Thermodynamic and kinetic effects of N3′→P5′ phosphoramidate modification on pyrimidine motif triplex DNA formation
Biochemistry, ISSN: 0006-2960, Vol: 40, Issue: 4, Page: 1063-1069
2001
- 23Citations
- 5Captures
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Metrics Details
- Citations23
- Citation Indexes23
- 23
- CrossRef16
- Captures5
- Readers5
Article Description
I have investigated the thermodynamic and kinetic effects of N3′→P5′ phosphoramidate (PN) backbone modification of triplex-forming oligonucleotide (TFO) on the pyrimidine motif triplex formation between a 23-bp target duplex and a 15-mer TFO using electrophoretic mobility shift assay, UV melting, isothermal titration calorimetry, and interaction analysis system. The thermodynamic and kinetic analyses have clearly indicated that the PN modification of TFO not only significantly increased the thermal stability of the pyrimidine motif triplex at neutral pH but also increased the binding constant of the pyrimidine motif triplex formation at room temperature and neutral pH by nearly 2 orders of magnitude. The consideration of the observed thermodynamic parameters has suggested that the more rigidity of the PN TFO in the free state relative to the unmodified TFO may enable the significant increase in the binding constant of the pyrimidine motif triplex formation at neutral pH. Kinetic data have also demonstrated that the observed PN modification-mediated promotion of pyrimidine motif triplex formation at neutral pH resulted from the considerable decrease in the dissociation rate constant rather than the increase in the association rate constant. This information will present an effective approach for designing chemically modified TFO with higher binding affinity in the triplex formation under physiological conditions, which may eventually lead to progress in therapeutic applications of the antigene strategy in vivo.
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