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Solid-support electron paramagnetic resonance (EPR) studies of A?40 monomers reveal a structured state with three ordered segments.

ABSTRACT: Alzheimer disease is associated with the pathological accumulation of amyloid-? peptide (A?) in the brain. Soluble A? oligomers formed during early aggregation process are believed to be neurotoxins and causative agents in Alzheimer disease. A? monomer is the building block for amyloid assemblies. A comprehensive understanding of the structural features of A? monomer is crucial for delineating the mechanism of A? oligomerization. Here we investigated the structures of A?40 monomer using a solid-support approach, in which A?40 monomers are tethered on the solid support via an N-terminal His tag to prevent further aggregation. EPR spectra of tethered A?40 with spin labels at 18 different positions show that A?40 monomers adopt a completely disordered structure under denaturing conditions. Under native conditions, however, EPR spectra suggest that A?40 monomers adopt both a disordered state and a structured state. The structured state of A?40 monomer has three more ordered segments at 14-18, 29-30, and 38-40. Interactions between these segments may stabilize the structured state, which likely plays an important role in A? aggregation.

SUBMITTER: Gu L

PROVIDER: S-EPMC3308762 | biostudies-literature | 2012 Mar

REPOSITORIES: biostudies-literature

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Solid-support electron paramagnetic resonance (EPR) studies of Aβ40 monomers reveal a structured state with three ordered segments.

Gu Lei L Ngo Sam S Guo Zhefeng Z

The Journal of biological chemistry 20120125 12

Alzheimer disease is associated with the pathological accumulation of amyloid-β peptide (Aβ) in the brain. Soluble Aβ oligomers formed during early aggregation process are believed to be neurotoxins and causative agents in Alzheimer disease. Aβ monomer is the building block for amyloid assemblies. A comprehensive understanding of the structural features of Aβ monomer is crucial for delineating the mechanism of Aβ oligomerization. Here we investigated the structures of Aβ40 monomer using a solid- ...[more]

PMID: 22277652

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Project description:PurposePatients with hyper- vs. hypo-inflammatory subphenotypes of acute respiratory distress syndrome (ARDS) exhibit different clinical outcomes. Inflammation increases the production of reactive oxygen species (ROS) and increased ROS contributes to the severity of illness. Our long-term goal is to develop electron paramagnetic resonance (EPR) imaging of lungs in vivo to precisely measure superoxide production in ARDS in real time. As a first step, this requires the development of in vivo EPR methods for quantifying superoxide generation in the lung during injury, and testing if such superoxide measurements can differentiate between susceptible and protected mouse strains.ProceduresIn WT mice, mice lacking total body extracellular superoxide dismutase (EC-SOD) (KO), or mice overexpressing lung EC-SOD (Tg), lung injury was induced with intraperitoneal (IP) lipopolysaccharide (LPS) (10 mg/kg). At 24 h after LPS treatment, mice were injected with the cyclic hydroxylamines 1-hydroxy-3-carboxy-2,2,5,5-tetramethylpyrrolidine hydrochloride (CPH) or 4-acetoxymethoxycarbonyl-1-hydroxy-2,2,5,5-tetramethylpyrrolidine-3-carboxylic acid (DCP-AM-H) probes to detect, respectively, cellular and mitochondrial ROS - specifically superoxide. Several probe delivery strategies were tested. Lung tissue was collected up to one hour after probe administration and assayed by EPR.ResultsAs measured by X-band EPR, cellular and mitochondrial superoxide increased in the lungs of LPS-treated mice compared to control. Lung cellular superoxide was increased in EC-SOD KO mice and decreased in EC-SOD Tg mice compared to WT. We also validated an intratracheal (IT) delivery method, which enhanced the lung signal for both spin probes compared to IP administration.ConclusionsWe have developed protocols for delivering EPR spin probes in vivo, allowing detection of cellular and mitochondrial superoxide in lung injury by EPR. Superoxide measurements by EPR could differentiate mice with and without lung injury, as well as mouse strains with different disease susceptibilities. We expect these protocols to capture real-time superoxide production and enable evaluation of lung EPR imaging as a potential clinical tool for subphenotyping ARDS patients based on redox status.

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Solid-support electron paramagnetic resonance (EPR) studies of A?40 monomers reveal a structured state with three ordered segments.

Publications

Solid-support electron paramagnetic resonance (EPR) studies of Aβ40 monomers reveal a structured state with three ordered segments.

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