College Of William And Mary

NIH Research Projects · FY 2025 · 2016-04

Project Abstract The ability to prepare well-defined protein/small-molecule/solid-support bioconjugates possesses significant advantages in the treatment and diagnosis of a variety of diseases. Moreover, the generation of multivalent conjugates can even further increase the utility and application of these therapeutics by bringing together multiple partners in a single molecule. However, several issues associated with their preparation preclude widespread application of these biomacromolecules. This proposal aims to address these issues via the use of noncanonical amino acid (ncAA) technologies to rapidly synthesize highly active and well-defined conjugates. Specifically, a variety of noncanonical amino acids will be synthetically prepared and genetically encoded into proteins to serve as functionalization handles. First, novel bioconjugation reactions will be developed and optimized under physiological conditions, including the Glaser Hay coupling between terminal alkynes and additional reactions based on the diyne moiety afforded by the Glaser Hay. Finally, the optimized reaction cascades can be combined to produce multivalent bioconjugates that provide numerous therapeutic properties inaccessible to standard bivalent conjugates. This includes the linking of an antibody targeting agent to a small molecule drug and a fluorophore to be able to track drug delivery; however, there are numerous other applications of multivalent conjugates that can also be envisioned. Ultimately, the methodologies will be transitioned to biologically relevant systems including ubiquitin binding proteins, esterases, antibodies, MEMO and Cas9/dCas9. The tools developed within the proposed research will not only significantly advance the fields of therapeutics and diagnostics, but also be extremely useful in the training of undergraduate researchers towards their future careers within the scientific arena.