A dumbbell-shaped, double-hairpin structure of DNA: a thermodynamic investigation - PubMed (original) (raw)
. 1987 Nov 3;26(22):7150-9.
doi: 10.1021/bi00396a042.
Affiliations
- PMID: 3427065
- DOI: 10.1021/bi00396a042
A dumbbell-shaped, double-hairpin structure of DNA: a thermodynamic investigation
D Erie et al. Biochemistry. 1987.
Abstract
We report the first calorimetric and spectroscopic investigation on a member of a new class of nucleic acid secondary structures in which both ends of a duplex core are closed by single-stranded loops. Such structures can be formed intramolecularly from appropriately designed base sequences. We have synthesized the 24-mer sequence shown, and we present calorimetric, spectroscopic, and electrophoretic (formula; see text) evidence that it adopts a dumbbell-shaped, double-hairpin structure. Our data allow us to reach the following conclusions: (1) The phosphodiester gap in the center of the core duplex of the dumbbell does not reduce the transition enthalpy relative to that measured for the corresponding octameric duplex d(GGAATTCC)2. (2) Incorporation of a 5'-phosphate group into the gap decreases the thermal stability of the dumbbell relative to its unphosphorylated sequence. On the basis of the salt dependence of this effect, we propose that the phosphorylation--induced decrease in thermal stability is electrostatic in origin. From the changes in the transition enthalpy and entropy, we suggest that the phosphorylation-induced decrease in thermal stability of the double hairpin arises from electrostatically induced based unstacking at the nick. (3) The thymine residues in the loop behave both electrostatically and enthalpically like denatured single strands. Published nuclear magnetic resonance studies reveal partial stacking of thymine residues in the loops of linear hairpin structures. If this feature persists in the double-hairpin structure, then the spatial overlap of thymine residues in the loops does not necessarily produce a favorable enthalpic contribution. (4) When both ends of the nicked octameric core duplex are constrained by loops of only four thymine residues, the dumbbell structure may adopt conformations in which the 5' and 3' ends at the nick are twisted relative to the helical axis and therefore are not in phase. Such conformations would account for the observed resistance of the double-hairpin structure to ligation, since the 3'OH and 5'P would no longer be collinear.
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