Applying protein specific biosensor tools towards the understanding of Alzheimer's disease
Abstract
Alzheimer’s Disease (AD) is a progressively debilitating disease that involves the inability to coordinate movement, loss of memory, and a decrease in cognition. It is the most common form of dementia that affects an astoundingly large amount of the elderly population today, and the numbers of affected people are increasing as the life expectancy of our population grows. Recent evidence has indicated that the protein cAbl may play a role in neuronal apoptosis in AD cell and animal models. In this thesis, the novel approach of using a cAbl protein-specific biosensor (cAbl biosensor) to observe the hypothesized increase in cAbl activation in AD models is proposed. First, the phosphorylation of the cAbl biosensor peptide in a cAbl over-expressed HEK cell line was demonstrated. The time dependency of peptide incubation and phosphorylation of the peptide varied, but it could be inhibited by the cAbl inhibitor, Gleevec. These findings demonstrate that the cAbl biosensor could be phosphorylated in a cAbl-dependent manner although further optimization is required. Next, the ability of a cAbl biosensor related peptide (SH3-TAT) to cross the cell membrane and localize inside N2A cells and PCTX cells, two cell lines that are commonly used in neuroscience research, was observed. These data demonstrated that the SH3-TAT could translocate into each of these cell lines. In the N2A cell line the localization of the SH3-TAT appeared to occur in small vesicles within the cytosol whereas SH3-TAT in PCTX neuronal cells could be found more evenly dispersed throughout the cell. Finally, phosphorylation of the cAbl biosensor in N2A and PCTX cells was successfully demonstrated. These findings, although preliminary, offer a step toward developing novel assays that will be an easy to use, quick, cost efficient, and accurate way to study the biological mechanisms of AD as well as diagnose and monitor the disease.
Degree
M.S.
Advisors
Parker, Purdue University.
Subject Area
Pharmacy sciences
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