DNA-DNA gyrase complex: the wrapping of the DNA duplex outside the enzyme

Digestion of the complex between double-stranded DNA and M. luteus or E. coli DNA gyrase with staphylococcal nuclease gives a 143 ± 3 base pair DNA fragment containing no single-chain scissions. Digestion of the same complex with bovine pancreatic DNAase I gives six discernible single-stranded DNA bands upon electrophoresis of the product in a denaturing gel. The lengths of these fragments, in number of nucleotides, are measured to be 47 ± 1, 57 ± 1, 67 ± 1, 77 ± 1, 86 ± 1 and 96 ± 1, respectively. These results support the notion that in the DNA-gyrase complex, a segment(s) of the DNA helix is wrapped around the enzyme. The wrapping of the DNA around the enzyme has been proposed previously based on the observation that in the absence of ATP, the linking number of a duplex DNA ring covalently closed by ligase in the presence of bound gyrase is higher than in the absence of gyrase (Liu and Wang, 1978). The coiling of DNA around the enzyme in the complex is believed to be intimately related to the ATP-dependent negative supercoiling of covalently closed duplex DNA ring by DNA gyrase. It has also been observed that digestion of pure double-stranded DNA by pancreatic DNAase I in the presence of calcium phosphate precipitate or solid hydroxylapatite gives a ladder of single-stranded DNA fragments of integral multiples of 10 nucleotides. This finding suggests that such a pancreatic DNAase I cleavage pattern is indicative of a DNA duplex lying on the outside of a surface.