Aggregation dynamics of a 150 kDa Aβ42 oligomer: Insights from cryo electron microscopy and multimodal analysis.

Protein misfolding is a widespread phenomenon that can result in the formation of protein aggregates, which are markers of various disease states, including Alzheimer's disease (AD). In AD, amyloid beta (Aβ) peptides are key players in the disease's progression, particularly the 40- and 42- residue variants, Aβ40 and Aβ42. These peptides aggregate to form amyloid plaques and contribute to neuronal toxicity.

Aggregation Dynamics of a 150 kDa Aβ42 Oligomer: Insights from Cryo Electron Microscopy and Multimodal Analysis.

Protein misfolding is a widespread phenomenon that can result in the formation of protein aggregates, which are markers of various disease states, including Alzheimer's disease (AD). In AD, amyloid beta (Aβ) peptides, particularly Aβ40 and Aβ42, are key players in the disease's progression, as they aggregate to form amyloid plaques and contribute to neuronal toxicity. Recent research has shifted attention from solely Aβ fibrils to also include Aβ protofibrils and oligomers as potentially critical pathogenic agents.

A common pathway for detergent-assisted oligomerization of Aβ42.

Amyloid beta (Aβ) aggregation is a slow process without seeding or assisted nucleation. Sodium dodecyl sulfate (SDS) micelles stabilize Aβ42 small oligomers (in the dimer to tetramer range); subsequent SDS removal leads to a 150-kD Aβ42 oligomer. Dodecylphosphorylcholine (DPC) micelles also stabilize an Aβ42 tetramer.

Oligomer Formation by Amyloid-β42 in a Membrane-Mimicking Environment in Alzheimer's Disease.

The brains of Alzheimer's disease (AD) patients contain numerous amyloid plaques that are diagnostic of the disease. The plaques are primarily composed of the amyloidogenic peptides proteins Aβ40 and Aβ42, which are derived by the processing of the amyloid pre-cursor protein (APP) by two proteases called β-secretase and γ-secretase. Aβ42 differs from Aβ40 in having two additional hydrophobic amino acids, ILE and ALA, at the C-terminus.

Solvent Deuterium Oxide Isotope Effects on the Reactions of Organophosphorylated Acetylcholinesterase.

Organophosphates (OPs) are esters of substituted phosphates, phosphonates or phosphoramidates that react with acetylcholinesterase (AChE) by initially transferring the organophosphityl group to a serine residue in the enzyme active site, concomitant with loss of an alcohol or halide leaving group. With substituted phosphates, this transfer is followed by relatively slow hydrolysis of the organophosphoryl AChE, or dephosphorylation, that is often accompanied by an aging reaction that renders the enzyme irreversibly inactivated. Aging is a dealkylation that converts the phosphate triester to a diester.

Cathepsin D regulates cerebral Aβ42/40 ratios via differential degradation of Aβ42 and Aβ40.

Background: Cathepsin D (CatD) is a lysosomal protease that degrades both the amyloid β-protein (Aβ) and the microtubule-associated protein, tau, and has been genetically linked to late-onset Alzheimer disease (AD). Here, we sought to examine the consequences of genetic deletion of CatD on Aβ proteostasis in vivo and to more completely characterize the degradation of Aβ42 and Aβ40 by CatD.

Methods: We quantified Aβ degradation rates and levels of endogenous Aβ42 and Aβ40 in the brains of CatD-null (CatD-KO), heterozygous null (CatD-HET), and wild-type (WT) control mice.

Out-of-Register Parallel β-Sheets and Antiparallel β-Sheets Coexist in 150-kDa Oligomers Formed by Amyloid-β(1-42).

We present solid-state NMR measurements of β-strand secondary structure and inter-strand organization within a 150-kDa oligomeric aggregate of the 42-residue variant of the Alzheimer's amyloid-β peptide (Aβ(1-42)). We build upon our previous report of a β-strand spanned by residues 30-42, which arranges into an antiparallel β-sheet. New results presented here indicate that there is a second β-strand formed by residues 11-24.

A Second Look at the Crystal Structures of Acetylcholinesterase in Complex with Tacrine Derivatives Provides Insights Concerning Catalytic Intermediates and the Design of Specific Insecticides.

