Localization of a fibrillar amyloid β-protein binding domain on its precursor

Abstract

Deposition of fibrillar amyloid-β protein (Aβ) in senile plaques and in the walls of cerebral blood vessels is a key pathological feature of Alzheimer's disease and certain related disorders. Fibrillar Aβ deposition is intimately associated with neuronal and cerebrovascular cell death both in vivo and in vitro. Similarly, accumulation of the Aβ protein precursor (AβPP) is also observed at sites of fibrillar Aβ deposition. Recently, we reported that fibrillar Aβ, but not unassembled Aβ, promotes the specific binding of AβPP through its cysteine-rich, amino-terminal region (Melchor, J. P., and Van Nostrand, W. E. (2000) J. Biol. Chem. 275, 9782-9791). In the present study we sought to determine the precise site on AβPP that facilitates its binding to fibrillar Aβ. A series of synthesized overlapping peptides spanning the cysteine-rich, amino-terminal region of AβPP were used as competitors for AβPP binding to fibrillar Aβ. A peptide spanning residues 105-119 of AβPP competitively inhibited AβPP binding to fibrillar Aβ in a solid-phase binding assay and on the surface of cultured human cerebrovascular smooth muscle cells. Alanine-scanning mutagenesis of residues 105-117 within glutathione S-transferase (GST)-AβPP-(18-119) revealed that His110, Val112, and Ile113 are key residues that facilitate AβPP binding to fibrillar Aβ. These specific residues belong to a common β-strand within this region of AβPP. Wild-type GST-AβPP-(18-119) protected cultured human cerebrovascular smooth muscle cells from Aβ-induced toxicity whereas H110A mutant GST-AβPP-(18-119) did not. Wild-type GST-AβPP-(18-119) bound to different isoforms of fibrillar Aβ and fibrillar amylin peptides whereas H110A mutant and I113A mutant GST-AβPP-(18-119) were substantially less efficient binding to each fibrillar peptide. We conclude that His110, Val112, and Ile113, residing in a common β-strand region within AβPP-(18-119), comprise a domain that mediates the binding of AβPP to fibrillar peptides.