Ultramild protein-mediated click chemistry creates efficient oligonucleotide probes for targeting and detecting nucleic acids.
Functionalized synthetic oligonucleotides are finding growing applications in research, clinical studies, and therapy. However, it is not easy to prepare them in a biocompatible and highly efficient manner. We report a new strategy to synthesize oligonucleotides with promising nucleic acid targeting and detection properties. We focus in particular on the pH sensitivity of these new probes and their high target specificity. For the first time, human copper(I)-binding chaperon Cox17 was applied to effectively catalyze click labeling of oligonucleotides. This was performed under ultramild conditions with fluorophore, peptide, and carbohydrate azide derivatives. In thermal denaturation studies, the modified probes showed specific binding to complementary DNA and RNA targets. Finally, we demonstrated the pH sensitivity of the new rhodamine-based fluorescent probes in vitro and rationalize our results by electronic structure calculations.
Department of Physics and Chemistry, University of Southern Denmark, Campusvej 55, 5230 Odense M (Denmark).
Lina J
LJ
Nåbo
Department of Chemistry, University of Copenhagen (Denmark), Thorvaldsensvej 40, 1871 Copenhagen (Denmark).
Charlotte S
CS
Madsen
Department of Chemistry, University of Copenhagen (Denmark), Thorvaldsensvej 40, 1871 Copenhagen (Denmark).
Knud J
KJ
Jensen
Department of Physics and Chemistry, University of Southern Denmark, Campusvej 55, 5230 Odense M (Denmark).
Jacob
J
Kongsted
Department of Physics and Chemistry, University of Southern Denmark, Campusvej 55, 5230 Odense M (Denmark). ias@sdu.dk.
Kira
K
Astakhova
D001386:Azides / Q000737:chemistry; D001483:Base Sequence; D002352:Carrier Proteins / Q000378:metabolism*; D057930:Click Chemistry; D000079922:Copper Transport Proteins; D004247:DNA / Q000032:analysis* / Q000737:chemistry*; D006801:Humans; D015345:Oligonucleotide Probes / Q000737:chemistry* / Q000235:genetics; D010455:Peptides / Q000737:chemistry; D012313:RNA / Q000032:analysis* / Q000737:chemistry*
D001386:Azides; C491006:COX17 protein, human; D002352:Carrier Proteins; D000079922:Copper Transport Proteins; D015345:Oligonucleotide Probes; D010455:Peptides; D012313:RNA; D004247:DNA
click chemistry; copper-binding proteins; fluorescent probes; nucleic acid hybridization; nucleotides