Metal nanocluster-modified nucleotides

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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.