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Type of Document	Dissertation
Author	Sakamoto, Kathleen Miho
URN	etd-12292003-134700
Persistent URL	http://resolver.caltech.edu/CaltechETD:etd-12292003-134700
Title	Targeting proteins for ubiquitination and degradation in the treatment of human disease
Degree	PhD
Option	Biology
Advisory Committee	Advisor Name Title Raymond J. Deshaies Committee Chair David Baltimore Committee Member Elliot Meyerowitz Committee Member Paul Sternberg Committee Member William Dunphy Committee Member
Keywords	ubiquitin ligase ubiquitination degradation human disease
Date of Defense	2003-12-18
Availability	unrestricted
Abstract Protein degradation is one of the tactics employed by the cell for irreversibly inactivating proteins. In eukaryotes, ATP-dependent protein degradation in the cytoplasm and nucleus is carried out by the 26S proteasome. Most proteins are targeted to the 26S proteasome by covalent attachment of a multiubiquitin chain. A key component of the enzyme cascade that results in attachment of the multiubiquitin chain to the target or labile protein is the ubiquitin ligase that controls the specificity of the ubiquitination reaction. Defects in ubiquitin-dependent proteolysis have been shown to result in a variety of human diseases, including cancer, neurodegenerative diseases, and metabolic disorders. The SCF (Skp1-Cullin-F-box-Hrt1) complex is a heteromeric ubiquitin ligase that multiubiquitinates proteins important for signal transduction and cell cycle progression. A technology was developed known as Protac (Proteolysis Targeting Chimeric Molecule), that acts as a bridge, bringing together the SCF ubiquitin ligase with a protein target, resulting in its ubiquitination and degradation. The Protac contains a peptide moiety at one end that is recognized by SCF that is chemically linked to the binding partner or ligand of the target protein. The first demonstration of the efficacy of Protac technology was the successful recruitment, ubiquitination, and degradation of the protein Methionine Aminopeptidase-2 (MetAP-2) through a covalent interaction between MetAP-2 and Protac. Subsequently, we demonstrated that Protacs could effectively ubiquitinate and degrade cancer-promoting proteins (estrogen and androgen receptors) through non-covalent interactions in vitro and in cells. Finally, cell-permeable Protacs can also promote the degradation of proteins in cells. Biologically, this work signifies the amazing versatility and flexibility of the ubiquitin-proteasome system. Technologically, this work represents the development of a novel ?chemical genetics? approach to selectively target proteins for degradation. Practically, this work is ?Proof of Concept? that exploiting the cell?s natural proteolytic machinery is a potential avenue for the treatment of human disease.
Files	Filename       Size       Approximate Download Time (Hours:Minutes:Seconds)     28.8 Modem   56K Modem   ISDN (64 Kb)   ISDN (128 Kb)   Higher-speed Access    Appendix1.pdf 374.52 Kb 00:01:44 00:00:53 00:00:46 00:00:23 00:00:01   Appendix2.pdf 1.38 Mb 00:06:23 00:03:17 00:02:52 00:01:26 00:00:07   Appendix3.pdf 577.27 Kb 00:02:40 00:01:22 00:01:12 00:00:36 00:00:03   Appendix4.pdf 1.86 Mb 00:08:36 00:04:25 00:03:52 00:01:56 00:00:09   Appendix5.pdf 631.92 Kb 00:02:55 00:01:30 00:01:18 00:00:39 00:00:03   Thesis.pdf 6.30 Mb 00:29:10 00:15:00 00:13:07 00:06:33 00:00:33