Why Does Substitution of Thymine by 6-Ethynylpyridone Increase the Thermostability of DNA Double Helices?

Abstract

Efficiency of 6-ethynylpyridone (E), a potential thymine (T) analogue, which forms high-fidelity base pairs with adenine (A) and gives rise to stabler DNA duplexes, with stability comparable to those containing canonical cytosine(C):guanine(G) base pairs, has been reported recently. Estimates of the interaction energies, involving geometry optimization at the DFT level (including middle range dispersion interactions) followed by single point energy calculation at MP2 level, in excellent correlation with the experimentally observed trends, show that E binds more strongly and more discriminately with A than T does. Detailed analysis reveals that the increase in base-base interaction arises out of conjugation of acetylenic π electrons with the ring π system of E, which results in not only an extra stabilizing C-H···π interaction in the EA pair, but also a strengthening of the conventional hydrogen bonds. However, the computed base-base interaction energy for the EA pair was found to be much less than that of the canonical CG pair, implying that the difference in the TA versus EA base pairing interaction alone cannot explain the large experimentally observed increase in the thermostability of DNA duplexes, where a TA pair is replaced with an EA pair. Our computations show that the conjugation of acetylenic π electrons with the ring π system also possibly plays a role in increasing the stacking potential of the EA pair, which in turn can explain its marked influence in the enhancement of duplex stability.

Reverse Watson-Crick purine-purine base pairs — the Sharp-turn motif and other structural consequences in functional RNAs

“…For geometry optimization and subsequent interaction energy calculations, we have selected Truhlar's M05-2X 39 functional and the 6-31G++(2d,2p) basis set 40 . M05-2X is a hybrid meta GGA functional with 56% Hartree-Fock exchange component and is known to perform remarkably well for noncovalently bonded molecules of biological relevance, 41 including nucleobases [42][43][44][45][46][47][48][49] . 6-31G++(2d,2p) is a split valance double ζ type basis set, with two sets of d type polarization function for all non-hydrogen atoms and two sets of p type polarization function for hydrogen atoms, and also include s-p diffused orbitals for non-hydrogen atoms, was used for ground state geometry optimization.…”

Stacking with the unnatural DNA base 6-ethynylpyridone

“…This implies that the increased stability of E:A observed experimentally cannot be explained fully by base-pair interactions, and that other interactions must also contribute. One hypothesis put forward is that the increased stability observed is due to enhanced stacking interactions within the DNA duplex structure [16]. We also note here that the stability of DNA is largely due to stacking interactions, rather than H-bonding interactions [17].…”

“…Table 1 compares the CP-corrected M06-2X/6-31+G(d)/PCM interaction energies of the H-bonded base pairs with the CP-corrected MP2/aug-cc-pVDZ single-point energies by Halder et al [16]. The M06-2X and MP2 results are in close agreement.…”

“…These observations indicate that the stability of the E:A base pair almost equals that of a C:G pair. A subsequent DFT study found that E does indeed bind with adenine with more affinity than thymine does; however, the calculations showed that the stability of the E:A pair is intermediate between those of the canonical T:A and C:G base pairs [16]. Thus, the computed stability of E:A does not fully explain the remarkable increased stability (similar to the C:G base pair) observed in the experiments.…”

“…A) can be classified as H-bonds [16]. In addition, NBO (Natural Bond Orbital) analysis and IR frequency data suggest the N-H•••O interaction is stronger in E:A than in T:A [16].…”