The effect of monovalent cations on the thermal stability of a small model DNA hairpin has been measured by capillary electrophoresis, using an oligomer with 16 thymine residues as an unstructured control. The melting temperature of the model hairpin increases approximately linearly with the logarithm of increasing cation concentration in solutions containing Na(+), K(+), Li(+), NH(4)(+), Tris(+), tetramethylammonium (TMA(+)), or tetraethylammonium (TEA(+)) ions, is approximately independent of cation concentration in solutions containing tetrapropylammonium (TPA(+)) ions, and decreases with the logarithm of increasing cation concentration in solutions containing tetrabutylammonium (TBA(+)) ions. At constant cation concentration, the melting temperature of the DNA model hairpin decreases in the order Li(+) ∼ Na(+) ∼ K(+) > NH(4)(+) > TMA(+) > Tris(+) > TEA(+) > TPA(+) > TBA(+). Isothermal studies indicate that the decrease in the hairpin melting temperature with increasing cation hydrophobicity is not due to saturable, site-specific binding of the cation to the random coil conformation, but to the concomitant increase in cation size with increasing hydrophobicity. Larger cations are less effective at shielding the charged phosphate residues in B-form DNA because they cannot approach the DNA backbone as closely as smaller cations. By contrast, larger cations are relatively more effective at shielding the phosphate charges in the random coil conformation, where the phosphate-phosphate distance more closely matches cation size. Hydrophobic interactions between alkylammonium ions interacting electrostatically with the phosphate residues in the coil may amplify the effect of cation size on DNA thermal stability.
Fluorescence spectroscopy studies of 7-Aminoactinomycin D (7-AMD) binding to dodecamers containing the Cre binding sequence demonstrate that binding affinity is directly related to both sequence context and cytosine methylation. Importantly, it was also found that methylation effects on binding affinity are sequence dependent. The native d[TTTCACGTGAAA] 2 and d[AA GAACGTTCTT] 2 samples show very similar binding affinities in their native forms, while native d[GAAAACGTTTTC] 2 has a dissociation constant fourfold higher than either of these samples. Upon methylation, d[TTTCA-m 5 -CGTGAAA] 2 shows the greatest change in the dissociation constant, increasing two-fold. The d[AAGAA-m 5 -CGTTCTT] 2 sequence has a 1.5-fold increase in the dissociation constant, while the d[GAAAACGTTTTC] 2 sequence shows no significant change in binding affinity upon methylation. NMR and FTIR evidence will be presented to further explain these phenomena and provide evidence that backbone conformations due to flanking sequence and methylation state are responsible for these differences.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.