UC Berkeley

The synthetic modification of proteins has significantly contributed in the fields of chemical biology and material science. Choosing an appropriate chemical strategy that involves the controlled coupling of biomolecules in aqueous solution and in a well-defined manner is key. This work describes the use of suitable bioconjugation reactions for a variety of applications. Firstly, oxidative coupling of o-aminophenols and anilines was used for the detection of nitrotyrosine-containing proteins. In this work, we demonstrated that the chemoselectivity of the tandem nitro reduction/oxidative coupling sequence was sufficiently high to label nitrated proteins among thousands of others in complex mixtures. Second, we constructed protein-polymer conjugates using 2-pyridinecarboxaldehyde reaction for intracellular protein delivery. The polymers were endowed with 2-pyridinecarboxaldehyde (2PCA) groups, which modify proteins selectively at their N-terminal positions through imidazolidinone formation. The polymer-modified proteins were delivered into the cytosolic compartment of various cancer cells and the function of the proteins was retained during the process. Importantly, mechanistic studies have revealed that the internalization pathways of the constructs are likely to occur through a membrane fusion mechanism, thus at least partially circumventing the protein inactivation that occurs through endocytosis. Finally, we adapted 2PCA and oxidative coupling reactions to create a Förster resonance energy transfer construct of an intrinsically disordered protein (IDP) to monitor protein folding. A library of IDP with different lengths and densities of polyethylene glycol (PEG) through lysine modifications was constructed. We demonstrated that the size and density of the PEG chains had little to no effect on the conformation of IDPs.