Browse
Submit Data
Databases
API
Help

Dataset Information

33 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

High resolution structural characterization of Aβ42 amyloid fibrils by magic angle spinning NMR.

ABSTRACT: 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 Aβ1-40 and Aβ1-42, respectively. While there have been numerous reports that structurally characterize fibrils of Aβ1-40, very little is known about the structure of amyloid fibrils of Aβ1-42, which are considered the more toxic alloform involved in AD. We have prepared isotopically (13)C/(15)N labeled AβM01-42 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 AβM01-42 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.

SUBMITTER: Colvin MT

PROVIDER: S-EPMC4623963 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Structural characterization of GNNQQNY amyloid fibrils by magic angle spinning NMR.

Project description:Several human diseases are associated with the formation of amyloid aggregates, but experimental characterization of these amyloid fibrils and their oligomeric precursors has remained challenging. Experimental and computational analysis of simpler model systems has therefore been necessary, for instance, on the peptide fragment GNNQQNY7?13 of yeast prion protein Sup35p. Expanding on a previous publication, we report here a detailed structural characterization of GNNQQNY fibrils using magic angle spinning (MAS) NMR. On the basis of additional chemical shift assignments we confirm the coexistence of three distinct peptide conformations within the fibrillar samples, as reflected in substantial chemical shift differences. Backbone torsion angle measurements indicate that the basic structure of these coexisting conformers is an extended ?-sheet. We structurally characterize a previously identified localized distortion of the ?-strand backbone specific to one of the conformers. Intermolecular contacts are consistent with each of the conformers being present in its own parallel and in-register sheet. Overall the MAS NMR data indicate a substantial difference between the structure of the fibrillar and crystalline forms of these peptides, with a clearly increased complexity in the GNNQQNY fibril structure. These experimental data can provide guidance for future work, both experimental and theoretical, and provide insights into the distinction between fibril growth and crystal formation.

| S-EPMC3026921 | biostudies-literature

Magic angle spinning NMR analysis of beta2-microglobulin amyloid fibrils in two distinct morphologies.

Project description:Beta(2)-microglobulin (beta(2)m) is the major structural component of amyloid fibrils deposited in a condition known as dialysis-related amyloidosis. Despite numerous studies that have elucidated important aspects of the fibril formation process in vitro, and a magic angle spinning (MAS) NMR study of the fibrils formed by a small peptide fragment, structural details of beta(2)m fibrils formed by the full-length 99-residue protein are largely unknown. Here, we present a site-specific MAS NMR analysis of fibrils formed by the full-length beta(2)m protein and compare spectra of fibrils prepared under two different conditions. Specifically, long straight (LS) fibrils are formed at pH 2.5, while a very different morphology denoted as worm-like (WL) fibrils is observed in preparations at pH 3.6. High-resolution MAS NMR spectra have allowed us to obtain (13)C and (15)N resonance assignments for 64 residues of beta(2)m in LS fibrils, including part of the highly mobile N-terminus. Approximately 25 residues did not yield observable signals. Chemical shift analysis of the sequentially assigned residues indicates that these fibrils contain an extensive beta-sheet core organized in a non-native manner, with a trans-P32 conformation. In contrast, WL fibrils exhibit more extensive dynamics and appear to have a smaller beta-sheet core than LS fibrils, although both cores seem to share some common elements. Our results suggest that the distinct macroscopic morphological features observed for the two types of fibrils result from variations in structure and dynamics at the molecular level.

| S-EPMC2919207 | biostudies-literature

Accurate determination of interstrand distances and alignment in amyloid fibrils by magic angle spinning NMR.

Project description:Amyloid fibrils are structurally ordered aggregates of proteins whose formation is associated with many neurodegenerative and other diseases. For that reason, their high-resolution structures are of considerable interest and have been studied using a wide range of techniques, notably electron microscopy, X-ray diffraction, and magic angle spinning (MAS) NMR. Because of the excellent resolution in the spectra, MAS NMR is uniquely capable of delivering site-specific, atomic resolution information about all levels of amyloid structure: (1) the monomer, which packs into several (2) protofilaments that in turn associate to form a (3) fibril. Building upon our high-resolution structure of the monomer of an amyloid-forming peptide from transthyretin (TTR(105-115)), we introduce single 1-(13)C labeled amino acids at seven different sites in the peptide and measure intermolecular carbonyl-carbonyl distances with an accuracy of ~0.11 A. Our results conclusively establish a parallel, in register, topology for the packing of this peptide into a ?-sheet and provide constraints essential for the determination of an atomic resolution structure of the fibril. Furthermore, the approach we employ, based on a combination of a double-quantum filtered variant of the DRAWS recoupling sequence and multispin numerical simulations in SPINEVOLUTION, is general and should be applicable to a wide range of systems.

| S-EPMC2959142 | biostudies-literature

Intermolecular structure determination of amyloid fibrils with magic-angle spinning and dynamic nuclear polarization NMR.

