The Intact Human Acetylcholinesterase C-Terminal Oligomerization Domain Is α-Helical in Situ and in Isolation, but a Shorter Fragment Forms β-Sheet-Rich Amyloid Fibrils and Protofibrillar Oligomers

A 14-residue fragment of the C-terminal oligomerization domain, or T-peptide, of human acetylcholinesterase (AChE) shares sequence homology with the amyloid-β peptide implicated in Alzheimer's disease and can spontaneously self-assemble into classical amyloid fibrils under physiological conditions [Greenfield, S. A., and Vaux, D. J. (2002) Neuroscience 113, 485−492; Cottingham, M. G., Hollinshead, M. S., and Vaux, D. J. (2002) Biochemistry 41, 13539−13547]. Here we demonstrate that the conformation of this AChE586 - 599 peptide, both before and after fibril formation, is different from that of a longer peptide, T40, corresponding to the entire 40-amino acid T-peptide (residues 575−614 of AChE). This peptide is prone to homomeric hydrophobic interactions, consistent with its role in AChE subunit assembly, and possesses an α-helical structure which protects against the development of the β-sheet-rich amyloidogenic conformation favored by the shorter constituent AChE586 - 599 fragment. Using a conformation-sensitive monoclonal antibody raised against the α-helical T40 peptide, we demonstrate that the conformation of the T-peptide domain within intact AChE is antigenically indistinguishable from that of the synthetic T40 peptide. A second monoclonal antibody raised against the fibrillogenic AChE586 - 599 fragment recognizes not only β-sheet amyloid aggregates but also SDS-resistant protofibrillar oligomers. A single-antibody sandwich ELISA confirms that such oligomers exist at micromolar peptide concentrations, well below that required for formation of classical amyloid fibrils. Epitope mapping with this monoclonal antibody identifies a region near the N-terminus of the peptide that remains accessible in oligomer and fibril alike, suggesting a model for the arrangement of subunits within AChE586 - 599 protofibrils and fibrils.