating that it is nucleolytically active. The ultimate mechanism through which the chimericprotein produces cell death is likely through the induction of apoptosis.