Comparative Biochemical and Structural Analysis of the Alzheimer’s Disease Related Proteins: Amyloid Precursor Protein and Notch1.

Abstract

BIOCHEMISTRY

Comparative Biochemical and Structural Analysis of the Alzheimer’s Disease Related Proteins: Amyloid Precursor Protein and Notch 1.

Catherine Laura Deatherage

Dissertation under the direction of Professor Charles R. Sanders, Ph.D.

Alzheimer’s disease is a neurodegenerative disorder that affects millions of people and costs billions of dollars every year. As of mid-2016, despite more than twenty years of research, there is no cure for Alzheimer’s disease. This is due, in part, to toxicity stemming from the protein Notch that occurs when the gamma-secretase inhibitors, designed to act as Alzheimer’s therapeutics by reducing cleavage of the amyloid precursor protein, inadvertently inhibit cleavage of Notch. The work presented in this dissertation describes structural and biochemical studies to determine ways to specifically regulate gamma-secretase. Using a combination of biochemistry, biophysics, and NMR spectroscopy, I expanded the body of knowledge regarding proteins that were thought to be either Alzheimer’s drug targets or to be a confounding factor in drug development. It was found that the gamma-secretase activating protein (GSAP) is unlikely to be a good drug target, as during its characterization, I was unable to replicate earlier results which led the conclusion that the protein is natively unfolded and does not interact with C99. I also was the first to express, purify, and characterize the transmembrane segment of Notch 1. The membrane tolerance, cholesterol binding, and the structure of the Notch 1 TM segment were also determined with NMR. These data lead to the hypothesis that the different structural properties of the transmembrane domain of Notch 1 versus that of the amyloid precursor protein may be exploited as the basis for the development of potential Alzheimer’s therapeutics that inhibit cleavage by gamma-secretase of the amyloid precursor protein while allowing normal (healthy) cleavage of the Notch 1 transmembrane domain.