Project description:We describe magic-angle spinning NMR experiments designed to elucidate the interstrand architecture of amyloid fibrils. Three methods are introduced for this purpose, two being based on the analysis of long-range (13)C-(13)C correlation spectra and the third based on the identification of intermolecular interactions in (13)C-(15)N spectra. We show, in studies of fibrils formed by the 86-residue SH3 domain of PI3 kinase (PI3-SH3 or PI3K-SH3), that efficient (13)C-(13)C correlation spectra display a resonance degeneracy that establishes a parallel, in-register alignment of the proteins in the amyloid fibrils. In addition, this degeneracy can be circumvented to yield direct intermolecular constraints. The (13)C-(13)C experiments are corroborated by (15)N-(13)C correlation spectra obtained from a mixed [(15)N,(12)C]/[(14)N,(13)C] sample which directly quantify interstrand distances. Furthermore, when the spectra are recorded with signal enhancement provided by dynamic nuclear polarization (DNP) at 100 K, we demonstrate a dramatic increase (from 23 to 52) in the number of intermolecular (15)N-(13)C constraints detectable in the spectra. The increase in the information content is due to the enhanced signal intensities and to the fact that dynamic processes, leading to spectral intensity losses, are quenched at low temperatures. Thus, acquisition of low temperature spectra addresses a problem that is frequently encountered in MAS spectra of proteins. In total, the experiments provide 111 intermolecular (13)C-(13)C and (15)N-(13)C constraints that establish that the PI3-SH3 protein strands are aligned in a parallel, in-register arrangement within the amyloid fibril.

| S-EPMC3190134 | biostudies-literature

Magic angle spinning NMR of viruses.

Project description:Viruses, relatively simple pathogens, are able to replicate in many living organisms and to adapt to various environments. Conventional atomic-resolution structural biology techniques, X-ray crystallography and solution NMR spectroscopy provided abundant information on the structures of individual proteins and nucleic acids comprising viruses; however, viral assemblies are not amenable to analysis by these techniques because of their large size, insolubility, and inherent lack of long-range order. In this article, we review the recent advances in magic angle spinning NMR spectroscopy that enabled atomic-resolution analysis of structure and dynamics of large viral systems and give examples of several exciting case studies.

| S-EPMC4413014 | biostudies-literature

Cell-free synthesis of amyloid fibrils with infectious properties and amenable to sub-milligram magic-angle spinning NMR analysis.

Project description:Structural investigations of amyloid fibrils often rely on heterologous bacterial overexpression of the protein of interest. Due to their inherent hydrophobicity and tendency to aggregate as inclusion bodies, many amyloid proteins are challenging to express in bacterial systems. Cell-free protein expression is a promising alternative to classical bacterial expression to produce hydrophobic proteins and introduce NMR-active isotopes that can improve and speed up the NMR analysis. Here we implement the cell-free synthesis of the functional amyloid prion HET-s(218-289). We present an interesting case where HET-s(218-289) directly assembles into infectious fibril in the cell-free expression mixture without the requirement of denaturation procedures and purification. By introducing tailored 13C and 15N isotopes or CF3 and 13CH2F labels at strategic amino-acid positions, we demonstrate that cell-free synthesized amyloid fibrils are readily amenable to high-resolution magic-angle spinning NMR at sub-milligram quantity.

| S-EPMC9646696 | biostudies-literature

High-resolution magic angle spinning NMR spectroscopy of human osteoarthritic cartilage.

