新型嵌段寡聚核苷酸-纳米金复合探针上DNA分子识别过程的热力学与动力学研究

There have been great interest to combined (bio)polymers and nanoparticles into a superstructure, which the physical properties of inorganic nanomaterials and the chemical flexibility/specificity of polymers can be used for obtaining possible unique functionalities. One of critical challenges in this area is understanding the behavior of biomolecules on nano-interface, which has long been considered difficult due to the instability and the control of conformation and density of biomolecules on surface. In this project, we aim to study the thermodynamic and kinetic of DNA hybridization on nano-interface based on diblock-DNA-AuNPs conjugates, which we developed recently. Conjugates of DNA and gold nanoparticles (AuNPs) typically exploit the strong Au?S chemistry to self-assemble thiolated oligonucleotides at AuNPs. However, it remains challenging to precisely control the orientation and conformation of surface-tethered oligonucleotides and finely tune the hybridization ability. We recently reported a novel strategy for spatially controlled functionalization of AuNPs with designed diblock oligonucleotides that are free of modifications. We have demonstrated that poly adenine (polyA) can serve as an effective anchoring block for preferential binding with the AuNP surface, and the appended recognition block adopts an upright conformation that favors DNA hybridization. The lateral spacing and surface density of DNA on AuNPs can also be systematically modulated by adjusting the length of the polyA block. We hope our study based on this newly developed system could provide deep insight for understanding the behaviour of biomolecules on nano-interface and provide new materials for the construction of high active biological molecular recognition interface.

纳米生物材料研究领域中一个关键问题是如何将生物分子（如DNA、蛋白质等）作为"分子探针"连接到纳米材料表面，并且避免此过程中界面上生物分子活性的损失，形成高效的分子识别界面。本项目基于申请人发展的一种新型嵌段DNA-纳米金偶联体制备方法（JACS，2012，134，11876，JACS亮点文章），构建一系列密度和构型可控的DNA-纳米金偶联体,以此为基础对DNA在纳米界面上分子识别的热力学和动力学等物理化学性质及生物分子活性进行深入的探讨和分析，建立理论模型。和传统的巯基化学修饰方法相比，这种制备DNA-纳米金的方法可以独立调控DNA在纳米金表面上的密度而不影响其构型，因此所制备的DNA-纳米金偶联体非常适合作为模型证实、修正和阐释DNA在纳米界面上分子识别的物理化学性质，用于单因素调节分析。本项目有望为生物纳米体系的构建提供理论基础，为高活性生物分子识别界面的构建及生物传感提供新型材料。

了解生物分子在纳米界面的状态对于生物分析是至关重要的，然而由于生物分子的不稳定性以及难以控制其位置和装载密度，这一研究面对许多挑战。在此，我们研究了DNA在AuNPs表面杂交的热力学和动力学，旨在提高DNA分析的效率。我们利用近期提出的基于polyA的组装策略实现了在AuNPs表面对DNA分子进行精确、定量的控制。PolyA 在这里做为锚定区会优先黏附到AuNPs表面，附带的识别区利用其垂直的构象用以与目标DNA分子杂交。通过调整polyA区域的长度可以系统地对AuNPs表面DNA的侧间距和密度进行调控。我们发现AuNPs上的双链体的稳定性随着polyA区长度的增加而增强。当polyA区长度达到40个碱基时，该体系的热力学性质与双链体在溶液中的热力学性质相似。二嵌段DNA-AuNPs体系的杂交率随着polyA区的增长而增大。我们认为该体系的高稳定性和优越的杂交表现归功于polyA将DNA-DNA和DNA-AuNPs相互作用力降至最小。我们的研究进一步揭示了生物分子在纳米界面上的行为，为建立高效、高速的生物传感器开启了新的契机。