Studies in pharmaceutical biotechnology : protein-protein interactions and beyond

Pharmaceutical biotechnology has been emerging as a defined, increasingly important area of science dedicated to the discovery and delivery of drugs and therapies for the treatment of various human diseases. In contrast to the advancement in pharmaceutical biotechnology, current drug discovery efforts are facing unprecedented challenges. Difficulties in identifying novel drug targets and developing effective and safe drugs are closely related to the complexity of the network of interacting human proteins. Protein-protein interactions mediate virtually all cellular processes. Therefore both identification and understanding of protein-protein interactions are essential to the process of deciphering disease mechanisms and developing treatments. Unfortunately, our current knowledge and understanding of the human interactome is largely incomplete. Most of the unknown protein-protein interactions are expected to be weak and/or transient, hence are not easily identified. These unknown or uncharacterized interactions could affect the efficacy and toxicity of drug candidates, contributing to the high rate of failure. In an attempt to facilitate the ongoing efforts in drug discovery, we describe herein a series of novel methods and their applications addressing the broad topic of protein-protein interactions. We have developed a highly efficient site-specific protein cross-linking technology mediated by the genetically incorporated non-canonical amino acid L-DOPA to facilitate the identification and characterization of weak protein-protein interactions. We also established a protocol to incorporate L-DOPA into proteins in mammalian cells to enable in vivo site-specific protein cross-kinking. We then applied the DOPA-mediated cross-linking methodology to design a protein probe which can potentially serve as a diagnostic tool or a modulator of protein-protein interactions in vivo. To deliver such engineered proteins or other bioanalytical reagents into single live cells, we established a laser-assisted cellular nano-surgery protocol which would enable detailed observations of cell-to-cell variability and communication. Finally we investigated a possible experimental scheme to genetically evolve a fluorescent peptide, which has tremendous potential as a tool in cellular imaging and dynamic observation of protein-protein interactions in vivo. We aim to contribute to the discovery and development of new drugs and eventually to the overall health of our society by adding the technology above to the array of currently available bioanalytical tools.