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THE FOLDING, MISFOLDING AND REFOLDING OF MEMBRANE PROTEINS AND THE DESIGN OF A PHOSPHATIDYLSERINE-SPECIFIC MEMBRANE SENSOR
by
Yujin E. Kim
A Dissertation Presented to the
FACULTY OF THE USC GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the Requirements for the Degree
DOCTOR OF PHILOSOPHY
(BIOCHEMISTRY AND MOLECULAR BIOLOGY)
December 2009
Copyright 2009 Yujin E. Kim
Object Description
| Title | The folding, misfolding and refolding of membrane proteins and the design of a phosphatidylserine-specific membrane sensor |
| Author | Kim, Yujin E. |
| Author email | yujinkim@usc.edu; yujinkim820@hotmail.com |
| Degree | Doctor of Philosophy |
| Document type | Dissertation |
| Degree program | Biochemistry & Molecular Biology |
| School | Keck School of Medicine |
| Date defended/completed | 2009-09-25 |
| Date submitted | 2009 |
| Restricted until | Unrestricted |
| Date published | 2009-10-29 |
| Advisor (committee chair) | Langen, Ralf |
| Advisor (committee member) |
Chen, Jeannie Chow, Robert Haworth, Ian Markland, Francis |
| Abstract | The interaction between proteins and membranes underlies many important biological processes. Proteins influence the structure and fluidity of membranes and likewise, membranes promote structural reorganization in proteins, mutually modifying their functions. In addition, protein-membrane interactions can lead to protein misfolding and the pathogenesis of disease. Despite the importance of these dynamic protein-membrane interactions, less is known about the structure of membrane-bound proteins in comparison to proteins in solution. As an alternative to conventional methods such as X-ray crystallography and NMR, we used electron paramagnetic resonance (EPR) spectroscopy in combination with site-directed spin labeling (SDSL) as the principle technique to study how different environmental conditions can promote the structural transitions that occur in two different membrane-binding proteins, annexin B12 (anxB12, anxXII) and α-synuclein (α-syn). AnxB12 is a member of a family of structurally conserved proteins that are abundantly expressed in most eukaryotic cells. The various proposed functions include vesicle trafficking, membrane fusion, ion channel formation, and cell signaling, indicating that membrane interactions are important for all annexin functions. In order to gain a better understanding of the mechanism of membrane binding and annexin function, the structures of two different membrane-bound states of AnxB12 were studied. In addition, using the data obtained from these studies, we engineered a polarity sensitive annexin-based biosensor applicable to the real-time detection of apoptotic membrane changes in cultured cells and in vivo. The second protein of interest, α-syn, is a natively unfolded cytosolic protein which is observed to bind preferentially to small vesicles (of similar size to synaptic vesicles) and is localized to the presynaptic nerve terminals.; The accumulation of misfolded cytoplasmic aggregates of α-syn is central to the pathology Parkinson’s disease (PD). Recent studies have suggested that the soluble oligomers of α-syn are a cause of cell toxicity and disease pathogenesis. One of the proposed mechanisms by which these oligomers are toxic is by membrane interaction and disruption. In order to gain a better understanding of how misfolded α-syn oligomers are assembled and their role in the disease relevant protein misfolding pathway, we studied the structures of different oligomeric states of α-syn by EPR techniques. |
| Keyword | EPR; α-synuclein; oligomers; protein misfolding; membrane protein structure; membrane binding |
| Language | English |
| Part of collection | University of Southern California dissertations and theses |
| Publisher (of the original version) | University of Southern California |
| Place of publication (of the original version) | Los Angeles, California |
| Publisher (of the digital version) | University of Southern California. Libraries |
| Provenance | Electronically uploaded by the author |
| Type | texts |
| Legacy record ID | usctheses-m2700 |
| Rights | Kim, Yujin E. |
| Repository name | Libraries, University of Southern California |
| Repository address | Los Angeles, California |
| Repository email | http://www.usc.edu/isd/libraries/services/ask_a_librarian/email/ |
| Filename | etd-Kim-3342 |
| Archival file | uscthesesreloadpub_Volume32/etd-Kim-3342.pdf |
Description
| Title | Page 1 |
| Full text | THE FOLDING, MISFOLDING AND REFOLDING OF MEMBRANE PROTEINS AND THE DESIGN OF A PHOSPHATIDYLSERINE-SPECIFIC MEMBRANE SENSOR by Yujin E. Kim A Dissertation Presented to the FACULTY OF THE USC GRADUATE SCHOOL UNIVERSITY OF SOUTHERN CALIFORNIA In Partial Fulfillment of the Requirements for the Degree DOCTOR OF PHILOSOPHY (BIOCHEMISTRY AND MOLECULAR BIOLOGY) December 2009 Copyright 2009 Yujin E. Kim |
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