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DEVELOPMENT OF FLUORESCENT CHOLESTEROL PROBES AND THEIR APPLICATION FOR CHOLESTEROL TRAFFICKING STUDIES
| Title: | DEVELOPMENT OF FLUORESCENT CHOLESTEROL PROBES AND THEIR APPLICATION FOR CHOLESTEROL TRAFFICKING STUDIES. |
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| Name(s): |
Rubio, Vicente Orlando , author Stawikowski, Maciej J., Thesis advisor Florida Atlantic University, Degree grantor Department of Chemistry and Biochemistry Charles E. Schmidt College of Science |
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| Type of Resource: | text | |
| Genre: | Electronic Thesis Or Dissertation | |
| Date Created: | 2024 | |
| Date Issued: | 2024 | |
| Publisher: | Florida Atlantic University | |
| Place of Publication: | Boca Raton, Fla. | |
| Physical Form: | application/pdf | |
| Extent: | 273 p. | |
| Language(s): | English | |
| Abstract/Description: | Cholesterol is a pivotal component of mammalian cell membranes and homeostasis. Due to its high concentration and heterogenous distribution in the brain, cholesterol is tightly regulated and its dyshomeostasis is frequently implicated in several neurodegenerative diseases. This dissertation reports the design, synthesis, and application of ten cholesterol naphthalimide probes (CND) to study cholesterol trafficking in live cells. The CND series was rationally designed by incorporating several of the structural features of endogenous cholesterol onto the naphthalimide (ND) scaffold conjugated via an ester bond. The modularity of the ND scaffold enabled all analogs to have the aliphatic tail of cholesterol, which is lacking in the most ubiquitously utilized probe, BODIPY-Cholesterol, a.k.a. Top-Fluor® Cholesterol (TFC). The CNDs were demonstrated to be optimal probes for cholesterol due to the ability to fluorescence exclusively in hydrophobic/membrane environments and exhibit low fluorescence in hydrophilic/aqueous environments. By incorporating a protonatable piperazine group on the C4 position of the ND scaffold, CND2 – CND4 possessed pH sensing capabilities, which were demonstrated to monitor intracellular vesicle turnover in neurons. The potential of the CNDs to bind to the lysosomal sterol transport protein NPC2 was investigated by molecular docking and molecular dynamics (MD) simulations. The docking pose with the CND’s aliphatic tail positioned inside the hydrophobic binding pocket was essential for mimicking endogenous cholesterol’s interactions and stabilizing the NPC2-ligand complex. Fluorescence confocal microscopy demonstrated a structure-dependent and cell-dependent intracellular distribution of the CND series in live cells. | |
| Identifier: | FA00014430 (IID) | |
| Degree granted: | Dissertation (PhD)--Florida Atlantic University, 2024. | |
| Collection: | FAU Electronic Theses and Dissertations Collection | |
| Note(s): | Includes bibliography. | |
| Subject(s): |
Cholesterol Fluorescent probes Naphthalimides Neurodegenerative Diseases |
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| Persistent Link to This Record: | http://purl.flvc.org/fau/fd/FA00014430 | |
| Use and Reproduction: | Copyright © is held by the author with permission granted to Florida Atlantic University to digitize, archive and distribute this item for non-profit research and educational purposes. Any reuse of this item in excess of fair use or other copyright exemptions requires permission of the copyright holder. | |
| Use and Reproduction: | http://rightsstatements.org/vocab/InC/1.0/ | |
| Host Institution: | FAU |
