The mechanism of type IA topoisomerase-mediated DNA topological transformations

Type IA DNA topoisomerases possess several domains forming a toroidal molec ule with a central hole large enough to accommodate single- or double-stran ded DNA. The sign inversion model predicts several protein-DNA intermediate s, including those in which DNA is trapped within the hole. Opposing cystei ne residues were incorporated into two independent domains surrounding the putative DNA binding cavity of E. coli topoisomerase III, creating a molecu le that can be covalently closed or opened by oxidizing or reducing the dis ulfide bond. The formation of the disulfide bond allowed the trapping of si ngle- and double-stranded DNA within the cavity of the enzyme and the ident ification of other intermediates proposed by the sign inversion model.