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: 3.19). 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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    • "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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    • "We define this chain as having a stagger of À1 since the C-terminal of chain i overlaps with the N-terminal of chain i À 1. First a 20mer was generated by placing 20 chains along z-axis with a spacing of 4.8 Å[7] [77] [78] between each other. A 180° rotation around the z-axis resulted in another 20mer which completes the 40mer. "
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    ABSTRACT: Various structural models for Amyloid β fibrils have been derived from a variety of experimental techniques. However, these models cannot differentiate between the relative position of the two arms of the β hairpin called the stagger. Amyloid fibrils of various hierarchical levels form left-handed helices composed of β sheets. However it is unclear if positive, negative and zero staggers all form the macroscopic left-handed helices. To address this issue we have conducted extensive molecular dynamics simulations of Amyloid β sheets of various staggers and shown that only negative staggers lead to the experimentally observed left-handed helices while positive staggers generate the incorrect right-handed helices. This result suggests that the negative staggers are physiologically relevant structure of the Amyloid β fibrils.
    FEBS letters 07/2013; 587(16). DOI:10.1016/j.febslet.2013.06.050 · 3.34 Impact Factor
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    • "Several structural models demonstrate that the locations of the Met35 red and Met35 ox side chains are different and can vary with the aggregation state [62]. The NMR solution structure of monomeric A␤ bound to an engineered affibody protein has the side chain solvent exposed [63], while NMR-derived models of early-formed aggregates (preglobulomer and globulomers ) [64] and solid state NMR data of the amyloid fibrils has the Met35 red packed in a hydrophobic interior [65] [66]. Methionine oxidation promotes increased bond polarity and dipole moment, plus allows the sulfur atom to act as a hydrogen bond acceptor [28], all of which can weaken hydrophobic and external quaternary contacts. "
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    ABSTRACT: Oxidative stress and amyloid-β (Aβ) formation are important processes that occur in Alzheimer's disease (AD). Amyloid formation is associated with the aggregation and precipitation of the Aβ peptide, while oxidative stress results from an imbalance in pro-oxidant/antioxidant homeostasis that produces harmful reactive oxygen species. The methionine-35 (Met35) residue of the Aβ peptide plays an important role in AD oxidative stress events and the associated neurotoxicity. We and other research groups previously demonstrated that in vitro oxidation of the Met35 side-chain to the sulfoxide (Met35red → Met35ox) impedes assembly and aggregation of monomeric Aβ peptide into protofibrils, the latter being the immediate precursors of amyloid plaques. Here, we report that Met35 oxidation state affects the stability of preexisting amyloid fibrils and plaques, where the Met35red → Met35ox process leads to changes in the morphology of filaments, protofibrils, mature fibrils, and loss of Congo red birefringence in senile plaques isolated from the brains of AD patients. The most notable differences were in fibril flexibility, as evidenced by changes from straight fibrils to irregularly shaped, rope-like fibrils. These findings suggest that the Met35 oxidation state and amyloid plaque formation may be intimately linked.
    Journal of Alzheimer's disease: JAD 05/2013; 37(1). DOI:10.3233/JAD-122389 · 4.15 Impact Factor
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