A NEW TEMPLATED BIO-ORTHOGONAL REACTION

The desire to explore biomolecules and biological processes within their natural contexts has driven the need to develop of reactions that are compatible with such environments. This pursuit presents a substantial challenge owing to the complexity of cellular systems. Reactions capable of proceeding within a biological system without disrupting native biological processes are termed “bioorthogonal reactions”. Components involved must exhibit rapid and selective reactivity in water, at phisiological pH and temperature, in contexts of high molecular crowding. Despite the remarkable advancements in bioorthogonal chemistries, very few reactions have proven to be efficient thus far at cellular level four ligation. Persistent challenges remain within current methodologies, particularly concerning issues related to interference amidst the numerous functionalities present in vivo, and the requirement to remain non-toxic to the biological system. Our aim is to improve a recently discovered bio-orthogonal templated reaction which requires “proximiy” as sole trigger and expand his reactivity, ultimately extending its applicability to cellular studies. The chosen reaction involves the ligation between 2,5-dioxopentanyl (DOP) moiety and an alpha nucleophile, resulting in the formation of piridazinium or pyrrole linkage (Fig.1). 1,2 This reaction, which has been successfully tested in cell lysate, has demonstrated its ability to proceed exclusively under the guidance of a template, such as (peptide) nucleic acids or coiled-coil systems, with high specificity and without off-target reactivity. Hence, we will present preliminar results regarding the synthesis of various alpha nucleophiles building blocks and ligation experiments performed on peptide nucleic acids (PNAs) models.