The effect of l-thymidine, acyclic thymine and 8-bromoguanine on the stability of model G-quadruplex structures

Abstract

Background: Guanine-rich oligonucleotides are capable of forming tetrahelical structures known as G-quadruplexes with interesting biological properties. We have investigated the effects of site-specific substitution in the loops and in the tetrads model G-quadruplexes using thymine glycol nucleic acid (GNA) units, L-thymidine and 8-Br-2'-deoxyguanosine.Methods: Modified oligonucleotides were chemically synthesized and spectroscopic techniques were used to determine the relative stability of the modified G-quadruplex. The double 8-BrdUmodified quadruplexes were further characterized by Nuclear Magnetic Resonance. Binding to thrombin of selected quadruplex was analyzed by gel electrophoresis retention assay. Results:The most interesting results were found with a 8-bromoG substitution that had the larger stabilization of the quadruplex. NMR studies indicate a tight relationship between the loops and the tetrads to accommodate 8-bromoG modifications within the TBA. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 single substitution with the highest thermal stabilization found in thrombin binding aptamers modified at the guanine residues and having good affinity for thrombin. Double 8-BrdG modification in anti positions of different tetrads produce a conformational flip from syn to anti conformation of 8-Br-dG to favor loop-tetrad interaction and preserve the overall TBA stability. ConclusionsGeneral Significance: Modified guanine-rich oligonucleotides are valuable tools for the search for G-quadruplex structures with higher thermal stability and may provide compounds with interesting protein-nucleic acid binding properties.

In What Ways Do Synthetic Nucleotides and Natural Base Lesions Alter the Structural Stability of G-Quadruplex Nucleic Acids?

“…In certain cases, br 8 dG could stabilize a GQ also in anti G positions. Aviñó et al [ 94 ] substituted the TBA GQ ( Figure 2 , 7 ) with br 8 G, which stabilized the GQ when substituted for the syn position G5, by 7.3°C, and interestingly also by the double substitutions for G2, G11, by 3.6°C, which were both anti positions. The TBA GQ remained antiparallel.…”

“…The TBA GQ remained antiparallel. The analog destabilized the GQ in the anti position G2 by 2.5°C and also by double substitutions of the anti positions G2 and G6 by 8.8°C [ 94 ].…”

“…The analog destabilized the TBA in positions in T4,T9 by 9–13°C, and stabilized it in T7 by 5.2°C. With double substitutions in T7,9, T3,12, and T4,T13, the TBA GQ was destabilized by 2–12°C [ 94 ]. Aaldering et al [ 57 ] reported on the impact of uU and the 3′-amino-uU on the structural dynamics and stability of TBA GQ by substituting them for the (nonstacking) loops T3, T7, and T12.…”

“…Thermal stabilities depended on the position of the L-nucleotides and the inversion sites [ 184 ]. Aviñó et al substituted the TBA thymines with L-Ts in loop positions 3, 4, and 7, and all destabilized the GQ by 0.7 to 4°C [ 94 ].…”

Impact of the Position of the Chemically Modified 5-Furyl-2′-Deoxyuridine Nucleoside on the Thrombin DNA Aptamer–Protein Complex: Structural Insights into Aptamer Response from MD Simulations

“…To better understand the behavior of internal fluorophores and aid the strategic optimization of new probes, molecular level information is required about the influence of the modification on the stability and structure of the unbound aptamer and target–aptamer complex. In this light, biochemical assays and biophysical (NMR, X-ray crystallography) studies have played critical roles in unveiling the molecular details of modified TBA–thrombin interactions [21,29,34,36,37,38,39,40,41,42,43,44,45,46,47,48,49]. For example, high-resolution X-ray crystal structures have been obtained for the modified TBA containing 5-(methyl-3-acetyl-3-amino-1-propenyl)-2′-deoxyuridine at the TT or TGT loops, both of which explain the seven-fold increase in the binding affinity relative to the native TBA based on stabilizing contacts of the T4-modified base at the aptamer–protein interface [21].…”