Open AccessDNA Aptamer–Target Binding Motif Revealed Using a Fluorescent Guanine Probe: Implications for Food Toxin Detection
Author(s) -
Kaila L. Fadock,
Richard A. Manderville
Publication year - 2017
Publication title -
acs omega
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.779
H-Index - 40
ISSN - 2470-1343
DOI - 10.1021/acsomega.7b00782
Subject(s) - aptamer , oligonucleotide , dna , g quadruplex , guanine , antiparallel (mathematics) , fluorescence , chemistry , biochemistry , stacking , biology , biophysics , microbiology and biotechnology , gene , nucleotide , physics , quantum mechanics , magnetic field , organic chemistry
DNA aptamers are single-stranded oligonucleotides that are generated by an in vitro selection method to bind targets with high affinity and specificity. Understanding molecular recognition by DNA aptamers is of fundamental importance in the development of biosensor applications. The small molecule ochratoxin A (OTA) is a fungal-derived food toxin, and OTA DNA aptamers have been established for the development of rapid detection platforms required for food safety. One such OTA aptamer (OTAA) is a guanine-rich DNA oligonucleotide that folds into an antiparallel G-quadruplex (GQ) upon OTA binding, although structural details of the GQ fold and its interaction with OTA are currently unknown. In the present study, the fluorescent nucleobase analogue, 8-thienyl-2'-deoxyguanosine (ThdG), was inserted into various G sites of OTAA to determine the probe impact on GQ folding and OTA binding affinity. Our results suggest that OTAA contains three lateral (l) loops connecting two stacked G-tetrads with an anticlockwise loop progression to afford a -(lll) GQ topology. The phenolic ring system of OTA undergoes π-stacking interactions with the G-tetrads of OTAA. Our results also demonstrate aptamer sites that can be modified with ThdG to afford a fluorescent light-up signal upon OTA binding.