Spatial separation of β-sheet domains of β-amyloid: Disruption of each β-sheet by N-methyl amino acids

Abstract

In a recent model of β-amyloid (Aβ) fibrils, based mainly on solid-state NMR data, a molecular layer consists of two β-sheets (residues 12-23 and 31-40 of Aβ1-40), folded onto one another by a connecting "bend" structure (residues 25-29) in the side-chain dimension. In this paper, we use two N-methyl amino acids to disrupt each of the two β-sheets individually (2NMe(NTerm), residues 17 and 19; and 2NMe(CTerm), residues 37 and 39), or both of them at the same time (4NMe, with the above four N-methylated residues). Our data indicate that incorporation of two N-methyl amino acids into one β-sheet is sufficient to disrupt that sheet while leaving the other, unmodified β-sheet intact and able to form fibrils. We show, however, that disruption of each of the two β-sheets has strikingly different effects on fibrillogenesis kinetics and fibril morphology. Both 2NMe(NTerm) and 2NMe(CTerm) form fibrils at similar rates, but more slowly than that of unmodified Aβ1-40. Electron microscopy shows that 2NMe(NTerm) forms straight fibrils with fuzzy amorphous material coating the edges, while 2NMe(CTerm) forms very regular, highly twisted fibrils-in both cases, distinct from the morphology of Aβ1-40 fibrils. Both 2NMe peptides show a "CMC" approximately four times greater than that of Aβ1-40. CD spectra of these peptides also evolve differently in time: whereas the CD spectra of 2NMe(NTerm) evolve little over 10 days, those of 2NMe(CTerm) show a transition to high β-sheet content at about day 4-5. We also show that disruption of both β-sheet domains, as in 4NMe, prevents fibril formation altogether, and renders Aβ1-40 highly water soluble and monomeric, and with solvent-exposed side chains. In summary, our data show (1) that the two β-sheet domains fold in a semiautonomous manner, since disrupting each one still allows the other to fold; (2) that disruption of the N-terminal β-sheet has a more profound effect on fibrillogenesis than disruption of the C-terminal β-sheet, suggesting that the former is the more critical for the overall structure of the fibril; and (3) that disruption of both β-sheet domains renders the peptide monomeric and unable to form fibrils. © 2006 American Chemical Society.