Genetic system for detecting, monitoring, and evolving sitespecific proteases: Development of therapeutic proteases targeting amyloid-beta peptide
Abstract
Site-specific proteases hold enormous potential as therapeutic agents due to their ability to initiate the degradation of peptides and proteins responsible for pathogenic states in human diseases. However, endowing proteases with tailor-made substrate specificities is the principle challenge to developing such therapeutic interventions. A bacterial genetic selection system and principles of directed evolution were utilized to develop proteases possessing enzymatic activities towards novel substrates. This system was initially validated and characterized by evolving a Tobacco Etch Virus protease mutant targeting a substrate weakly recognized by the wild-type enzyme. The candidates that emerged were characterized by qualitative growth tests, as well as by quantitative assays that employ transcriptional reporters and fluorogenic substrates. This strategy was implemented towards the development of therapeutic proteases that recognize the amyloid-beta peptide, a peptide catabolite linked to the pathogenesis of Alzheimer’s Disease.
Degree
Ph.D.
Advisors
Savinov, Purdue University.
Subject Area
Genetics|Biochemistry
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