Over recent decades, crystallographic software for data processing and structure refinement has improved dramatically, resulting in more accurate and detailed crystal structures. It is, therefore, sometimes valuable to have a second look at "old" diffraction data, especially when earlier interpretation of the electron density maps was rather difficult. Here, we present updated crystal structures of acetylcholinesterase (AChE) originally published in [Harel et al.

Rate-limiting step in the decarbamoylation of acetylcholinesterases with large carbamoyl groups.

Carbamates are esters of substituted carbamic acids that react with acetylcholinesterase (AChE) by initially transferring the carbamoyl group to a serine residue in the enzyme active site accompanied by loss of the carbamate leaving group followed by hydrolysis of the carbamoyl enzyme. This hydrolysis, or decarbamoylation, is relatively slow, and half-lives of carbamoylated AChEs range from 4 min to more than 30 days. Therefore, carbamates are effective AChE inhibitors that have been developed as insecticides and as therapeutic agents.

VEGF receptor-1 modulates amyloid β 1-42 oligomer-induced senescence in brain endothelial cells.

Aggregated amyloid β (Aβ) peptides in the Alzheimer's disease (AD) brain are hypothesized to trigger several downstream pathologies, including cerebrovascular dysfunction. Previous studies have shown that Aβ peptides can have antiangiogenic properties, which may contribute to vascular dysfunction in the early stages of the disease process. We have generated data showing that brain endothelial cells (ECs) exposed to toxic Aβ1-42 oligomers can readily enter a senescence phenotype.

Decarbamoylation of acetylcholinesterases is markedly slowed as carbamoyl groups increase in size.

Carbamates are esters of substituted carbamic acids that react with acetylcholinesterase (AChE) by initially transferring the carbamoyl group to a serine residue in the enzyme active site accompanied by loss of the carbamate leaving group followed by hydrolysis of the carbamoyl enzyme. This hydrolysis, or decarbamoylation, is relatively slow, and half-lives of carbamoylated AChEs range from 4 min to more than 30 days. Therefore, carbamates are effective AChE inhibitors that have been developed as insecticides and as therapeutic agents.

Comparison of the Binding of Reversible Inhibitors to Human Butyrylcholinesterase and Acetylcholinesterase: A Crystallographic, Kinetic and Calorimetric Study.

Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) hydrolyze the neurotransmitter acetylcholine and, thereby, function as coregulators of cholinergic neurotransmission. Although closely related, these enzymes display very different substrate specificities that only partially overlap. This disparity is largely due to differences in the number of aromatic residues lining the active site gorge, which leads to large differences in the shape of the gorge and potentially to distinct interactions with an individual ligand.

Distinct spatiotemporal accumulation of N-truncated and full-length amyloid-β42 in Alzheimer's disease.

Accumulation of amyloid-β peptides is a dominant feature in the pathogenesis of Alzheimer's disease; however, it is not clear how individual amyloid-β species accumulate and affect other neuropathological and clinical features in the disease. Thus, we compared the accumulation of N-terminally truncated amyloid-β and full-length amyloid-β, depending on disease stage as well as brain area, and determined how these amyloid-β species respectively correlate with clinicopathological features of Alzheimer's disease. To this end, the amounts of amyloid-β species and other proteins related to amyloid-β metabolism or Alzheimer's disease were quantified by enzyme-linked immunosorbent assays (ELISA) or theoretically calculated in 12 brain regions, including neocortical, limbic and subcortical areas from Alzheimer's disease cases (n = 19), neurologically normal elderly without amyloid-β accumulation (normal ageing, n = 13), and neurologically normal elderly with cortical amyloid-β accumulation (pathological ageing, n = 15).

Blood-Based Oligomeric and Other Protein Variant Biomarkers to Facilitate Pre-Symptomatic Diagnosis and Staging of Alzheimer's Disease.

Oligomeric forms of amyloid-β (Aβ), tau, and TDP-43 play important roles in Alzheimer's disease (AD), and therefore are promising biomarkers. We previously generated single chain antibody fragments (scFvs) that selectively bind disease-related variants of these proteins including A4, C6T, and E1, which bind different oligomeric Aβ variants; D11C, which binds oligomeric tau; and AD-TDP1 and AD-TDP2, which bind disease related TDP-43 variants. To determine the utility of these disease-related variants as early biomarkers, we first analyzed 11 human sera samples obtained ∼2 years prior to an initial mild cognitive impairment (MCI) diagnosis.

