Biostable aptamers with antagonistic properties to the neuropeptide nociceptin/orphanin FQ
| Autor: | Clemens Gillen, Hermann Nawrath, Dirk Faulhammer, Bernd Eschgfäller, Sebastian D. Vulcu, Werner Englberger, Sven Klussmann, Petra Burgstaller, Sandra Stark, Stefan Vonhoff, Frank Kleinjung, Jeannette Erfurth, Werner Schröder, Johanna Rupp |
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| Rok vydání: | 2004 |
| Předmět: |
Xenopus
Aptamer Molecular Sequence Data GTPgammaS Neuropeptide Ligands Tritium Nociceptin Receptor Article chemistry.chemical_compound Animals Humans Receptor Molecular Biology Base Sequence biology Oligonucleotide biology.organism_classification In vitro Nociceptin receptor Opioid Peptides Biochemistry chemistry Receptors Opioid Oocytes Biophysics Nucleic Acid Conformation RNA |
| Zdroj: | RNA. 10:516-527 |
| ISSN: | 1469-9001 1355-8382 |
| DOI: | 10.1261/rna.5186504 |
| Popis: | The neuropeptide nociceptin/orphanin FQ (N/OFQ), the endogenous ligand of the opioid receptor-like 1 (ORL1) receptor, has been shown to play a prominent role in the regulation of several biological functions such as pain and stress. Here we describe the isolation and characterization of N/OFQ binding biostable RNA aptamers (Spiegelmers) using a mirror-image in vitro selection approach. Spiegelmers arel-enantiomeric oligonucleotide ligands that display high affinity and specificity to their targets and high resistance to enzymatic degradation compared tod-oligonucleotides. A representative Spiegelmer from the selections performed was size-minimized to two distinct sequences capable of high affinity binding to N/OFQ. The Spiegelmers were shown to antagonize binding of N/OFQ to the ORL1 receptor in a binding-competition assay. The calculated IC50values for the Spiegelmers NOX 2149 and NOX 2137a/b were 110 nM and 330 nM, respectively. The competitive antagonistic properties of these Spiegelmers were further demonstrated by their effective and specific inhibition of G-protein activation in two additional models. The Spiegelmers antagonized the N/OFQ-induced GTPγS incorporation into cell membranes of a CHO-K1 cell line expressing the human ORL1 receptor. In oocytes fromXenopus laevis,NOX 2149 showed an antagonistic effect to the N/OFQ-ORL 1 receptor system that was functionally coupled with G-protein-regulated inwardly rectifying K+channels. |
| Databáze: | OpenAIRE |
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