1 research outputs found
Detection of Ligand-Induced Conformational Changes in Oligonucleotides by Second-Harmonic Generation at a Supported Lipid Bilayer Interface
There is a high demand for characterizing
oligonucleotide structural
changes associated with binding interactions as well as identifying
novel binders that modulate their structure and function. In this
study, second-harmonic generation (SHG) was used to study RNA and
DNA oligonucleotide conformational changes associated with ligand
binding. For this purpose, we developed an avidin-based biotin capture
surface based on a supported lipid bilayer membrane. The technique
was applied to two well-characterized aptamers, both of which undergo
conformational changes upon binding either a protein or a small molecule
ligand. In both cases, SHG was able to resolve conformational changes
in these oligonucleotides sensitively and specifically, in solution
and in real time, using nanogram amounts of material. In addition,
we developed a competition assay for the oligonucleotides between
the specific ligands and known, nonspecific binders, and we demonstrated
that intercalators and minor groove binders affect the conformation
of the DNA and RNA oligonucleotides in different ways upon binding
and subsequently block specific ligand binding in all cases. Our work
demonstrates the broad potential of SHG for studying oligonucleotides
and their conformational changes upon interaction with ligands. As
SHG offers a powerful, high-throughput screening approach, our results
here also open an important new avenue for identifying novel chemical
probes or sequence-targeted drugs that disrupt or modulate DNA or
RNA structure and function