Phage-inspired coated nucleic acids for aptamers and their polymerases – PICNA-APT

The aim of this project is to expand the chemical universe of biopolymers used for aptamer synthesis, selection and amplification, by using a much larger library of elementary bricks modified on the nucleobase. The resulting aptamers will have an increased stability and specificity.
The choice of base modifications will be largely inspired but not limited to what is observed in some phages that use them to escape their host endonucleases.
Indeed, we will derivatize DNA or RNA polymers with amino acids and glycans at the nucleobase level such that Watson-Crick base-pairing will be kept untouched, thereby developing entirely new informational biopolymers, whose information can be amplified by variants of natural DNA or RNA polymerases that we will develop.
In practice, we will synthesize a whole new family of base-substituted nucleotide triphosphates, mainly by grafting amino-acids, glycans or combination thereof, at position C5 for pyrimidines or position C7 for (7-deaza) purines, while adopting a 2'OMe chemical group on the ribose for nuclease resistance.
We will then test a library of DNA and RNA polymerases that is available in the coordinator's lab for their capacity of incorporating with high fidelity these new nucleotides. This set includes a new family of simplified RNA polymerases that was recently fully characterized by one of us, both structurally and functionally, which is amenable to protein engineering and highly tolerant to base and sugar modifications in the incoming nucleotide substrate. Structural and functional information will be used to propose positions to mutate and generate libraries of mutants to make the initial hits more efficient. Directed evolution will be used to find replicative enzymes with high enzymatic capabilities and fidelity for hyper-modified aptamers amplification.
Finally, we will use these new tools to explore new chemical diversity in aptamers and amplify them in a specific application with potential use in human health.