François J C, Hélène C
Laboratoire de Biophysique, Muséum National d'Histoire Naturelle, INSERM U 201, CNRS UA 481, Paris, France.
Biochemistry. 1995 Jan 10;34(1):65-72. doi: 10.1021/bi00001a008.
A new approach is described to design antisense oligonucleotides targeted against single-stranded nucleic acids containing hairpin structures by use of both Watson-Crick and Hoogsteen hydrogen bond interactions for recognition. The oligonucleotide has two different domains, one allowing double helix formation involving Watson-Crick base pairs and the other one forming a triple helix involving Hoogsteen-type base triplets in the major groove of a hairpin stem. Spectroscopic and gel retardation experiments provided evidence for such Watson-Crick/Hoogsteen (WC/H) recognition of hairpin structures in single-stranded DNA. An antisense oligonucleotide designed to form only Watson-Crick base pairs was unable to disrupt the stable stem structure of the target under conditions where the oligonucleotide designed with the Watson-Crick/Hoogsteen interactions could bind efficiently to the hairpin-containing target. The addition of one nucleotide to the oligonucleotide at the junction between the double helix and triple helix regions in WC/H complexes had an effect on stability which was dependent on the relative orientation of the Watson-Crick and Hoogsteen domains in the target. An oligodeoxynucleotide-phenanthroline conjugate targeted against such a hairpin-containing DNA fragment induced specific cleavage in the double-stranded stem. This WC/H approach may be useful in designing artificial regulators of gene expression.
本文描述了一种新方法，通过利用沃森-克里克和 hoogsteen 氢键相互作用进行识别，来设计针对含有发夹结构的单链核酸的反义寡核苷酸。该寡核苷酸有两个不同结构域，一个允许通过沃森-克里克碱基对形成双螺旋，另一个在发夹茎的大沟中形成涉及 hoogsteen 型碱基三联体的三螺旋。光谱和凝胶阻滞实验为单链 DNA 中发夹结构的这种沃森-克里克/hoogsteen（WC/H）识别提供了证据。在寡核苷酸与发夹结构靶标能有效结合的条件下，设计仅形成沃森-克里克碱基对的反义寡核苷酸无法破坏靶标的稳定茎结构。在 WC/H 复合物的双螺旋和三螺旋区域交界处向寡核苷酸添加一个核苷酸，对稳定性有影响，这取决于靶标中沃森-克里克和 hoogsteen 结构域的相对取向。针对这种含发夹结构的 DNA 片段设计的寡脱氧核苷酸-菲咯啉缀合物在双链茎中诱导特异性切割。这种 WC/H 方法可能有助于设计基因表达的人工调节因子。
