Accumulation of the neurotoxic Abeta peptide is considered to be a major etiological aspect of Alzheimer's disease (AD). Thus, strategies that reduce or normalize Abeta levels are important to consider as avenues for the treatment of AD. Since biosynthetic and clearance activities regulate the steady state level of Abeta, either or both of these activities could be manipulated to reduce Abeta levels. IDE is a protease that degrades Abeta. Thus, enhancing IDE activity could theoretically improve Abeta clearance and slow AD progression. This study is focused on understanding the enzymatic properties of IDE as they pertain to Abeta degradation and the cleavage of other amyloidogenic peptides. First, we will screen compound libraries for small molecule activators of IDE using a biochemical assay to monitor cleavage of a FRET-based substrate. The assay is readily amenable for high-throughput methods, and our approach is similar to one successfully used for the identification of activators and inhibitors of other enzymes that we study. The kinetic parameters of activated IDE will be further characterized using our FRET-based substrate and other known substrates of IDE, such as Abeta, amylin, and insulin, using established assays. Activators of IDE will represent important new tools for understanding the function of IDE, and may hold potential for enhancing Abeta clearance via drug-based therapeutic strategies. Second, we will investigate hyperactive mutants of IDE. We will determine whether cysteine mutant alleles of IDE that are hyperactive toward insulin are also hyperactive toward other IDE substrates, including Abeta. Concurrently, we will isolate novel hyperactive alleles of IDE by taking advantage of computational methods and/or a yeast system that we have developed for high throughput molecular genetics of IDE. We have already demonstrated the utility of our yeast system in investigating structure-function relationships of IDE. We expect that hyperactive alleles could be used to enhance Abeta clearance via enzyme-based therapeutic strategies. By detailing the function of IDE, we intend to gain a better understanding of the role of IDE in Abeta clearance and its potential as a therapeutic agent for AD..
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