Project description:Osteoarthritis (OA) is a degenerative disease of the joints and results in changes in the biochemical composition of cartilage. Previous studies have been undertaken that have used high-resolution NMR spectroscopy to study the biochemical composition of porcine, canine and bovine cartilage. In the present study, high-resolution magical angle spinning (HR-MAS) NMR spectroscopy at 11.7 T has been used to characterize metabolites and detect differences in the spectral signature of human knee articular cartilage from non-OA healthy cadaver knees and samples acquired from severe OA patients at the time of total knee replacement surgery. A statistically significant difference in the alanine (1.47 p.p.m.), N-acetyl (2.04 p.p.m.), choline (3.25 p.p.m.) and glycine (3.55 p.p.m.) metabolite levels was observed between healthy and OA specimens. The results of the present study indicate that a decrease in the intensity of N-acetyl resonance occurs in the later stages of OA. A positive correlation of the N-acetyl levels as measured by (1)H HR-MAS NMR spectroscopy with the total proteoglycan content in the same cartilage specimens as measured by the glycosaminoglycan (GAG) assay was observed. This indicates that N-acetyl can serve as an important bio-marker of OA disease progression. A decrease in the alanine concentration in OA may be attributed to the degradation of the collagen framework with disease progression and eventual loss of the degradation products that are transported from cartilage into the synovial cavity.

| S-EPMC3299852 | biostudies-other

Structural biology of supramolecular assemblies by magic-angle spinning NMR spectroscopy.

Project description:In recent years, exciting developments in instrument technology and experimental methodology have advanced the field of magic-angle spinning (MAS) nuclear magnetic resonance (NMR) to new heights. Contemporary MAS NMR yields atomic-level insights into structure and dynamics of an astounding range of biological systems, many of which cannot be studied by other methods. With the advent of fast MAS, proton detection, and novel pulse sequences, large supramolecular assemblies, such as cytoskeletal proteins and intact viruses, are now accessible for detailed analysis. In this review, we will discuss the current MAS NMR methodologies that enable characterization of complex biomolecular systems and will present examples of applications to several classes of assemblies comprising bacterial and mammalian cytoskeleton as well as human immunodeficiency virus 1 and bacteriophage viruses. The body of work reviewed herein is representative of the recent advancements in the field, with respect to the complexity of the systems studied, the quality of the data, and the significance to the biology.

| S-EPMC5483179 | biostudies-literature

Metabolite localization in living drosophila using High Resolution Magic Angle Spinning NMR.

Project description:We have developed new methods enabling in vivo localization and identification of metabolites through their (1)H NMR signatures, in a drosophila. Metabolic profiles in localized regions were obtained using HR-MAS Slice Localized Spectroscopy and Chemical Shift Imaging at high magnetic fields. These methods enabled measurement of metabolite contents in anatomic regions of the fly, demonstrated by a decrease in ?-alanine signals in the thorax of flies showing muscle degeneration.

| S-EPMC4402646 | biostudies-literature

High-resolution molecular structure of a peptide in an amyloid fibril determined by magic angle spinning NMR spectroscopy.

Project description:Amyloid fibrils are self-assembled filamentous structures associated with protein deposition conditions including Alzheimer's disease and the transmissible spongiform encephalopathies. Despite the immense medical importance of amyloid fibrils, no atomic-resolution structures are available for these materials, because the intact fibrils are insoluble and do not form diffraction-quality 3D crystals. Here we report the high-resolution structure of a peptide fragment of the amyloidogenic protein transthyretin, TTR(105-115), in its fibrillar form, determined by magic angle spinning NMR spectroscopy. The structure resolves not only the backbone fold but also the precise conformation of the side chains. Nearly complete (13)C and (15)N resonance assignments for TTR(105-115) formed the basis for the extraction of a set of distance and dihedral angle restraints. A total of 76 self-consistent experimental measurements, including 41 restraints on 19 backbone dihedral angles and 35 (13)C-(15)N distances between 3 and 6 A were obtained from 2D and 3D NMR spectra recorded on three fibril samples uniformly (13)C, (15)N-labeled in consecutive stretches of four amino acids and used to calculate an ensemble of peptide structures. Our results indicate that TTR(105-115) adopts an extended beta-strand conformation in the amyloid fibrils such that both the main- and side-chain torsion angles are close to their optimal values. Moreover, the structure of this peptide in the fibrillar form has a degree of long-range order that is generally associated only with crystalline materials. These findings provide an explanation of the unusual stability and characteristic properties of this form of polypeptide assembly.

| S-EPMC321745 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data