Project Summary Age is universally regarded as the primary risk factor in most amyloid-related disease syndromes. There is a significant unmet need for novel, disease-modifying therapeutic strategies to target amyloidosis syndromes. In CNS diseases such as Alzheimer's disease and Parkinson's disease, and in systemic diseases such as Senile Systemic Amyloidosis and Light-chain Amyloidosis, disease-specific proteins misfold to acquire a pathological cross-β-sheet conformation characteristic of amyloidosis. Although the molecular pathophysiology of amyloid varies depending on the specific disease state, it is a common shared feature that properly folded amylogenic proteins or peptides with important physiological function become highly toxic upon misfolding. Interestingly, despite a lack of primary structure homology, amyloid proteins share this common cross-β-sheet secondary structure and similar quaternary structure when they misfold and aggregate. Molecular chaperones are naturally occurring inhibitors of amyloid protein aggregation, and understanding their detailed biochemistry could substantially advance the development of new treatments. Some chaperone-like amyloid binding proteins (CLABPs) can target amyloids from multiple sources and therefore are hypothesized to make use of the common cross-β-sheet secondary structures. These CLABPs display so-called pan-amyloid activity. However, only a small number of pan-amyloid CLABPs have been identified to date, and even fewer target soluble, early aggregates such as protofibrils. The molecular interactions between CLABPs and transient protofibrils remain unknown, constituting a major obstacle for the development of high affinity pan-amyloid therapeutics. To address this problem, the naturally occurring CLABP, NUCB1, will be thoroughly investigated. To date, this is the only CLABP that has pan-amyloid, protofibril capping activity. Using domain and mutational analyses, the critical components that are necessary and sufficient for NUCB1 to cap protofibrils will be defined. Understanding the NUCB1 capping mechanism will provide a unique opportunity to develop an innovative approach to prevent the formation of toxic amyloid protofibril conformations in human disease states. A NUCB1-immunoglobulin fusion protein will be engineered that retains pan-amyloid, protofibril binding activity. This proof-of-concept molecule has potentially impactful use as a tool for detecting protofibrils in situ, or ex vivo, and will be tested as a novel therapeutic strategy in models of age-related amyloidosis syndromes. In summary, the extraordinary properties of the naturally occurring CLABP, NUCB1, will be exploited to create a novel biotherapeutic that can “cap” and detoxify amyloid protofibrils derived from many different amyloidogenic proteins and peptides associated with human diseases of aging.