Experimental Constraints on Quaternary Structure in Alzheimer's β-Amyloid Fibrils †

Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892-0520, USA.
Biochemistry (Impact Factor: 3.02). 02/2006; 45(2):498-512. DOI: 10.1021/bi051952q
Source: PubMed

ABSTRACT We describe solid-state nuclear magnetic resonance (NMR) measurements on fibrils formed by the 40-residue beta-amyloid peptide associated with Alzheimer's disease (Abeta(1-40)) that place constraints on the identity and symmetry of contacts between in-register, parallel beta-sheets in the fibrils. We refer to these contacts as internal and external quaternary contacts, depending on whether they are within a single molecular layer or between molecular layers. The data include (1) two-dimensional 13C-13C NMR spectra that indicate internal quaternary contacts between side chains of L17 and F19 and side chains of I32, L34, and V36, as well as external quaternary contacts between side chains of I31 and G37; (2) two-dimensional 15N-13C NMR spectra that indicate external quaternary contacts between the side chain of M35 and the peptide backbone at G33; (3) measurements of magnetic dipole-dipole couplings between the side chain carboxylate group of D23 and the side chain amine group of K28 that indicate salt bridge interactions. Isotopic dilution experiments allow us to make distinctions between intramolecular and intermolecular contacts. On the basis of these data and previously determined structural constraints from solid-state NMR and electron microscopy, we construct full molecular models using restrained molecular dynamics simulations and restrained energy minimization. These models apply to Abeta(1-40) fibrils grown with gentle agitation. We also present evidence for different internal quaternary contacts in Abeta(1-40) fibrils grown without agitation, which are morphologically distinct.

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    • "We used an A fibrillar structure from a solid-state NMR fibril model of full-length Aβ (PDB entry: 2LMO) (Petkova et al. 2006). Each GSPE molecule was prepared for Rosetta docking simulations, as described previously (Jiang et al. 2013). "
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    ABSTRACT: Epidemiological evidence that red wine consumption negatively correlates with risk of AD has led to experimental studies demonstrating that grape seed extracts inhibit the aggregation and oligomerization of Aβ in vitro and ameliorate neuropathology and behavioral deficits in a mouse model of AD. The active agent in the extracts is a mixed population of polyphenolic compounds. To evaluate the relative potency of each of these compounds, HPLC was used to fractionate the mixture into monomers, dimers, and oligomers. Each fraction was analyzed for its effect on Aβ conformational dynamics (circular dichroism), oligomerization (zero-length photochemical cross-linking), aggregation kinetics (Thioflavin T fluorescence), and morphology (electron microscopy). The relative activities of each fraction were determined on the basis of molar concentration (mol/L) or mass concentration (g/L). When molar concentration, the number concentration of each polyphenolic compound, was considered, the oligomer fraction was the most potent inhibitor of Aβ oligomerization and aggregation. However, when mass concentration, the number concentration of phenolic groups, was considered, monomers were the most potent inhibitors. To understand these ostensibly contradictory results, a model of polyphenol:Aβ complexation was developed. This model, which was found to be consistent with published X-ray crystallographic studies, offers an explanation for the effects of functional group polyvalency on inhibitor activity. Our data emphasize the importance of an in-depth understanding of the mechanism(s) underlying "concentration dependence" in inhibitor systems involving polyfunctional agents. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    Journal of Neurochemistry 07/2015; DOI:10.1111/jnc.13270 · 4.28 Impact Factor
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    • "First, mass-per-length values obtained from scanning TEM indicate that fibrils can be composed of one to five protofilaments (the minimal fibrillar entities) [80, 81]. Second, distinct orientations and modes of lateral association of protofilaments by different patterns of inter-residue interactions determine if protofilaments are oriented side-by-side [50, 82], offset from one another [76, 77], or winded around a hollow core [79]. Third, solid-state NMR demonstrated that agitated (striated) and quiescent (twisted) fibrils differ in the residues participating in the β-strands and such variations in the underlying protofilament substructure can contribute to polymorphism [59, 83]. "
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    ABSTRACT: The aggregation and deposition of the amyloid-β peptide (Aβ) in the brain has been linked with neuronal death, which progresses in the diagnostic and pathological signs of Alzheimer's disease (AD). The transition of an unstructured monomeric peptide into self-assembled and more structured aggregates is the crucial conversion from what appears to be a harmless polypeptide into a malignant form that causes synaptotoxicity and neuronal cell death. Despite efforts to identify the toxic form of Aβ, the development of effective treatments for AD is still limited by the highly transient and dynamic nature of interconverting forms of Aβ. The variability within the in vivo "pool" of different Aβ peptides is another complicating factor. Here we review the dynamical interplay between various components that influence the heterogeneous Aβ system, from intramolecular Aβ flexibility to intermolecular dynamics between various Aβ alloforms and external factors. The complex dynamics of Aβ contributes to the causative role of Aβ in the pathogenesis of AD.
    Cellular and Molecular Life Sciences CMLS 05/2014; 71(18). DOI:10.1007/s00018-014-1634-z · 5.81 Impact Factor
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    • "Solid-state NMR experiments showed that Aβ40 assemblies adopted a well-defined structure, in which residues 22–39 formed β-sheets, whereas the N-terminus (residues 1–20) was unstructured. The characteristic Asp23–Lys28 turn, which previously had been identified as a key structural feature in Aβ fibrils [89] [90], monomers [91], and oligomers [92], was present in these Aβ assemblies [84]. Overall, the NMR data suggested a model in which EGCG binds to unfolded Aβ40 by aromatic interactions. "
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    ABSTRACT: Abnormal protein folding and self-assembly causes over 30 cureless human diseases for which no disease-modifying therapies are available. The common side to all these diseases is formation of aberrant toxic protein oligomers and amyloid fibrils. Both types of assemblies are drug targets, yet each presents major challenges to drug design, discovery, and development. In this review, we focus on two small molecules that inhibit formation of toxic amyloid protein assemblies - the green-tea derivative (-)-epigallocatechin-3-gallate (EGCG), which was identified through a combination of epidemiologic data and a compound library screen, and the molecular tweezer CLR01, whose inhibitory activity was discovered in our group based on rational reasoning, and subsequently confirmed experimentally. Both compounds act in a manner that is not specific to one particular protein and thus are useful against a multitude of amyloidogenic proteins, yet they act via distinct putative mechanisms. CLR01 disrupts protein aggregation through specific binding to lysine residues, whereas the mechanisms underlying the activity of EGCG are only recently beginning to unveil. We discuss current in vitro and, where available, in vivo literature related to EGCG and CLR01's effects on amyloid beta-protein, alpha-synuclein, transthyretin, islet amyloid polypeptide, and calcitonin. We also describe the toxicity, pharmacokinetics, and mechanism of action of each compound.
    12/2013; 4(4-4):385-409. DOI:10.2478/s13380-013-0137-y
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