Conformation and dynamics of a left-handed Z-DNA hairpin: studies of d(CGCGCGTTTTCGCGCG) in solution

Abstract

The physical properties of the DNA oligomer d(CGCGCGTTTTCGCGCG) in solvents containing 4 M NaClO4 and 0.1 M NaCl were investigated by proton NMR, optical melting, and circular dichroism spectroscopy. Results of these investigations are as follows: (i) The DNA hexadecamer exists as a unimolecular hairpin in either high or low salt. (ii) In high salt the stem region of the hairpin is in the left-handed Z conformation. (iii) In either high or low salt, the duplex stem of the hairpin is stabilized against melting by approximately 40 degrees C compared to the linear core duplex. The added stability of the hairpin is entropic in origin. (iv) In high salt, as the temperature is elevated, the equilibrium structure of the duplex stem of the hairpin shifts from the Z to the B conformation before melting. (v) In low salt, when the DNA duplex exists in the B conformation, attachment of a T4 single-strand loop to one end only slightly decreases (by 14%) the correlation time of the CH5-CH6 interproton vector. In high salt, when the DNA duplex exists in the Z conformation, the correlation time of the CH5-CH6 interproton vector decreases by 51%. Since these viscosity-corrected correlation times are taken to be indicators of duplex motions on the nanosecond time scale, this result directly suggests a larger amplitude of these motions is present in the duplex stem of the hairpin when it exists in the Z conformation.

Thermal stability, structural features, and B‐to‐Z transition in DNA tetraloop hairpins as determined by optical spectroscopy in d(CG)3T4(CG)3 and d(CG)3A4(CG)3 oligodeoxynucleotides

“…The main results of this series of investigations can be briefly described as follows: In all cases the presence of the tetraloop leads to a considerable increase of thermal stability compared to that measured in a reference double helix containing the same base repeat as in the stem of hairpins. As previously mentioned,40 the added stability of the hairpin is of entropic origin. Based on UV and NMR results,37 the stem of d(CG) 5 T 4 (CG) 5 hairpin undergoes a B‐to‐Z formtransition in particular conditions: high concentration of Na + or Mg 2+ and dehydration (ethanol) conditions. NMR results have shown that the oligomer d(CG) 2 T 4 (CG) 2 forms a tetraloop hairpin with a B form stem 38. The use of distance geometry calculations have permitted the obtention of the T 4 tetraloop hairpin three‐dimensional (3D) structure (up to now the only existing structural model of this hairpin in solution).…”

“…The choice of the ‐TTTT‐ (hereafter referred to as T 4 ) tetraloop is related to the fact that it is one of the most documented and perhaps the unique one for which we find a substantial amount of thermodynamic and structural data in the literature 35–42. The analysis of the T 4 tetraloop hairpin was started by a systematic optical analysis of d(ATCCTA‐T n ‐TAGGAT) oligomers,35 with n = 1–5 35, 36.…”

“…In these studies, n = 4 was found to be the optimal number for T n loops in regard to their thermal stability. A series of other valuable publications have described the formation of T 4 tetraloop hairpins in d(CG) m T 4 (CG) m sequences, with m = 2, 3, and 5 37–42. In fact, the idea was to induce a B‐to‐Z form transition in the stem of these hairpins.…”

A 6 bp Z‐DNA hairpin binds two Zα domains from the human RNA editing enzyme ADAR1

“…In contrast, previous studies [9] utilized polyd(CG) to determine the a¤nity of ZK to Z-DNA. The two d(CG) 3 strands of this hairpin were tethered through a T 4 loop, which increases the melting temperature by 40³C ensuring that a stable duplex stem is present at room temperature [19]. Analytical ultracentrifugation veri¢ed that d(CG) 3 T 4 (CG) 3 shows the molecular weight of a hairpin (measured: 4850, theoretical: 4864) rather than that of higher molecular weight duplexes with T 4 bulges.…”

Free-energy landscape of a hyperstable RNA tetraloop

“…The Z-RNA, in particular, revealed many interesting facets about the capacity of our force field to recapitulate the heterogeneity of folded RNA motifs. The presence of an alternating purine-pyrimidine sequence and high salt concentration has been established as a prerequisite for Z-form backbones for both DNA (37) and RNA (5); additional studies of DNA tetraloops have shown B-to Z-backbone transitions in high salt concentrations (5 M NaCl) (38,39), and combinations of pressure and salt (6 kbar; 5 M NaCl) (46) have been shown to induce A to Z transitions in r(CG) 6 duplexes. From our analyses, pressure is suggested to be a destabilizing factor in the formation of Z-RNA; however, high local density of counterions clearly stabilizes Z-RNA, which was indicated by consistent ion association in increased pressure and consistent coordination within the stem minor groove.…”