Development of Abasic Site-Containing DNA Duplex Aptamers for Biologically Important Substrates

适配子是一种短小的核酸结合物种，由于它们能够结合大量的配体，包括氨基酸、含核糖的辅因子和药物，因此被认为是分子识别事件的候选对象。这些适配子对靶配体的高亲和力和特异性通常是通过分子形状互补、氢键和堆积作用的组合来实现的。这类结合事件通常涉及配体诱导的适体结构变化，导致形成与配体结合有关的独特二级结构，其中内环或茎环结构代表结合事件的活性部位。然而，为了进一步设计用于小配体的rna和dna适配子，尽管其他类型的非碱基配对部分具有结合口袋的潜力，但很少有人关注。本文报道了一类新的用于选择性和st…的dna双链适配子更多的与生物重要配体的结合，其中双链中的碱性(AP)位被用作与生物重要配体结合的活性腔，其中双链中的碱性(AP)位被用作结合事件的活性腔。对于基于AP位点的DNA适配子，人们强烈预期，通过在AP位点两侧堆积两个核苷酸，靶标配体被插入AP位点，并伴随着与AP位点相反的受体核苷酸形成氢键。事实上，在本研究项目中，我们已经成功地开发出对核黄素(解离常数K_d=1.9μM)、茶碱(K_d=31μM)、咖啡因(K_d=20μM)和腺苷(检测范围10~150μM)具有高选择性的这类适体。与典型的RNA适配子相比，基于AP位点的DNA适配子具有合成简单、成本低、化学稳定性高等优点。这些功能将允许各种类型的分析应用。较少

Aptamers, short nucleic acid-binding species, have emerged as promising candidates for molecular recognition events on account of their significant ability to bind large number of ligands, including amino acids, ribose-containing cofactors and drugs. High affinity and specificity of these aptamers toward target ligands are generally achieved by a combination of molecular shape complementarity, hydrogen-bonding and stacking interactions. Such binding events typically involve the ligand-induced structural changes of the aptamers, resulting in the formation of unique secondary structures responsible to the ligand binding, for which the internal - or stem-loop structures are representative of active sites for binding events. However, for the further design of both RNA and DNA aptamers for small ligands, little attention has been paid to other types of non-base-pairing moieties despite the potential as binding pockets.Here we report on a new class of DNA duplex aptamers for selective and st … More rong binding to biologically important ligands, where an abasic (AP) site in the duplex is utilized as an active cavity for binding to biologically important ligands, where an abasic (AP) site in the duplex is utilized as an active cavity for binding events. For the AP site-based DNA aptamers, it is strongly expected that, by stacking with two nucleotides flanking the AP site, a target ligand is intercalated into the AP site, and this is accompanied by the formation of hydrogen bonds with a receptor nucleotide opposite the AP site. Indeed, we have successfully developed this type of aptamers with high selectivity for riboflavin (dissociation constant K_d=1.9μM), theophylline (K_d=31μM), caffeine (K_d=20μM), and adenosine (detection range: 10~150μM) in this research project. As compared to typical RNA aptamers, our AP site-based DNA aptamers have some advantages, such as their easy and low-cost synthesis, and higher chemical stability. These features would allow various kinds of analytical applications. Less