The effect of sodium, potassium and ammonium ions on the conformation of the dimeric quadruplex formed by the Oxytricha nova telomere repeat oligonucleotide d(G4T4G4)

Abstract

The DNA sequence d(G(4)T(4)G(4)) [Oxy-1.5] consists of 1.5 units of the repeat in telomeres of Oxytricha nova and has been shown by NMR and X-ray crystallographic analysis to form a dimeric quadruplex structure with four guanine-quartets. However, the structure reported in the X-ray study has a fundamentally different conformation and folding topology compared to the solution structure. In order to elucidate the possible role of different counterions in this discrepancy and to investigate the conformational effects and dynamics of ion binding to G-quadruplex DNA, we compare results from further experiments using a variety of counterions, namely K(+), Na(+)and NH(4)(+). A detailed structure determination of Oxy-1.5 in solution in the presence of K(+)shows the same folding topology as previously reported with the same molecule in the presence of Na(+). Both conformations are symmetric dimeric quadruplexes with T(4)loops which span the diagonal of the end quartets. The stack of quartets shows only small differences in the presence of K(+)versus Na(+)counterions, but the T(4)loops adopt notably distinguishable conformations. Dynamic NMR analysis of the spectra of Oxy-1.5 in mixed Na(+)/K(+)solution reveals that there are at least three K(+)binding sites. Additional experiments in the presence of NH(4)(+)reveal the same topology and loop conformation as in the K(+)form and allow the direct localization of three central ions in the stack of quartets and further show that there are no specific NH(4)(+)binding sites in the T(4)loop. The location of bound NH(4)(+)with respect to the expected coordination sites for Na(+)binding provides a rationale for the difference observed for the structure of the T(4)loop in the Na(+)form, with respect to that observed for the K(+)and NH(4)(+)forms.

Two-repeat Tetrahymena Telomeric d(TGGGGTTGGGGT) Sequence Interconverts Between Asymmetric Dimeric G-quadruplexes in Solution

“…23 Methyl and H6 protons of T6 and T7 were assigned by 13 C-filtered experiments using sitespecific 2% 13 C, 15 N-labeled samples 21,22 (see Figure S1 of the Supplementary Material).…”

“…[12][13][14][15][16][17][18] The first structure from this family was the Gquadruplex structure of the two-repeat Oxytricha telomeric d(G 4 T 4 G 4 ) sequence solved by NMR. 12,13 This sequence forms in the presence of various monovalent cations, including Na + or K + , a dimeric G-quadruplex with diagonal loops at opposite ends. The G-tetrad core of this Oxytricha G-quadruplex is similar to that of the Tetrahymena sequence from this study with respect to strand orientations, syn/anti distributions and hydrogen bond directionalities.…”

“…6 Eight samples (2% 15 N-labeled at individual G positions) and two samples (2% 15 N, 13 C-labeled at T6 or T7) were synthesized at a 2 μmol scale with lowenrichment labeled phosphoramidites prepared as described. 21,22 In all cases, samples were dialyzed successively against 50 mM NaCl and against water.…”

Characterization of Metal Ion-Nucleic Acid Interactions in Solution

“…Only very few metal binding motifs have been characterized so far that are able to slow down exchange so significantly. Among them are the strong binding sites in the stem of G-quadruplex structures, for which lifetimes can be exceptionally long on the NMR chemical shift timescale [227,228] and which have been studied extensively due to their compact size.…”

Highly Polymorphic G-quadruplexes in the c-MYC Promoter

“…Guanine tetrad is formed by Hoogsteen base pairing of four guanines, and stabilized by coordinated monovalent ions such as K + and Na + . 6,7 When a complementary C-rich strand coexists, additional stabilizing factors, for example negative-supercoiling, G-quadruplex stabilizing proteins or chemicals, are required for the formation of G-quadruplexes, which has higher Gibbs free energy than a Watson-Crick DNA duplex. 8 However, once it is formed, it can be maintained for a long time to perform its biological roles.…”