Aneta T. Petkova

A structural model for Alzheimer's β-amyloid fibrils based on experimental constraints from solid state NMR

We present a structural model for amyloid fibrils formed by the 40-residue β-amyloid peptide associated with Alzheimers disease (Aβ1–40), based on a set of experimental constraints from solid state NMR spectroscopy. The model additionally incorporates the cross-β structural motif established by x-ray fiber diffraction and satisfies constraints on Aβ1–40 fibril dimensions and mass-per-length determined from electron microscopy. Approximately the first 10 residues of Aβ1–40 are structurally disordered in the fibrils. Residues 12–24 and 30–40 adopt β-strand conformations and form parallel β-sheets through intermolecular hydrogen bonding. Residues 25–29 contain a bend of the peptide backbone that brings the two β-sheets in contact through sidechain-sidechain interactions. A single cross-β unit is then a double-layered β-sheet structure with a hydrophobic core and one hydrophobic face. The only charged sidechains in the core are those of D23 and K28, which form salt bridges. Fibrils with minimum mass-per-length and diameter consist of two cross-β units with their hydrophobic faces juxtaposed.

Cross polarization in the tilted frame: assignment and spectral simplification in heteronuclear spin systems

A frequency selective heteronuclear polarization transfer technique is introduced for rotating solids. In this method, radiofrequency fields comparable with the frequency offsets are applied to establish Hartmann-Hahn cross polarization that therefore depends explicitly on the resonance offset of the nuclei involved. Under these conditions, spectrally induced filtering in combination with cross polarization (SPECIFIC CP) can be achieved and is demonstrated to be useful for spectral simplification or assignment in heteronuclear spin pairs. The design principles are outlined and demonstrated experimentally on 13C, 15N labelled amino acids.

Supramolecular Structure in Full-Length Alzheimer's β-Amyloid Fibrils: Evidence for a Parallel β-Sheet Organization from Solid-State Nuclear Magnetic Resonance

Abstract We report constraints on the supramolecular structure of amyloid fibrils formed by the 40-residue β -amyloid peptide associated with Alzheimers disease (A β 1–40 ) obtained from solid-state nuclear magnetic resonance (NMR) measurements of intermolecular dipole-dipole couplings between 13 C labels at 11 carbon sites in residues 2 through 39. The measurements are carried out under magic-angle spinning conditions, using the constant-time finite-pulse radiofrequency-driven recoupling (fpRFDR-CT) technique. We also present one-dimensional 13 C magic-angle spinning NMR spectra of the labeled A β 1–40 samples. The fpRFDR-CT data reveal nearest-neighbor intermolecular distances of 4.8±0.5A for carbon sites from residues 12 through 39, indicating a parallel alignment of neighboring peptide chains in the predominantly β -sheet structure of the amyloid fibrils. The one-dimensional NMR spectra indicate structural order at these sites. The fpRFDR-CT data and NMR spectra also indicate structural disorder in the N-terminal segment of A β 1–40 , including the first nine residues. These results place strong constraints on any molecular-level structural model for full-length β -amyloid fibrils.

Backbone and side chain assignment strategies for multiply labeled membrane peptides and proteins in the solid state.

We demonstrate that the SPECIFIC CP technique can be used to obtain heteronuclear correlation (HETCOR) spectra of peptide backbones with greater efficiency than conventional HETCOR methods. We show that similar design principles can be employed to achieve selective homonuclear polarization transfer mediated through dipolar or scalar couplings. Both approaches are demonstrated in a tripeptide with uniform 15N and 13C labeling, and with uniform 15N labeling and natural abundance 13C. In other applications, the high efficiency of the heteronuclear SPECIFIC CP transfer allows discrimination of single amide signals in the 248-residue membrane protein bacteriorhodopsin (bR). In particular, variations are detected in the ordering of the Ala81-Arg82 peptide bond among the photocycle intermediates of bR and SPECIFIC CP is used to correlate 15N and 13C signals from the three Val-Pro peptide bonds.

Solid State NMR of Fibrillar Beta Amyloid Isoforms

The 40-43 residue-long beta amyloid peptides aggregate in the extracellular neuronal space to form amyloid plaques, which contain numerous beta amyloid fibrils. Biological interest in amyloid fibrils arises from their occurrence in amyloid diseases, such as Alzheimers disease (AD). The truncated pyroglutamate beta amyloid (p3-42) and beta amyloid (4-42) peptides are two of the most dominant isoforms found in excised amyloid material from AD brains. We investigate the self-seeding and cross-seeding behavior of the different amyloid beta isoforms, and utilize magic angle spinning NMR techniques to study the molecular structure of the fibrils formed by the amyloid beta (p3-42) and (4-42) peptides. The structures of these fibrils are compared to the available structures of the wild type and mutant beta amyloid (1-40) and (1-42) fibrils.