antiviral drugs; G-quadruplex; i-motif; nucleic acids; developmental biology; organic chemistry; virology; nucleosides; cell biology; fluorescent probes
Mata Guillaume, Schmidt Olivia P., Luedtke Nathan W. (2016), A fluorescent surrogate of thymidine in duplex DNA, in Chem. Commun.
, 52(25), 4718-4721.
Seyfried Martin S., Alzeer Jawad, Luedtke Nathan W. (2016), Molecular Design and Synthesis of a Planar Telomestatin Analogue, in European Journal of Organic Chemistry
, 2016(2), 367-372.
Neef Anne B., Pernot Lucile, Schreier Verena N., Scapozza Leonardo, Luedtke Nathan W. (2015), A Bioorthogonal Chemical Reporter of Viral Infection, in Angewandte Chemie International Edition
, 54(27), 7911-7914.
Mata Guillaume, Luedtke Nathan W. (2015), Fluorescent Probe for Proton-Coupled DNA Folding Revealing Slow Exchange of i-Motif and Duplex Structures, in Journal of the American Chemical Society
, 137(2), 699-707.
Rieder Ulrike, Luedtke Nathan W. (2014), Alkene-Tetrazine Ligation for Imaging Cellular DNA, in Angewandte Chemie International Edition
, 53(35), 9168-9172.
Neef Anne B., Luedtke Nathan W. (2014), An Azide-Modified Nucleoside for Metabolic Labeling of DNA, in ChemBioChem
, 15(6), 789-793.
Vummidi Balayeshwanth R., Noreen Faiza, Alzeer Jawad, Moelling Karin, Luedtke Nathan W. (2013), Photodynamic Agents with Anti-metastatic Activities, in ACS Chemical Biology
, 8(8), 1737-1746.
Mata Guillaume, Luedtke Nathan W. (2013), Synthesis and Solvatochromic Fluorescence of Biaryl Pyrimidine Nucleosides, in Organic Letters
, 15(10), 2462-2465.
Wang I-Hsuan, Suomalainen Maarit, Andriasyan Vardan, Kilcher Samuel, Mercer Jason, Neef Anne, Luedtke Nathan W., Greber Urs F. (2013), Tracking Viral Genomes in Host Cells at Single-Molecule Resolution, in Cell Host & Microbe
, 14(4), 468-480.
As compared to the extensive methodologies developed for proteins, relatively few imaging techniques are available for nucleic acids - all of which are limited by their low sensitivity to alternatively folded structures, large perturbations to native systems, and/or inability to be applied in unmodified cells and organisms. To address unresolved questions in developmental biology and virology, we propose the synthesis and evaluation of new fluorescent probes for characterizing the structure, function, and flow of nucleic acids in vivo. In this proposal, synthetic nucleosides are used to deliver bioorthogonal functional groups (azide, alkyne, vinyl, or aryl halide) into cellular or viral genomes by metabolic incorporation. Subsequent bioorthogonal chemical reactions will be used to further modify the DNA to introduce fluorophores, affinity tags, and/or photocrosslinking groups. In addition to the development of these chemical tools, we propose important biological experiments that have been difficult or even impossible to accomplish using existing technologies, for example: (1) Azide-alkyne “click” reactions that furnish highly fluorescent nucleobase analogs will be used in FRET experiments to probe for non-canonical DNA structures like G-quadruplexes in vivo. (2) Modified nucleosides that exhibit non-toxic labeling of cellular genomes will be used in DNA “birth dating” experiments to evaluate the controversial “immortal DNA strand” hypothesis in Zebrafish. (3) Virus-specific genomic labeling will be conducted using modified nucleotides that are incorporated into virus genomes by error-prone viral polymerases, but rejected by high-fidelity cellular polymerases. This way, the newly synthesized viruses can be selectively labeled and visualized in whole animals without the need for specific antibodies or genetically modified viruses. In addition to visualization, we also propose new crosslinking and capture techniques that will enable unambiguous characterization of the labeled nucleic acids and their associated proteins. By using purified viruses that contain chemically-labeled genomes to infect cells, we will generate a catalog of all cellular factors that interact with the viral genomes during early- and late-phase herpes virus replication. Together these studies will shed new light on virus-host interactions, and may reveal new opportunities for therapeutic intervention.