Small-Molecule Aptamers
Many detection methods take advantage of a compound's conformational change. By isolating DNA aptamers that exhibit this change upon binding to a small molecule, we can develop simple sensors and high-throughput screens for biomarkers, illicit drugs and other targets of interest.
Aptamers are nucleic acid probes that are designed to selectively and strongly bind to a variety of targets, such as small molecules, proteins, and other biomolecules.1 Compared to antibodies, aptamers offer a variety of advantages, such as ease of production, ability to be used in non-physiological conditions, and ability to be generated for targets that do not generate an immunogenic response within the body.
Aptamers are generated from a process known as SELEX (Selective Evolution of Ligands by Exponential Enrichment). Selection of aptamers for small molecules is a challenging process; the miniscule difference in size between the free aptamer and most aptamer-small molecule complexes makes isolation of high affinity aptamers difficult.2
This project proposes generation of aptamers for small molecule targets, where the isolation mechanism is designed to isolate aptamers that undergo folding upon binding to the target of interest. Using size exclusion chromatography-based SELEX (SEC-SELEX, illustrated above), we will isolate aptamers that not only bind to the target, but undergo a conformational change upon binding. By isolating folding aptamers for small molecule targets, these aptamers will be highly adaptable to a variety of sensing platforms, both for rapid detection of single analytes in the field as well as high-throughput analysis in a laboratory setting.
Some targets of interest include 3-hydroxykynurenine, a molecule whose upregulation in urine has been tied to the development of several neurodegenerative diseases, and illict drug compounds such as amphetamines. Screening/sensing methods include fluorescence polarization, fluorescence quenching, fluorescence resonance energy transfer (FRET), and voltammetry.
1. Song, S. P.; Wang, L. H.; Li, J.; Zhao, J. L.; Fan, C. H., Aptamer-based biosensors. Trac-Trends in Analytical Chemistry 2008, 27 (2), 108-117.
2. McKeague, M.; DeRosa, M. C., Challenges and Opportunities for Small Molecule Aptamer Development. Journal of Nucleic Acids 2012, 2012, 20.