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Journal Article

ADLOC: an aptamer-displacement assay based on luminescent oxygen channeling

MPS-Authors

Famulok,  M.
External Organizations;
Max Planck Fellow Chemical Biology, Center of Advanced European Studies and Research (caesar), Max Planck Society;

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Citation

Niebel, B., Lentz, C., Pofahl, M., Mayer, G., Hoerauf, A., Pfarr, K. M., et al. (2010). ADLOC: an aptamer-displacement assay based on luminescent oxygen channeling. Chemistry, 16(36), 11100-7. doi:10.1002/chem.201001192.


Cite as: http://hdl.handle.net/11858/00-001M-0000-0028-6432-F
Abstract
Functional nucleic acids, such as aptamers and allosteric ribozymes, can sense their ligands specifically, thereby undergoing structural alterations that can be converted into a detectable signal. The direct coupling of molecular recognition to signal generation enables the production of versatile reporters that can be applied as molecular probes for various purposes, including high-throughput screening. Here we describe an unprecedented type of a nucleic acid-based sensor system and show that it is amenable to high-throughput screening (HTS) applications. The approach detects the displacement of an aptamer from its bound protein partner by means of luminescent oxygen channeling. In a proof-of-principle study we demonstrate that the format is feasible for efficient identification of small drug-like molecules that bind to a protein target, in this case to the Sec7 domain of cytohesin. We extended the approach to a new cytohesin-specific single chain DNA aptamer, C10.41, which exhibits a similar binding behavior to cytohesins but has the advantage of being more stable and easier to synthesize and to modify than the RNA-aptamer M69. The results obtained with both aptamers indicate the general suitability of the aptamer-displacement assay based on luminescent oxygen channelling (ADLOC) for HTS. We also analyzed the potential for false positive hits and identified from a library of 18,000 drug-like small molecules two compounds as strong singlet-oxygen quenchers. With full automation and the use of commercially available plate readers, we estimate that the ADLOC-based assay described here could be used to screen at least 100,000 compounds per day.