Triazolinedione Modification of Prenylated Peptides, Proteins, and DNA: Toward a Single-Site Multilabeling Approach for Biomolecules

In efforts to shed light on the complexity of biological processes, attaching several or different payloads onto a biomolecular target of interest has become an interesting tool within the field of bioconjugation. Herein, we report on the exploitation of prenylated (bio)-molecules in a 1,2,4-triazole-3,5-(4<i>H</i>)-dione-based labeling strategy to develop a two-step single-site multiple labeling methodology that allows the introduction of up to three identical or different property-enhancing moieties. The methodology was first demonstrated on the small molecule targets farnesol and S-geranyl-2-thiouridine to be then applied to multi-(functional) labeling of amino acids and peptides. We further demonstrate the usefulness of this approach to achieve branched lipidation of peptides, a strategy recently receiving more attention for enhanced cellular membrane localization and vaccine development. In addition, proof-of-concept experiments were performed involving single-site multiple labeling of large biomolecules such as farnesylated proteins and geranylated DNA.