Lai Xiangdong, Zhao Weiwei, Jiang Lihua, Li Jing, Munawer Muhammad Faizan, Lai Jiejuan, Juarez Jesus Alejandro Martinez, Ud Din Miraj, Zhang Xiaoyang, Song Zhongquan, Wu Tao, Ge Yiyue, Jiang Hui, Liu Xiaohui, Wang Xuemei
State Key Laboratory of Digital Medical Engineering, School of Biological Science and Medical Engineering, Southeast University, Nanjing, 210096, China.
State Key Laboratory Breeding Base for The Protection and Utilization of Biological Resources in Tarim Basin, College of Life Science and Technology, Tarim University, Alar, Xinjiang, 843300, China.
Mater Today Bio. 2025 Jun 23;33:102020. doi: 10.1016/j.mtbio.2025.102020. eCollection 2025 Aug.
Both the limited research about structure-function relationship and the ill-defined process of conformational dynamic change greatly impede the development of aptamer engineering transformation and seriously restrict the practical applications of aptamers. In this work, an optimization strategy combining exonuclease III (Exo III) digestion and in silico simulation was presented for the first time for constructing high-affinity and functional aptamers and clarifying the three-dimensional (3D) structure of aptamer-target complexes and the conformational dynamic conversion in the process of aptamer recognizing its target. As a demonstration, the parent aptamer (Apt2) against SARS-CoV-2 spike subunit 1 (S1) was mutated or truncated at the predicted binding sites to produce eight derivatives (Seq1-Seq8). The progeny Seq3 exhibited a higher affinity for S1 and a better blocking effect on S1-angiotensin-converting enzyme 2 (ACE2) interaction compared to Apt2. Subsequently, Seq3 sealed the pores of nickel-doped zeolitic imidazolate framework-8 (NZIF-8) loaded with Rhodanine (Rho) to fabricate the aptasensor (NZIF-8-Rho-Apt) for inactivated virus detection, showing excellent performances in spiked actual samples. Therefore, this post systematic evolution of ligands by exponential enrichment (post-SELEX) is a very effective and general strategy for acquiring functionally-optimized aptamers.
关于结构-功能关系的有限研究以及构象动态变化过程的不明确，极大地阻碍了适体工程转化的发展，并严重限制了适体的实际应用。在这项工作中，首次提出了一种将核酸外切酶III（Exo III）消化与计算机模拟相结合的优化策略，用于构建高亲和力和功能性适体，并阐明适体-靶标复合物的三维（3D）结构以及适体识别其靶标的过程中的构象动态转换。作为示范，针对严重急性呼吸综合征冠状病毒2（SARS-CoV-2）刺突亚基1（S1）的亲本适体（Apt2）在预测的结合位点进行突变或截短，以产生八个衍生物（Seq1-Seq8）。与Apt2相比，子代Seq3对S1表现出更高的亲和力，并且对S1-血管紧张素转换酶2（ACE2）相互作用具有更好的阻断作用。随后，Seq3封闭了负载罗丹宁（Rho）的镍掺杂沸石咪唑框架-8（NZIF-8）的孔，以制造用于灭活病毒检测的适体传感器（NZIF-8-Rho-Apt），在加标的实际样品中显示出优异的性能。因此，这种指数富集配体的后筛选（post-SELEX）是获得功能优化适体的一种非常有效且通用的策略。
