Campo, Cristian del ; Leeder, Wolf-Matthias ; Reißig, Paul ; Göringer, H. Ulrich (2020)
Analyzing editosome function in high-throughput.
In: Nucleic acids research, 48 (17)
doi: 10.1093/nar/gkaa658
Artikel, Bibliographie
Kurzbeschreibung (Abstract)
Mitochondrial gene expression in African trypanosomes and other trypanosomatid pathogens requires a U-nucleotide specific insertion/deletion-type RNA-editing reaction. The process is catalyzed by a macromolecular protein complex known as the editosome. Editosomes are restricted to the trypanosomatid clade and since editing is essential for the parasites, the protein complex represents a near perfect target for drug intervention strategies. Here, we report the development of an improved in vitro assay to monitor editosome function. The test system utilizes fluorophore-labeled substrate RNAs to analyze the processing reaction by automated, high-throughput capillary electrophoresis (CE) in combination with a laser-induced fluorescence (LIF) readout. We optimized the assay for high-throughput screening (HTS)-experiments and devised a multiplex fluorophore-labeling regime to scrutinize the U-insertion/U-deletion reaction simultaneously. The assay is robust, it requires only nanogram amounts of materials and it meets all performance criteria for HTS-methods. As such the test system should be helpful in the search for trypanosome-specific pharmaceuticals.
| Typ des Eintrags: | Artikel |
|---|---|
| Erschienen: | 2020 |
| Autor(en): | Campo, Cristian del ; Leeder, Wolf-Matthias ; Reißig, Paul ; Göringer, H. Ulrich |
| Art des Eintrags: | Bibliographie |
| Titel: | Analyzing editosome function in high-throughput. |
| Sprache: | Englisch |
| Publikationsjahr: | 25 September 2020 |
| Titel der Zeitschrift, Zeitung oder Schriftenreihe: | Nucleic acids research |
| Jahrgang/Volume einer Zeitschrift: | 48 |
| (Heft-)Nummer: | 17 |
| DOI: | 10.1093/nar/gkaa658 |
| Kurzbeschreibung (Abstract): | Mitochondrial gene expression in African trypanosomes and other trypanosomatid pathogens requires a U-nucleotide specific insertion/deletion-type RNA-editing reaction. The process is catalyzed by a macromolecular protein complex known as the editosome. Editosomes are restricted to the trypanosomatid clade and since editing is essential for the parasites, the protein complex represents a near perfect target for drug intervention strategies. Here, we report the development of an improved in vitro assay to monitor editosome function. The test system utilizes fluorophore-labeled substrate RNAs to analyze the processing reaction by automated, high-throughput capillary electrophoresis (CE) in combination with a laser-induced fluorescence (LIF) readout. We optimized the assay for high-throughput screening (HTS)-experiments and devised a multiplex fluorophore-labeling regime to scrutinize the U-insertion/U-deletion reaction simultaneously. The assay is robust, it requires only nanogram amounts of materials and it meets all performance criteria for HTS-methods. As such the test system should be helpful in the search for trypanosome-specific pharmaceuticals. |
| ID-Nummer: | pmid:32756897 |
| Fachbereich(e)/-gebiet(e): | 10 Fachbereich Biologie 10 Fachbereich Biologie > Genregulation und RNA-Therapeutika |
| Hinterlegungsdatum: | 31 Aug 2020 09:48 |
| Letzte Änderung: | 02 Aug 2021 13:34 |
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