Knud Jørgen Jensen, Professor, University of Copenhagen explained that his research is focused on installing new properties on oligonucleotides and using oligonucleotides to construct molecules that behave like proteins. He suggested that advancing single molecule studies and incorporating novel fluorescence techniques are key to mimicking protein behaviour.
Jensen aimed to create folded stable structures resembling proteins by combining orthogonal self-assembly principles such as oligonucleotide self-assembly and coiled coil peptide formations. By using strain-promoted alkyne-azide coupling, the team successfully linked peptides to oligonucleotides. This method precisely created constructs that replicated protein stability and folding behaviour. Jensen said: “This was a close collaboration with Jesper Wengel, the inventor of LNA. For the peptide moiety, we relied on what's called a coiled coil.” Leveraging coiled coil peptides allows researchers to organise ligands or oligos in multivalent arrangements.
The gel studies confirmed the successful assembly of peptide-oligonucleotide conjugates (POCs). The melting studies showed that the interactions between oligonucleotides and peptides reinforced each other’s stability, resulting in high melting points.
Further analysis utilized small-angle X-ray scattering (SAXS) and molecular dynamics simulations to study the 3D structures of the conjugates in solution. Jensen explained: “So SAXS by itself does not give atomic resolution, but you can get an idea of the internal structure by using MD simulations.” These methods revealed tightly folded structures with notable stability, even at termini and linkage regions. Jensen explained that a kink in the construct was observed at lower concentrations, while higher concentrations led to dimerization due to hydrophobic and hydrogen bonding interactions in the peptide segment.
Jensen then discussed the incorporation of fluorescent DNA-silver nanoclusters into the peptide-oligonucleotide system. These clusters, stabilised by DNA, demonstrated tunable fluorescent properties and were successfully conjugated to peptides and proteins such as insulin. Spectroscopic analysis confirmed that the optical properties of the silver clusters were preserved after conjugation. Then imaging studies demonstrated their ability to stain cells with overexpressed insulin receptors selectively.
This talk outlines the dynamic nature of peptide-oligonucleotide conjugates, they can be applied to a range of processes like drug delivery, artificial protein design, and the study of RNA conjugate endosomal escape. This research adopts a creative approach to combining the best of peptide and oligonucleotide chemistry.