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Transition characteristics and thermodynamic analysis of DNA duplex formation: a quantitative consideration for the extent of duplex association.

ABSTRACT: Transition characteristics and thermodynamic properties of the single-stranded self-transition and the double-stranded association were investigated and analyzed for 9-, 15- and 21-bp non-self-complementary DNA sequences. The multiple transition processes for the single-stranded self-transition and the double-stranded association were further put forth. The experimental results confirmed that the double-stranded association transition was generally imperfect and the thermodynamic properties of the single-stranded self-transition would exert an influence on a duplex formation. Combining ultraviolet melting experiments in various molar ratios, the extent of duplex association was estimated for three double-stranded DNAs. In our experimental range, the extent of duplex association decreases with increasing the number of base pairs in DNA sequences, which suggest that the short oligonucleotides may proceed in a two-state transition while the long oligonucleotides may not. When the extent of duplex association was considered, the true transition enthalpies of a duplex formation derived from UV and differential scanning calorimetry measurements were in good agreement.

SUBMITTER: Wu P

PROVIDER: S-EPMC115173 | biostudies-literature | 2000 Dec

REPOSITORIES: biostudies-literature

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Transition characteristics and thermodynamic analysis of DNA duplex formation: a quantitative consideration for the extent of duplex association.

Wu P P Sugimoto N N

Nucleic acids research 20001201 23

Transition characteristics and thermodynamic properties of the single-stranded self-transition and the double-stranded association were investigated and analyzed for 9-, 15- and 21-bp non-self-complementary DNA sequences. The multiple transition processes for the single-stranded self-transition and the double-stranded association were further put forth. The experimental results confirmed that the double-stranded association transition was generally imperfect and the thermodynamic properties of t ...[more]

PMID: 11095688

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Project description:DNA interstrand cross-links have important biological consequences and are useful biotechnology tools. Phenylselenyl substituted derivatives of thymidine (1) and 5-methyl-2'-deoxycytidine (5) produce interstrand cross-links in duplex DNA when oxidized by NaIO4. The mechanism involves a [2,3]-sigmatropic rearrangement of the respective selenoxides to the corresponding methide type intermediates, which ultimately produce the interstrand cross-links. Determination of the rate constants for the selenoxide rearrangements indicates that the rate-determining step for cross-linking is after methide formation. Cross-linking by the thymidine derivative in duplex DNA shows a modest kinetic preference when flanked by pyrimidines as opposed to purines. In contrast, the rate constant for cross-link formation from 5 opposite dG in duplex DNA is strongly dependent upon the flanking sequence and, in general, is at least an order of magnitude slower than that for 1 in an otherwise identical sequence. Introduction of mispairs at the base pairs flanking 5 or substitution of the opposing dG by dI significantly increases the rate constant and yield for cross-linking, indicating that stronger hydrogen bonding between the methide derived from it and dG compared to dA and the respective electrophile derived from 1 limits reaction by increasing the barrier to rotation into the required syn-conformation. Incorporation of 1 or 5 in triplex forming oligonucleotides (TFOs) that utilize Hoogsteen base pairing also yields interstrand cross-links. The dC derivative produces ICLs approximately 10x faster than the thymidine derivative when incorporated at the 5'-termini of the TFOs and higher yields when incorporated at internal sites. The slower, less efficient ICL formation emanating from 1 is attributed to reaction at N1-dA, which requires local melting of the duplex. In contrast, 5 produces cross-links by reacting with N7-dG. The cross-linking reactions of 1 and 5 illustrate the versatility and utility of these molecules as mechanistic probes and tools for biotechnology.

Dataset Information

Transition characteristics and thermodynamic analysis of DNA duplex formation: a quantitative consideration for the extent of duplex association.

Publications

Transition characteristics and thermodynamic analysis of DNA duplex formation: a quantitative consideration for the extent of duplex association.

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