Lee Hui-Ting, Arciniegas Santiago, Marky Luis A
Department of Pharmaceutical Sciences, University of Nebraska Medical Center, 986025 Nebraska Medical Center, Omaha, Nebraska 68198-6025, USA.
J Phys Chem B. 2008 Apr 17;112(15):4833-40. doi: 10.1021/jp710926h. Epub 2008 Mar 22.
Nucleic acid oligonucleotides (ODNs), as drugs, present an exquisite selectivity and affinity that can be used in antigene and antisense strategies for the control of gene expression. In this work we try to answer the following question: How does the molecularity of a DNA triplex affect its overall stability and melting behavior? To this end, we used a combination of temperature-dependent UV spectroscopy and calorimetric (differential scanning calorimetry) techniques to investigate the melting behavior of DNA triplexes with a similar helical stem, TC+TC+TC+T/AGAGAGA/TCTCTCT, but formed with different strand molecularity. We determined standard thermodynamic profiles and the differential binding of protons and counterions accompanying their unfolding. The formation of a triplex is accompanied by a favorable free energy term, resulting from the typical compensation of favorable enthalpy-unfavorable entropy contributions, i.e., the folding of a particular triplex is enthalpy driven. The magnitude of the favorable enthalpy contributions corresponds to the number and strength of the base-triplet stacks formed, which are helped by stacking contributions due to the incorporation of dangling ends or loops. Triplex stability is in the following order: monomolecular > bimolecular > trimolecular; this is explained in terms of additional stacking contributions due to the inclusion of loops. As expected, acidic pH stabilized all triplexes by allowing protonation of the cytosines in the third strand; however, the percentage of protonation increases as the molecularity decreases. The results help to choose adequate solution conditions for the study of triplexes containing different ratios of CGC+ and TAT base triplets and to aid in the design of oligonucleotide sequences as targeting reagents that could effectively react with mRNA sequences involved in human diseases, thereby increasing the feasibility of using the antisense strategy for therapeutic purposes.
核酸寡核苷酸（ODNs）作为药物，具有出色的选择性和亲和力，可用于反基因和反义策略来控制基因表达。在这项工作中，我们试图回答以下问题：DNA三链体的分子性如何影响其整体稳定性和熔解行为？为此，我们结合使用了温度依赖性紫外光谱法和量热法（差示扫描量热法），来研究具有相似螺旋茎TC+TC+TC+T/AGAGAGA/TCTCTCT但由不同链分子性形成的DNA三链体的熔解行为。我们确定了标准热力学曲线以及伴随其解折叠的质子和抗衡离子的差异结合。三链体的形成伴随着有利的自由能项，这是由有利的焓贡献和不利的熵贡献的典型补偿导致的，即特定三链体的折叠是由焓驱动的。有利的焓贡献的大小对应于形成的碱基三联体堆积的数量和强度，由于包含悬垂末端或环而产生的堆积贡献有助于此。三链体稳定性的顺序如下：单分子>双分子>三分子；这可以通过由于包含环而产生的额外堆积贡献来解释。正如预期的那样，酸性pH通过使第三条链中的胞嘧啶质子化来稳定所有三链体；然而，质子化百分比随着分子性的降低而增加。这些结果有助于为研究含有不同比例CGC+和TAT碱基三联体的三链体选择合适的溶液条件，并有助于设计作为靶向试剂的寡核苷酸序列，这些序列可以与涉及人类疾病的mRNA序列有效反应，从而提高将反义策略用于治疗目的的可行性。
