背景:生物标志物检测策略有,近年来,一直在朝着适体形式的基于核酸的检测系统发展,在临床前和研究环境中显示出希望的短寡核苷酸序列。一种这样的适体是M5-15,一种针对人α突触核蛋白(α-syn)产生的DNA适体,所述α-syn是路易体和帕金森病(PD)相关的痴呆的致病因子。虽然这个适体显示出了希望,计算机模拟方法已经证明了产生对其靶标具有比体外产生的序列更高亲和力的适体的能力。方法:使用DINAMelt服务器-QuickFold网络服务器,根据其热力学稳定性筛选Python脚本随机生成的DNA序列库。用MFold检查所选择的序列以产生二级结构数据,所述二级结构数据用于使用RNA合成器软件产生3D数据。更进一步,校正结构,用DNA替换RNA,利用CSD-Discovery-GOLD软件进行一系列分子对接实验,对α-syn适体进行虚拟筛选.结果:在本文中,我们提出了一种称为TMG-79的替代方法,与M5-15相比,它对靶标具有更大的亲和力(M5-15=-15.9kcal/mol,TMG-79=-17.77kcal/mol)以及顶部姿势之间更好的ChemPLP适应性评分(M5-15=32.33,TMG-79=53.32)。结构分析表明,虽然有相似之处,与M5-15相比,TMG-79的更大的潜在柔性可以促进对α-syn的更大亲和力。结论:适体产生的计算机模拟方法具有彻底改变适体设计领域的潜力。我们认为TMG-79的进一步开发和体外验证将使其成为未来诊断和研究使用的可行候选者。
Background: Biomarker detection strategies have, in recent years, been moving towards nucleic acid-based detection systems in the form of aptamers, short oligonucleotide sequences which have shown promise in pre-clinical and research settings. One such aptamer is M5-15, a DNA aptamer raised against human alpha synuclein (α-syn) the causative agent in Lewy body and Parkinson\'s disease (PD) associated dementia. While this aptamer has shown promise, in silico methodologies have demonstrated a capacity to produce aptamers that have higher affinities for their targets than in vitro generated sequences. Methods: A Python script random generated library of DNA sequences were screened based on their thermodynamic stability with the use of DINAMelt server-QuickFold web server. The selected sequences were examined with MFold in order to generate secondary structure data that were used to produce 3D data with the use of RNA composer software. Further on, the structure was corrected and RNA was replaced with DNA and the virtual screening for α-syn aptamer took place with a series of molecular docking experiments with the use of CSD-Discovery-GOLD software. Results: Herein we propose an alternative in silico generated aptamer we call TMG-79 which demonstrates greater affinity for the target compared to M5-15 (M5-15 = -15.9 kcal/mol, TMG-79 = -17.77 kcal/mol) as well as better ChemPLP fitness scoring between the top poses (M5-15 = 32.33, TMG-79 = 53.32). Structural analysis suggests that while there are similarities, the greater potential flexibility of TMG-79 could be promoting greater affinity for the α-syn compared to M5-15. Conclusions: In silico methods of aptamer generation has the potential to revolutionise the field of aptamer design. We feel that further development of TMG-79 and validation in vitro will make it a viable candidate for diagnostic and research use in the future.