Advisory Committee Chair
Om P Srivastava
Advisory Committee Members
Kent T Keyser
Alecia K Gross
Dennis J Pillion
David R Whikehart
Document Type
Dissertation
Date of Award
2011
Degree Name by School
Doctor of Philosophy (PhD) School of Optometry
Abstract
Cataract is an opacification of the lens causing reduction in vision over time. It is one of the leading causes of vision loss among adults age 55 and older. The lens is a transparent structure that focuses incoming light onto the retina for visual perception. It has a high protein content, attributed mainly to water-soluble proteins known as crystallins. With age, crystallins undergo post-translational modifications (PTMs) that alter their structural and/or functional properties and, in turn, lead to cataract formation. The purpose of this research was to take a top-down approach in understanding changes in lens proteins with age, starting with determining the overall changes in crystallins occurring in cortical and nuclear regions of the lens, followed by determining effects of PTMs (deamidation and truncation) on structural/functional properties of αB-crystallin, and concluding with studies of molecular properties of individual N-terminal domain (NTD), core domain (CD), and C-terminal extension (CTE) of αA- and αB-crystallins. In the first study, a combination of laser capture microdissection (LCM), 2-D Difference Gel Electrophoresis (2-D DIGE), and Matrix-Assisted Laser Desorption/Ionization Time-of-Flight (MALDI-TOF) and Electrospray Ionization Quadripole Linear Ion-Trap Liquid Chromatography (ESI-QTRAP LC-MS/MS) mass spectrometry were used to map and identify the in situ protein profile and PTMs of crystallins in the cortical and nuclear regions of aging human lenses. Crystallin fragmentation resulting from truncation began in the cortical region and continued into the nuclear region, whereas aggregation was predominantly observed in the nuclear region. One way to study the effects of PTMs is to create mutants that mimic in vivo modifications (i.e., truncation and deamidation) and examine their altered structural and functional properties. In the second study, the structural and functional properties of deamidated, N- or C-terminally deleted, and deamidated plus N- or C-terminally deleted mutants of αB-crystallin were analyzed. Relative to WT αB, the N146 deamidation and N-terminal deletion caused greater structural and functional changes in the crystallin. In the third study the molecular properties of NTD, CD, and CTE constructs of αA- and αB-crystallins were analyzed. The individual α-crystallin regions exhibited varied biophysical properties and each region alone retained some level of chaperone function.
Recommended Citation
Asomugha, Chinwe, "Post-translational Modifications, Structure and Function of Human Lens Crystallins" (2011). All ETDs from UAB. 1046.
https://digitalcommons.library.uab.edu/etd-collection/1046
