Effects of methylation on a synthetic polynucleotide: the B--Z transition in poly(dG-m5dC).poly(dG-m5dC).

Abstract

We have compared the behavior in solution of the synthetic polynucleotide poly(dG-m5dC)poly(dG-m5dC) with that of the unmethylated polynucleotide poly(dG-dC)poly(dGdC). In solutions containing high concentrations of salt, poly(dGdC)-poly(dG-dC) is known to exhibit altered circular dichroic and absorption spectra correlated with formation of a left-handed Z DNA structure. poly(dG-m5dC) behaves similarly, but the spectral transition from the B to the Z form occurs at much lower salt concentrations, close to usual physiological conditions. Divalent and polyvalent ions are particularly effective: The B-Z transition of poly(dG-m5dC) (dG-m5dC) can be induced at a Mg2e concentration three orders of magnitude lower than that required for the unmethylated polymer. We have also studied mixed copolymers containing both dC and m5dC. Our results suggest that the sequence m5dC-dG, which occurs in eukaryotic DNA, can have a disproportionately large effect on the B-Z transition.transcriptional activity of the gene (7). For these reasons, we wished to compare the properties of the methylated C-G synthetic polymer with the unmethylated compound.We have found that, as judged by circular dichroism and ultraviolet absorption, poly(dG-m5dC)poly(dG-m5dC) also undergoes a transition from the B to the Z form, but at much lower salt concentrations, so that the Z form is stable under typical physiological conditions. Furthermore, in mixed alternating copolymers of dG-dC and dG-m5dC, the presence of the latter sequence has a disproportionate effect in lowering the amount of divalent ion required to stabilize the Z form. Our results provide some information about the forces that govern the B-Z transition and suggest that there are circumstances in which the Z form might be observed in vivo.It is now accepted that DNA of appropriate nucleotide sequence is capable of forming left-handed double helical structures with Watson-Crick base pairing, as well as the more familiar righthanded structures. X-ray crystallographic studies of the synthetic oligonucleotide d(CpGpCpGpCpG), demonstrated the left-handed, or Z-DNA, conformation (1). Subsequent studies of the smaller oligonucleotide d(CpGpCpG) suggested that there are at least two related Z conformations (2, 3). Diffraction patterns consistent with the Z form have also been observed (4) with fibers of the high molecular weight alternating copolymer poly(dG-dC)-poly(dG-dC) and with poly(dAdC)poly~dG-dT).The solution properties of poly(dG-dC)-poly(dG-dC) were described and analyzed by Pohl and Jovin some years ago (5). They showed that at high salt concentrations (above 2.5 M NaCl or 0.7 M MgCl2) the polynucleotide is converted to a new form by a cooperative, intramolecular process. This new form displays a circular dichroic spectrum that is inverted compared to that of the B form, and an ultraviolet absorption spectrum with increased molar extinction compared to the B form in the range 280-300 nm. The recent x-ray diffraction results, as well as nuclear magnetic resonance studies in sol...

Interaction of Z form poly(dG-dC). poly(dG-dC) with divalent metal ions: localization of the binding sites by I.R. spectroscopy

“…The difference in the spectroscopic effect induced by Co2+ and Ni2+ compared to the effect of Mg2+ on the 1467 cnm1 band seems to be well correlated with the efficiency of the metal ions to induce the B to Z transition in solution (1,16,18,32) and allows us to propose an explanation for the particular role of the nucleophile site in the mechanism of this transition. In conditions in which the B conformation in solution is mainly observed, the thermal agitation through the DNA respiration can modify locally the dG conformation into a syn geometry without breaking the Watson-Crick base pairing (2,7,33).…”

“…poly(dG-dC) in solution, the nature and the concentration of various ions has been previously studied by U.V. absorbance and circular dichroism (1,(16)(17)(18)(19). On the other hand, the important role played in vivo and in vitro by metal ions in processes involving nucleic acids has been investigated for many years (for review, see ref.…”

Cationic porphyrins as probes of DNA structure

“…While the early work in the area employed simple aquated metal salts in studies with DNA (1) more recent efforts have utilized metal compounds containing a variety of different types of coordinated ligands. For example, cationic complex ions containing coordinated ammonia and various substituted o-phenanthrolines have been found useful in studies of DNA conformation (2)(3)(4). With interest stimulated largely by the antitumor antibiotic bleomycin (5), metal complexes have also been designed which can cleave DNA.…”

2'-deoxy-5-azacytidine increases binding of cisplatin to DNA by a mechanism independent of DNA hypomethylation

“…Alternatively, topologic changes may be involved. Stretches of alternating purines and pyrimidines are known to adopt the unusual left-handed Z DNA conformation (Wang et al, 1982) and this topologic form is stabilised by methylation of cytosines (Behe & Felsenfeld, 1981). Methylation of a critical promotor region may produce or stabilise a DNA conformation that is no longer recognised by the transcriptional machinery.…”