Dirhodium Metallopeptides for Catalytic Protein Modification & Inhibition of Protein-Protein Interactions
Ball, Zachary T
Doctor of Philosophy
The fundamental goal of chemical biology is to develop tools for interrogation of the activity and function of biomolecules, ultimately leading to understanding of the inner workings of biological systems. This thesis discusses three contributions towards expanding the chemical biology toolbox: i) an enzyme-like specific protein modification method, ii) a chemical blotting protocol for analyzing the products of protein modification reactions, iii) potent and selective inhibitors of protein-protein interactions. Inspired by the exquisite control and selectivity seen in enzymatic transformations, dirhodium metallopeptides were developed that catalyze selective labeling of natural proteins in their native environment. These tailored metallopeptides are highly specific for the targeted proteins, allowing quantitative modification of a target protein at micromolar concentrations even in the presence of a large number of potentially reactive biomolecules. The utility of the method for biophysical studies was demonstrated by labeling Yes protein kinase directly in cell lysate. In order to analyze products of in-lysate modification reactions, a chemical blotting protocol was established, and luminogenic probes were designed for use in chemical blotting and cell imaging applications. Design of inhibitors for protein-protein interactions is a formidable challenge due to the structural and chemical properties of protein interfaces involved in these interactions. Exploiting the potential of the dirhodium core to participate in cooperative binding, inhibitors for SH3 protein domains were developed by attaching dirhodium at different positions on a parent peptide ligand with modest SH3-affinity. The rationally designed library of dirhodium metalloinhibitors yielded the tightest known inhibitor for Lyn SH3; Lyn, a tyrosine kinase, represents an important target to further our understanding of a variety of cellular dysfunctions. Taken together, these contributions represent a set of powerful tools in chemical biology and can serve as a foundation for continued advancement of the field.
Protein modification; metalloinhibitor design; SH3 protein domains