Left-handed Z-DNA regions are present in negatively supercoiled bacteriophage PM2 DNA

Bacteriophage PM2 DNA is a 10-kb covalently closed circular (ccc) molecule with a reported superhelical density of sigma = -0.12. Here we describe the binding of anti-Z-DNA antibodies to PM2 form I DNA under high and low salt conditions. The binding to PM2 DNA has been demonstrated by competitive radioimmunoassay (RIA), retardation of the DNA:antibody complexes in agarose gels and visualization by electron microscopy. The antibody binding is dependent on the degree of negative supercoiling. Thus, PM2 form II and form III did not bind the antibody. The low salt RIA results indicated the presence of 200-400 bp of left-handed DNA per PM2 molecule. This could reduce the effective superhelical density to sigma = -0.04 to -0.08, a range comparable with those found for other ccc DNAs in vivo. Electron microscopy revealed that a maximum of 22 antibody molecules bind to PM2. Single-site restriction with HpaII of the fixed DNA:antibody complex showed a cluster of four to five antibody molecules bound near one end of the linear DNA molecule. The evidence presented indicates that PM2 DNA contains regions of left-handed conformation under physiological conditions (low salt concentration) as well as at high salt concentrations. In addition, electrophoretic analyses of PM2 topoisomers indicate the presence of left-handed regions at superhelical densities less than that of isolated PM2 DNA.