| dc.identifier.uri | http://hdl.handle.net/11401/77823 | |
| dc.description.sponsorship | This work is sponsored by the Stony Brook University Graduate School in compliance with the requirements for completion of degree. | en_US |
| dc.format | Monograph | |
| dc.format.medium | Electronic Resource | en_US |
| dc.language.iso | en_US | |
| dc.publisher | The Graduate School, Stony Brook University: Stony Brook, NY. | |
| dc.type | Dissertation | |
| dcterms.abstract | Lipid rafts are tightly packed saturated lipid and cholesterol-rich liquid-ordered (Lo) membrane domains that are believed to co-exist with loosely packed disordered domains composed mostly of unsaturated lipids. Rafts control numerous protein-protein and protein-lipid interactions at the cell surface. In this study, the sterol-dependent, pore-forming cytolysin perfringolysin O (PFO) was used to investigate how transmembrane (TM) protein association with lipid rafts is regulated and how lipid asymmetry affects transmembrane protein structure. First, to test the hypothesis that mismatch between protein TM segment length and bilayer width controls TM protein affinity for rafts, the raft affinity of PFO mutants with different length of TM segments was measured by Förster Resonance Energy Transfer (FRET) and confocal microscopy. The results indicate that the affinity of PFO for ordered lipid domains is increased by matching between TM segment lengths and bilayer width in the ordered domains. Since this effect of match is seen for PFO, which has multiple TM segments and not for single TM segment proteins, we propose that hydrophobic mismatch and multimeric state can control TM protein association with lipid rafts. Another hypothesis is that tight binding of TM proteins to lipids that have a strong affinity for ordered domains promotes raft-protein interaction. To test this hypothesis, PFO raft affinity when bound to cholesterol was compared to that when PFO is bound to coprostanol, a sterol which has a structure unfavorable for raft association. The results from the FRET assay indicate that PFO binding with ordered domains is promoted upon the association of the PFO-bound sterol with ordered domains. Finally, the effects of lipid asymmetry on PFO conformational behavior were investigated. Using hydroxylpropyl-alpha-cyclodextrin based lipid exchange, we developed a novel method to prepare large unilamellar model membrane vesicles that closely resemble mammalian plasma membranes in terms of their lipid asymmetry, i.e. which have a different lipid composition in their inner and outer leaflets. PFO conformational behavior in plasma membrane-like asymmetric vesicles was found to be distinct both from that in symmetric vesicles with the same lipid composition, and from that in vesicles containing either only inner leaflet lipids or only outer leaflet lipids. These studies show that lipid asymmetry can strongly influence the behavior of a membrane-inserted protein. | |
| dcterms.available | 2017-09-26T17:00:35Z | |
| dcterms.contributor | Deutsch, Dale | en_US |
| dcterms.contributor | London, Erwin | en_US |
| dcterms.contributor | Scarlata, Suzanne | en_US |
| dcterms.contributor | Brown, Deborah | en_US |
| dcterms.contributor | Thanassi, David. | en_US |
| dcterms.creator | Lin, Qingqing | |
| dcterms.dateAccepted | 2017-09-26T17:00:35Z | |
| dcterms.dateSubmitted | 2017-09-26T17:00:35Z | |
| dcterms.description | Department of Biochemistry and Structural Biology. | en_US |
| dcterms.extent | 134 pg. | en_US |
| dcterms.format | Monograph | |
| dcterms.format | Application/PDF | en_US |
| dcterms.identifier | http://hdl.handle.net/11401/77823 | |
| dcterms.identifier | Lin_grad.sunysb_0771E_11482.pdf | en_US |
| dcterms.issued | 2013-05-01 | |
| dcterms.language | en_US | |
| dcterms.provenance | Submitted by Jason Torre (fjason.torre@stonybrook.edu) on 2017-09-26T17:00:35Z
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Lin_grad.sunysb_0771E_11482.pdf: 4398802 bytes, checksum: 4aae9314c02770d368f14492213afebe (MD5) | en |
| dcterms.provenance | Made available in DSpace on 2017-09-26T17:00:35Z (GMT). No. of bitstreams: 1
Lin_grad.sunysb_0771E_11482.pdf: 4398802 bytes, checksum: 4aae9314c02770d368f14492213afebe (MD5)
Previous issue date: 2013-05-01 | en |
| dcterms.publisher | The Graduate School, Stony Brook University: Stony Brook, NY. | |
| dcterms.subject | Biochemistry | |
| dcterms.title | Transmembrane Protein Behavior in " Artificial Plasma Membranes" Reveals How Protein Binding to Lipid Rafts is Regulated and How Lipid Asymmetry Affects Protein Structure | |
| dcterms.type | Dissertation | |
