Date of Award

1-1-2015

Document Type

Dissertation

Degree Name

Ph.D.

Organizational Unit

Chemistry and Biochemistry

First Advisor

Michelle K. Knowles, Ph.D.

Second Advisor

Dinah Loerke

Third Advisor

Martin Margittai

Fourth Advisor

Alex Huffman

Fifth Advisor

Joseph Angleson

Keywords

Membrane curvature, Protein behavior, Plasma membrane

Abstract

Variations in the physical deformation of the plasma membrane play a significant role in the sorting and behavior of the proteins that occupy it. Determining the interplay between membrane curvature and protein behavior required the development and thorough characterization of a model plasma membrane with well defined and localized regions of curvature. This model system consists of a fluid lipid bilayer that is supported by a dye-loaded polystyrene nanoparticle patterned glass substrate. As the physical deformation of the supported lipid bilayer is essential to our understanding of the behavior of the protein occupying the bilayer, extensive characterization of the structure of the model plasma membrane was conducted. Neither the regions of curvature in the vicinity of the polystyrene nanoparticles or the interaction between a lipid bilayer and small patches of curved polystyrene are well understood, so the results of experiments to determine these properties are described. To do so, individual fluorescently labeled proteins and lipids are tracked on this model system and in live cells. New methods for analyzing the resulting tracks and ensemble data are presented and discussed. To validate the model system and analytical methods, fluorescence microscopy was used to image a peripheral membrane protein, cholera toxin subunit B (CTB). These results are compared to results obtained from membrane components that were not expected to show an preference for membrane curvature: an individual fluorescently-labeled lipid, lissamine rhodamine B DHPE, and another protein, streptavidin associated with biotin-labeled DHPE. The observed tendency for cholera toxin subunit B to avoid curved regions of curvature, as determined by new and established analytical methods, is presented and discussed.

Publication Statement

Copyright is held by the author. User is responsible for all copyright compliance.

Rights Holder

Philip P. Cheney

Provenance

Received from ProQuest

File Format

application/pdf

Language

en

File Size

235 p.

Discipline

Chemistry, Biophysics, Physical Chemistry

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