Poster abstracts

Poster number 6 submitted by Papa Nii Asare-Okai

Exploring the mechanism of FID using ESI-MS

Papa Nii Asare-Okai (Chemistry Department, Wayne State University)

Abstract:
Fluorescent intercalator displacement (FID) is becoming an important tool for identifying new nucleic acid binding ligands. Its success is based on the fact that it can be fashioned into a versatile high-throughput assay that can be used for assessing the relative binding affinities of compounds to nucleic acids in a simple and efficient manner, and requiring no significant expertise. FID is becoming an increasingly attractive method because it is a tag-less approach; neither the RNA nor the small molecule under investigation has to be modified.
In this study, an FID method for screening RNA-binding ligands was established, using TO-PRO as the fluorescent intercalator. Electrospray ionization mass spectrometry (ESI-MS) was used to investigate its mechanism, and the assay was then successfully applied to screening a variety of RNA-binding ligands.
Our assay was able to differentiate ligands that bind to a variety of RNA constructs used in this study (A-site, TAR, H31 and H69) with moderate selectivity. Furthermore, the FID results were compared to those obtained using ESI-MS. Our results provide molecular evidence that correlates the reduction in fluorescence observed in the FID assay with the displacement of a dye molecule from RNA.

Keywords: Fluorescent intercalator displacement, Electrospray ionization mass spectrometry, RNA