Computational Design and Experimental Characterization of Protein Oligomers

Citation

Huang, Po-Ssu (2004) Computational Design and Experimental Characterization of Protein Oligomers. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/3DZW-2R54. https://resolver.caltech.edu/CaltechETD:etd-06012004-214823

Abstract

Previous efforts in designing protein binding interfaces have focused on altering binding specificities. These methods fall short, however, when applied to the design of novel binding sites due to difficulties in accurately modeling protein backbones. The goal of this project is to create dimers from monomeric proteins. We developed a special docking algorithm that positions the member protein subunits to a plausible configuration with respect to each other using parameters determined from known complex structures. The docking procedure treats the proteins as rigid bodies and uses Fourier correlation theorem and fast Fourier transform to efficiently search for dimers with the highest interfacial surface complementarities. Using the docked structures as scaffolds for design and employing hydrophobic surface residues to drive dimer formation, we have demonstrated two successful designs, one heterodimer and one homodimer, using protein G and engrailed homeodomain respectively as the starting monomeric proteins. The designed dimers were characterized using circular dichroism, nuclear magnetic resonance, analytical ultracentrifugation, and X-ray crystallography methods. This is the first report of computationally designed de novo protein homodimers generated using a combination of protein docking and protein design tools. These results suggest that this strategy can be used to address the protein recognition problem, and is generally applicable to creating novel binding sites with compatible binding partners.

Thesis Files