Amyloid fibrils associated with Alzheimer's disease and a wide range of other neurodegenerative diseases have a cross beta-sheet structure, where main chain hydrogen bonding occurs between beta-strands in the direction of the fibril axis. The surface of the beta-sheet has pronounced ridges and grooves when the individual beta-strands have a parallel orientation and the amino acids are in-register with one another. Here we show that in Abeta amyloid fibrils, Met35 packs against Gly33 in the C-terminus of Abeta40 and against Gly37 in the C-terminus of Abeta42. These packing interactions suggest that the protofilament subunits are displaced relative to one another in the Abeta40 and Abeta42 fibril structures. We take advantage of this corrugated structure to design a new class of inhibitors that prevent fibril formation by placing alternating glycine and aromatic residues on one face of a beta-strand. We show that peptide inhibitors based on a GxFxGxF framework disrupt sheet-to-sheet packing and inhibit the formation of mature Abeta fibrils as assayed by thioflavin T fluorescence, electron microscopy, and solid-state NMR spectroscopy. The alternating large and small amino acids in the GxFxGxF sequence are complementary to the corresponding amino acids in the IxGxMxG motif found in the C-terminal sequence of Abeta40 and Abeta42. Importantly, the designed peptide inhibitors significantly reduce the toxicity induced by Abeta42 on cultured rat cortical neurons.

The ␤-amyloid peptide (A␤) is the major constituent of the amyloid core of senile plaques found in the brain of patients with Alzheimer disease. A␤ is produced by the sequential cleavage of the amyloid precursor protein (APP) by ␤-and ␥-secretases. Cleavage of APP by ␥-secretase also generates the APP intracellular C-terminal domain (AICD) peptide, which might be involved in regulation of gene transcription. APP contains three Gly-XXX-Gly (GXXXG) motifs in its juxtamembrane and transmembrane (TM) regions. Such motifs are known to promote dimerization via close apposition of TM sequences. We demonstrate that pairwise replacement of glycines by leucines or isoleucines, but not alanines, in a GXXXG motif led to a drastic reduction of A␤40 and A␤42 secretion. ␤-Cleavage of mutant APP was not inhibited, and reduction of A␤ secretion resulted from inhibition of ␥-cleavage. It was anticipated that decreased ␥-cleavage of mutant APP would result from inhibition of its dimerization. Surprisingly, mutations of the GXXXG motif actually enhanced dimerization of the APP C-terminal fragments, possibly via a different TM ␣-helical interface. Increased dimerization of the TM APP C-terminal domain did not affect AICD production.The progressive deposition of ␤-amyloid peptide (A␤) 3 leading to the formation of senile plaques is an invariant feature of Alzheimer disease. A␤ is a 39 -43-amino acid peptide, with two major isoforms of 40 and 42 amino acids (1, 2). A␤ is produced by the amyloidogenic cleavage of its precursor, the amyloid precursor protein or APP (3).The amyloidogenic processing of APP is initiated by ␤-cleavage within the lumenal/extracellular domain of the protein.The ␤-cleavage of APP is performed by the BACE proteins (BACE1 and -2) that are integral membrane proteins belonging to the aspartyl protease family (4 -8). ␤-Cleavage produces a 99-amino acid, membrane-anchored APP C-terminal fragment (␤CTF), which is further cleaved by the ␥-secretase activity to generate A␤. The ␥-secretase activity is contained in a high molecular weight multiprotein complex formed at least by the following proteins: a presenilin (PS1 or PS2), nicastrin (Nct), Pen-2, and Aph-1 (9). The activity of the ␥-secretase complex is also required for the generation of the intracellular fragment named (APP intracellular C-terminal domain (AICD). AICD was shown to translocate to the nucleus (10, 11), and there is growing experimental evidence suggesting a role for AICD in the regulation of gene transcription (12-17) even if the identity of APP target genes remains a matter of debate (18). The ␥-secretase complex, therefore, plays a central role in the onset and progression of Alzheimer disease not only because proteolysis of ␤CTF controls the production of A␤, but it also controls the intracellular signaling associated with APP, which in turn might regulate the expression of genes involved in the disease. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "ad...