We here report the synthesis and characterization of small interfering RNAs with aryl trifluoromethyl diazirine moieties in the 3'-overhang regions, which allow sensitive detection of interacting proteins during assembly of the effector ribonucleoprotein complex by irradiation with minimally destructive long-wavelength ultraviolet light.
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The function of cellular RNA is modulated by a host of post-transcriptional chemical modifications installed by dedicated RNA-modifying enzymes. RNA modifications are widespread in biology, occurring in all kingdoms of life and in all classes of RNA molecules. They regulate RNA structure, folding, and protein-RNA interactions, and have important roles in fundamental gene expression processes involving mRNA, tRNA, rRNA, and other types of RNA species.
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Curr Protoc Nucleic Acid Chem
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Department of Chemistry, Princeton University, Princeton, New Jersey.
Post-transcriptional modifications play an important role in RNA biology. In particular, the addition of small chemical groups to the nucleobases of mRNA can affect how modified transcripts are processed in the cell, thereby impacting gene expression programs. In order to study the molecular mechanisms underlying these modifications, it is necessary to characterize their 'readers', that is, proteins that directly bind to these modifications to mediate their functional consequences; this is a major challenge because we lack approaches to precisely manipulate RNA chemistry in the cell and because protein-modified RNA interactions can be low affinity.
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Bioorg Med Chem Lett
September 2018
United Graduate School of Agricultural Science, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan; Faculty of Applied Biological Sciences, 1-1 Yanagido, Gifu 501-1193, Japan; Center of Highly Advanced Integration of Nano and Life Sciences, Gifu University (G-CHAIN), 1-1 Yanagido, Gifu 501-1193, Japan. Electronic address:
We designed and synthesized a photo-reactive and tag-free RNA probe for the identification of microRNA (miRNA) targets. To synthesize the RNA probe, we designed a novel nucleoside analog 1-O-[3-ethynyl-5-(3-trifluoromethyl-3H-diazirine-3-yl)]benzyl-β-d-ribofuranose containing aryl trifluoromethyl diazirine and ethynyl moieties. The RNA probe containing this analog was observed to form crosslinks with complementary RNA by UV irradiation and was rapidly tagged by Cu-catalyzed azide alkyne cycloaddition (CuAAC).
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J Am Chem Soc
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Department of Chemistry, Princeton University, Princeton, New Jersey 08544, United States.
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J Org Chem
March 2014
Course of Applied Life Science, Faculty of Applied Biological Sciences and ‡United Graduate School of Agricultural Science, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.
Here, we report the applicability of diazirine-containing RNA photo-cross-linking probes for the identification of microRNA (miRNA) targets. The RNA cross-linking probes were synthesized by substituting the RNA nucleobases with nucleoside analogues such as 1-O-[3-(3-trifluoromethyl-3H-diazirin-3-yl)]benzyl-β-d-ribofuranose or 1-O-[4-(3-trifluoromethyl-3H-diazirin-3-yl)]benzyl-β-D-ribofuranose that carry aryl trifluoromethyl diazirine moieties. The probes were successfully cross-linked with synthetic RNAs containing the four natural nucleosides on the opposite site of the nucleoside analogues.
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