PEPTIDE SYNTHESIS, OLIGOMER PREPARATION AND CONFORMATION-DEPENDENT ANTIBODY CORE

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Even though there is compelling evidence for a causal role of amyloid forming proteins in disease, the type of amyloid and its assembly state is controversial and remains to be established. Prefibrillar amyloid oli- gomers are soluble spherical aggregates of approximately 3-10 nm in diameter that have been observed for many different types of amyloids by electron and atomic force microscopy. One of the significant advances in our understanding of prefibrillar oligomers was the finding that amyloid oligomers have a common peptide backbone structure that is distinct from amyloid fibrils based on the observation that a conformation depend- ent antibody specifically recognizes a common epitope on amyloid oligomers, but not fibrils, monomers or natively folded proteins for many different types of proteins. This indicates that the antibody recognizes a ge- neric polypeptide backbone epitope that is independent of the amino acid sequence, but yet is shared in common among all types of amyloid oligomers. The anti-oligomer antibody also generically inhibits the toxic- ity of soluble oligomers examined in vitro. Since different amyloid oligomers share a common structure and they are generically toxic to cells regardless of what protein they are derived from, this suggests that they have the same primary mechanism of toxicity in degenerative diseases. A growing body of evidence sug- gests that membrane permeabilization by amyloid oligomers may represent the common, primary mecha- nism of pathogenesis of amyloid related degenerative diseases. The overall goal of this program project is to elucidate the mechanism of membrane permeabilization by amyloid prefibrillar oligomers. I support of the specific aims of the projects, Core C will provide homogene- ous preparations of prefibrillar Aft oligomers to the projects as well as fluorescent and deuterated Aftoli- gomers. Core C will also provide preparations of alternative assembly states, such as amyloid fibrils and an- nular protofibrils as controls. Core C also provides conformation dependent antibodies that specifically rec- ognize prefibrillar oligomers, fibrils and annular protofibrils for use in blocking membrane permeabilization and verifying the conformational status of the oligomer preparations during and after the experiments.