Structure of the replicative form of bacteriophage φX174: V. Interconversions between twisted, extended and randomly coiled forms of cyclic DNA

Abstract

As is already known, double-stranded DNA of bacteriophage φX174 occurs in a twisted cyclic double helix (21 s). The two strands making up the double helix are separately continuous. The tertiary twists in the molecule are released by the introduction of single-strand chain breaks by pancreatic DNase. The product formed is an extended cyclic helix (17 s). A third form of double-stranded φX174 DNA, known as coiled cyclic double-stranded DNA (40 s in m-NaCl at neutral pH) is obtained upon alkaline denaturation of the twisted cyclic helix. In this paper, quantitative proof is given that the conversion of the twisted cyclic helix to the extended cyclic form is induced by only one single and random chain break. The conformational change is accompanied by a twofold decrease in biological activity. The further biological inactivation of the extended ring form occurs at a rate which suggests that approximately one out of 20 single-strand chain breaks in double-stranded DNA is lethal. The introduction of single-strand chain breaks into the cyclic coil (40 s) leads to the spontaneous restoration of the helical structure. An extended cyclic helix (17 s) is formed as indicated by ultracentrifugal analysis and electron microscopy. It has been shown that the previously reported but unexplained high resistance to ultraviolet inactivation of the denatured cyclic coil is due to host-cell reactivation. The required DNA double helix may arise from the cyclic coil by the action of nucleases known to be involved in host-cell reactivation.