Z DNA and loop structures by immunoelectronmicroscopy of supercoiled pRW751, a plasmid containing left-handed helices

Abstract

Single and multiple loops were seen when the plasmid pRW751 was allowed to react with anti-Z-DNA or with a Z-specific crosslinking agent. Loop formation was dependent upon negative supercoiling and the presence of Z-specific antibody or crosslinking agent. Restriction enzyme mapping located 18 sites at the bottoms of loops, in addition to the two (dG-dC) _n inserts of pRW751. No more than 5 loops were seen in any of the measured molecules; thus, not all potential Z-sites assume the Z conformation at any particular time. Stretches of alternating purine-pyrimidine sequences occur at all 20 sites. Almost all of the Z sites could be mapped to regions located at the beginnings or ends of reading frames or at various regulatory sites. Our findings support the concept that supercoiling brings distant sequences to within 5Å of one another, allowing joint participation in regulatory processes controlled by DNA-binding proteins.