G-triplex structure and formation propensity

Abstract

The occurrence of a G-triplex folding intermediate of thrombin binding aptamer (TBA) has been recently predicted by metadynamics calculations, and experimentally supported by Nuclear Magnetic Resonance (NMR), Circular Dichroism (CD) and Differential Scanning Calorimetry (DSC) data collected on a 3′ end TBA-truncated 11-mer oligonucleotide (11-mer-3′-t-TBA). Here we present the solution structure of 11-mer-3′-t-TBA in the presence of potassium ions. This structure is the first experimental example of a G-triplex folding, where a network of Hoogsteen-like hydrogen bonds stabilizes six guanines to form two G:G:G triad planes. The G-triplex folding of 11-mer-3′-t-TBA is stabilized by the potassium ion and destabilized by increasing the temperature. The superimposition of the experimental structure with that predicted by metadynamics shows a great similarity, with only significant differences involving two loops. These new structural data show that 11-mer-3′-t-TBA assumes a G-triplex DNA conformation as its stable form, reinforcing the idea that G-triplex folding intermediates may occur in vivo in human guanine-rich sequences. NMR and CD screening of eight different constructs obtained by removing from one to four bases at either the 3′ and the 5′ ends show that only the 11-mer-3′-t-TBA yields a relatively stable G-triplex.

Divalent cations and molecular crowding buffers stabilize G-triplex at physiologically relevant temperatures

“…Under the same conditions, TBA11 in presence of K + revealed a similar CD spectrum profile with a positive peak around 288 and a negative peak around 264 nm, indicating the formation of antiparallel G-triplex by TBA11. These features are identical to those reported by Limogelli et al in which G-triplex formation was confirmed by NMR investigations78. However, it should be noted that the CD signal intensity of the TBA11 G-triplex was much lower than that of the TBA G-quadruplex, suggesting the probability of G-triplex formation is less than that of G-quadruplex.…”

“…The truncated form of TBA, TBA11, has been shown to form a G-triplex structure by Limogelli and co-workers previously78. The CD spectra of TBA in a dilute solution containing K + or Na + had a positive peak at approximately 292 nm and a negative peak at approximately 269 nm (Figure 2).…”

“…A G-rich sequence with only three G-tracts may form a G-triplex with stacked G-triads, in which a central guanine connects with two other guanines by Hoogsteen-like hydrogen-bonds (Figure 1)78910. The formation of these G-triplexes has been hypothesized111213, and experiments have suggested that they might be intermediates during folding or unfolding of G-quadruplexes91314151617.…”

“…The formation of these G-triplexes has been hypothesized111213, and experiments have suggested that they might be intermediates during folding or unfolding of G-quadruplexes91314151617. Recently, Limongelli et al demonstrated the formation of G-triplexes in the G-rich sequence, 5'-GGTTGGTGTGG-3', using nuclear magnetic resonance (NMR) experiment in a dilute solution78. However, the melting temperature of the observed G-triplex was 33.5°C, suggesting that the structure is not stable at physiological temperatures.…”

Discovery of “folded DNA” structures in human cells: Potential drug targets

“…A guanine‐rich sequence consisting of only three adjacent guanines can form a fold, in this case a stack consisting of a central guanine hydrogen‐bonded to two other guanines . These may constitute independent folds, or they might instead be intermediates that are formed during the folding and unfolding of G‐quadruplexes .…”