High Resolution Structural Characterization of Aβ_(42) Amyloid Fibrils by Magic Angle Spinning NMR

摘要

The presence of amyloid plaques composed of amyloid beta (Aβ) fibrils is a hallmark of Alzheimer's disease (AD). The Aβ peptide is present as several length variants with two common alloforms consisting of 40 and 42 amino acids, denoted Aβ_(1-40) and Aβ_(1-42), respectively. While there have been numerous reports that structurally characterize fibrils of Aβ_(1-40), very little is known about the structure of amyloid fibrils of Aβ_(1-42), which are considered the more toxic alloform involved in AD. We have prepared isotopically ~(13)C/~(15)N labeled Aβ_(M01-42) fibrils in vitro from recombinant protein and examined their ~(13)C-~(13)C and ~(13)C-~(15)N magic angle spinning (MAS) NMR spectra. In contrast to several other studies of Aβ fibrils, we observe spectra with excellent resolution and a single set of chemical shifts, suggesting the presence of a single fibril morphology. We report the initial structural characterization of Aβ_(M01-42) fibrils utilizing ~(13)C and ~(15)N shift assignments of 38 of the 43 residues, including the backbone and side chains, obtained through a series of cross-polarization based 2D and 3D ~(13)C-~(13)C, ~(13)C-~(15)N MAS NMR experiments for rigid residues along with J-based 2D TOBSY experiments for dynamic residues. We find that the first ～5 residues are dynamic and most efficiently detected in a J-based TOBSY spectrum. In contrast, residues 16-42 are easily observed in cross-polarization experiments and most likely form the amyloid core. Calculation of ψ and φ dihedral angles from the chemical shift assignments indicate that 4 β-strands are present in the fibril's secondary structure.