Metal nanocluster-modified nucleotides

在过去的二十年里，金属纳米团簇(NCS)在生物成像、生物传感、治疗应用和催化等领域取得了重大进展。贵金属纳米管如AgNCs和AuNCs，由于其无毒、生物相容性和光稳定性等优点，已经成为比有机染料和量子点更好的发光材料。硫醇修饰的AUNCs在催化、发光、化学传感等领域得到了广泛的研究，具有广阔的应用前景。DNA聚合酶接受染料修饰的脱氧核苷三磷酸(DNTPs)的能力被开发在许多重要的生物技术应用中。这些方法包括下一代测序方法、单分子测序以及DNA和聚合酶链式反应扩增产物的标记，例如用于微阵列分析。此外，利用DNA的固有性质与化学引入的功能相结合，提供了一种新的基于核酸的杂化材料的入口。尽管金属纳米管具有令人印象深刻的性质，但据我们所知，这些纳米管还没有被用作dNTPs的修饰和随后作为DNA聚合酶的底物。将以协作的方式解决这一问题。我们建议设计新型的核苷酸修饰的水溶性纳米簇，以便通过DNA聚合酶的作用来掺入和标记DNA。我们将用合适的硫醇基团作为配体来合成嘌呤和嘧啶，以获得所需的NC偶联物。在开发出有效的合成路线后，将对NC-dNTP和NC-核苷偶联物进行光谱表征。同时，将研究NC-dNTP结合物对DNA聚合酶的作用，目的是将NC修饰的核苷酸酶结合到新生的DNA中。如果需要，将对DNA聚合酶进行改造，以改善NC-dNTP结合物的底物性质。通过反复几轮的合成和物理和生化研究，基于NC-dNTP的系统将获得比现有系统更好的(光)物理和生化性能，并将具有广泛的应用前景。

In the last two decades, metal nanoclusters (NCs) have made significant advances in the field of bioimaging, biosensing, therapeutic applications, and catalysis. Noble metal NCs like AgNCs and AuNCs, owing to their non-toxicity, biocompatibility, and photostability, have emerged as better luminescent materials over organic dyes and quantum dots. Thiol-capped AuNCs have been extensively studied in the field of catalysis, luminescence, and chemical sensing and exhibit very promising properties, calling for broad applications. The capability of DNA polymerases to accept dye-modified deoxynucleoside triphosphates (dNTPs) is exploited in many important biotechnological applications. These include next-generation sequencing approaches, single molecule sequencing, and labeling of DNA and PCR amplificates e.g. for microarray analysis. Furthermore, utilizing the intrinsic properties of DNA in combination with chemically introduced functionalities, provide an entry to new classes of nucleic acids-based hybrid material. Despite the impressive properties of metal NCs, these have, up to our knowledge, not yet exploited as modification for dNTPs and subsequently as substrate for DNA polymerases. In a collaborative approach will address this issue. We propose to design novel water-soluble nanoclusters modified with nucleotides in order to incorporate and label DNA by the action of DNA polymerases. We will be synthesize purines and pyrimidines with a suitably disposed thiol group used as ligand for obtaining the desired NC conjugates. After development of efficient synthetic routes, the NC-dNTP and NC-nucleoside conjugates will be spectroscopically characterized. In parallel, the NC-dNTP conjugates will be investigated towards their action on DNA polymerases with the aim to enzymatically incorporate the NC-modified nucleotides into the nascent DNA. If required, DNA polymerases will be engineered in order to improve the substrate properties of the NC-dNTP conjugates. By iterative rounds of synthesis and physical and biochemical studies, NC-dNTP-based systems will be obtained that have superior (photo)physical and biochemical properties than the systems available now and will allow broad applications.