Boatz, Jennifer C.
(2019)
Aggregate Polymorphism in Protein Deposition Diseases: Investigations by Magic Angle Spinning Solid State NMR and Transmission Electron Microscopy.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
This is the latest version of this item.
Abstract
The deposition of normally soluble protein can occur in any organ in the human body and is associated with tissue dysfunction, cell death, and the progression of disease. Protein aggregation is concomitant with blindness as an outcome of cataract, life-threatening organ failure as a consequence of amyloidosis, and pronounced degeneration of the brain.
The mutation responsible for Huntington’s disease causes an expansion of the polyglutamine domain of huntingtin exon 1 that directly promotes misfolding and refolding of huntingtin and huntingtin N-terminal fragments into amyloid-like fibrils in the basal striatum and cortex of the brain. Several fibril polymorphs have been identified, however the relationship between neurotoxicity and amyloid polymorphism is poorly understood. The P23T mutant of gamma-D-crystallin is associated with cataract formation in the eyes of very young children. Crystallins have been shown to form amyloid-like, native-like, as well as amorphous looking aggregates in vitro, accordingly it is unclear which class of aggregates P23T gamma-D-crystallin is most likely to form in cataract. Apolipoprotein A-I is a known anti-atherosclerotic factor and oxidation at methionine residues enhances its function. However, this oxidation also induces aggregation in vascular amyloidosis, which is interlinked with atherosclerosis progression. It is unclear whether apolipoprotein A-I aggregates misfold into amyloid-like fibrils as is usually the case in amyloidosis.
Magic angle spinning solid state NMR (MAS ssNMR) is ideally suited to provide atomic resolution information on the structure and dynamics of insoluble, non-crystalline protein aggregates. Transmission electron microscopy (TEM) allows for the visualization of morphological features of aggregates that cannot be observed by optical microscopy and can be used to identify polymorphs and aid in distinguishing between different classes of aggregates. In this dissertation, I use both MAS ssNMR and TEM in addition to other biophysical and structural techniques to investigate the differences in structure and dynamics between polymorphs of huntingtin exon 1, P23T gamma-D-crystallin, and apolipoprotein A-I. Enabled by my experiments, I narrow down the potential molecular mechanisms involved in these three distinct types of protein deposition diseases. I show that depending on the milieu, proteins have the potential for varied amyloidogenic and non-amyloidogenic self-assembly.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
3 January 2019 |
| Date Type: |
Publication |
| Defense Date: |
23 October 2018 |
| Approval Date: |
3 January 2019 |
| Submission Date: |
18 December 2018 |
| Access Restriction: |
1 year -- Restrict access to University of Pittsburgh for a period of 1 year. |
| Number of Pages: |
225 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
School of Medicine > Molecular Biophysics and Structural Biology |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
amyloid, huntingtin, cataract, apolipoprotein, aggregation, crystallin, Huntington's Disease |
| Date Deposited: |
03 Jan 2019 20:26 |
| Last Modified: |
03 Jan 2020 06:15 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/35802 |
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Aggregate Polymorphism in Protein Deposition Diseases: Investigations by Magic Angle Spinning Solid State NMR and Transmission Electron Microscopy. (deposited 03 Jan 2019 20:26)
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