Salt-induced Z-A-Z transition sequence in the mixed ribo-deoxyribo copolymer poly(rG-dC) X poly(rG-dC).

Abstract

The left-handed helical conformation of various polynucleotides has been seen in solution at higher salt concentration than has the right-handed helical conformation of the corresponding polymer. We report here, however, studies of conformational transitions in the mixed polymer poly-(rG-dC)-poly (rG-dC). This polynucleotide appears to exist in a Z conformation under low-salt conditions, converts to an A helix at intermediate salt, and reverts to a left-handed helix at high-salt concentration.The left-handed helical Z conformation (1) has been demonstrated to occur in the synthetic polydeoxynucleotide poly-(dG-dC)-poly(dG-dC) when the concentration of NaCl in solution is increased to >2.5 M (2). High concentrations of other mono-and oligovalent cations also can promote the transition (2, 3). Much smaller concentrations (<1 mM) of oligovalent ions are needed to cause the methylated analog of the above polymer, poly(dG-m5dC)-poly(dG-m5dC), to flip to the Z form (3). Other chemical modifications of poly-(dG-dC)poly(dG-dC), such as bromination (4) or alkylation (5-7), also can lower the critical concentration of cation necessary for the transition. However, in all of these examples, and also in the case of alternating purine-pyrimidine polymers containing A-T base pairs (8, 9), the Z conformation is the "high-salt" form-i.e., if the salt concentration is decreased, the conformational equilibrium is shifted toward the right-handed helical B form. An explanation that has been offered for this is that phosphate residues on different strands approach each other more closely in the Z form than in the B form, so that the Z conformation is favored by the electrostatic shielding afforded by higher salt concentrations (1). Another argument for the high-salt conformation being the Z form comes from the observation that Z DNA is less hydrated than B DNA (10), so that the Z conformation is favored by the lowered water activity of high-salt solutions.We report here the study of conformational transitions in the mixed ribo-deoxyribo copolymer poly(rG-dC)-poly(rGdC). We chose to study this polymer because ribonucleotides favor the C3'-endo conformation, and this sugar conformation was reported to occur in the guanosine residues of Z-DNA crystals (1). We have determined that, unlike previously studied polynucleotides, poly(rG-dC)-poly(rG-dC) appears to exist in a Z conformation at low-salt concentration (5 mM Tris), converts to the A conformation at intermediate levels of counterion, and again flips to a Z form at high NaCl concentration. MATERIALS AND METHODSSynthesis of the Polymer. Escherichia coli DNA polymerase I large fragment (Klenow fragment) was purchased either from Bethesda Research Laboratories or New England Biolabs. Poly(dI-dC)poly(dI-dC) was obtained from P-L Biochemicals, as were NTPs and dNTPs. Radioactive materials were purchased from ICN.The synthesis of the polymer was carried out as follows. The reaction medium contained in 1 ml 40 mM N,N-bis(2-hydroxyethyl)glycine (Bicine) buffer (pH 8.95), 5 mM Mn C12...

Nuclear aggregates of polyamines are supramolecular structures that play a crucial role in genomic DNA protection and conformation

“…Another important consequence of s‐NAP complexation into m‐NAP is the strong increase of electrostatic forces that the latter compound exerts on DNA. It is known that electrostatic forces play a greater role in the A–Z transition than they do in the B–Z transition, because the difference in the linear charges density is greater between the A and the Z forms than between the B and the Z forms [51]. For this reason, we constructed a model in which s‐NAP interacts with A‐DNA and, among the non‐Z→Z transitional possibilities, we chose the A→Z possibility (Fig.…”

Nuclear aggregates of polyamines

“…The aggregation of s-NAP into m-NAP, however it occurs, brings about a significant increase in the charge on the DNA. It was shown that electrostatic forces play a greater role in A -Z transition than they do in B -Z transition, because the difference in linear charge density is greater between the A and the Z forms than between the B and the Z forms (18). Thus, we presented a model in which s-NAP interacts with A-DNA, and the s-NAP aggregation into (or its replacement with) m-NAP induces an A to Z transition on DNA (Fig.…”

Influence of tetraalkyl ammonium lons on the structure of poly (rG-dC).poly (rG-dC): unexpected transitions among the Z, A and B conformations

“…The far ultraviolet CD spectrum of poly (rG-dC)Opoly (rG-dC) in the presence of (tetramethyl ammonium ion] ranging from 0.5 mM to 3 M is very similar to that for the Z form of the polynucleotide previously seen in solutions of very low or very high [NaCl] (1). To be certain that the CD spectrum did reflect a Z form the circular dichroism spectrum of the polymer was recorded down to 180 nm.…”

“…Instead, as shown in Fig. 2, both ions caused the transition of the polymer from the low salt Z form to the A form at about 2-5 mM counterion concentration, more efficiently than the primary amine Tris (1). Tetrabutyl amonium ion, the largest counterion we tested, would not support the low K.…”

“…In the present study we examine the effect of tetraalkyl ammonium ions on poly (rG-dC)Opoly (rG-dC), a polymer containing 50% ribose and 50% deoxyribose residues. This polymer was previously shown to undergo two right-left hand helical transitions at different concentrations of NaCl: an A to high salt Z transition and an A to low salt Z transition (1). The presence of moderate concentrations of tetraalkyl ammonium ions causes a re-ordering of transitions of the polymer, and an additional conformational transition, to a B form, is observed.…”