Development of acetophenone ligands as potential neuroimaging agents for cholinesterases.

Association of cholinesterase with β-amyloid plaques and tau neurofibrillary tangles in Alzheimer's disease offers an opportunity to detect disease pathology during life. Achieving this requires development of radiolabelled cholinesterase ligands with high enzyme affinity. Various fluorinated acetophenone derivatives bind to acetylcholinesterase with high affinity, including 2,2,2-trifluoro-1-(3-dimethylaminophenyl)ethanone (1) and 1-(3-tert-butylphenyl)-2,2,2-trifluoroethanone (2).

Hopeahainol A binds reversibly at the acetylcholinesterase (AChE) peripheral site and inhibits enzyme activity with a novel higher order concentration dependence.

Natural product inhibitors of AChE are of interest both because they offer promise as inexpensive drugs for symptomatic relief in Alzheimer's disease and because they may provide insights into the structural features of the AChE catalytic site. Hopeahainol A is an uncharged polyphenol AChE inhibitor from the stem bark of Hopea hainanensis with a constrained, partially dearomatized bicyclic core. Molecular modeling indicates that hopeahainol A binds at the entrance of the long but narrow AChE active site gorge because it is too bulky to be accommodated within the gorge without severe distortion of the gorge as depicted in AChE crystal structures.

Opposing roles of the triggering receptor expressed on myeloid cells 2 and triggering receptor expressed on myeloid cells-like transcript 2 in microglia activation.

Mutations in triggering receptor expressed on myeloid cells 2 (TREM2), which has been proposed to regulate the inflammatory responses and the clearance of apoptotic neurons and/or amyloid-β, are genetically linked to increased risk for late-onset Alzheimer's disease (AD). Interestingly, a missense variant in TREM-like transcript 2 (TREML2), a structurally similar protein encoded by the same gene cluster with TREM2 on chromosome 6, has been shown to protect against AD. However, the molecular mechanisms by which TREM2 and TREML2 regulate the pathogenesis of AD, and their functional relationship, if any, remain unclear.

Selective Targeting of Extracellular Insulin-Degrading Enzyme by Quasi-Irreversible Thiol-Modifying Inhibitors.

Many therapeutically important enzymes are present in multiple cellular compartments, where they can carry out markedly different functions; thus, there is a need for pharmacological strategies to selectively manipulate distinct pools of target enzymes. Insulin-degrading enzyme (IDE) is a thiol-sensitive zinc-metallopeptidase that hydrolyzes diverse peptide substrates in both the cytosol and the extracellular space, but current genetic and pharmacological approaches are incapable of selectively inhibiting the protease in specific subcellular compartments. Here, we describe the discovery, characterization, and kinetics-based optimization of potent benzoisothiazolone-based inhibitors that, by virtue of a unique quasi-irreversible mode of inhibition, exclusively inhibit extracellular IDE.

Antiparallel β-Sheet Structure within the C-Terminal Region of 42-Residue Alzheimer's Amyloid-β Peptides When They Form 150-kDa Oligomers.

Understanding the molecular structures of amyloid-β (Aβ) oligomers and underlying assembly pathways will advance our understanding of Alzheimer's disease (AD) at the molecular level. This understanding could contribute to disease prevention, diagnosis, and treatment strategies, as oligomers play a central role in AD pathology. We have recently presented a procedure for production of 150-kDa oligomeric samples of Aβ(1-42) (the 42-residue variant of the Aβ peptide) that are compatible with solid-state nuclear magnetic resonance (NMR) analysis, and we have shown that these oligomers and amyloid fibrils differ in intermolecular arrangement of β-strands.

Structures of human acetylcholinesterase bound to dihydrotanshinone I and territrem B show peripheral site flexibility.

Acetylcholinesterase is a critical enzyme that regulates neurotransmission by degrading the neurotransmitter acetylcholine in synapses of the nervous system. It is an important target for both therapeutic drugs that treat Alzheimer's disease and chemical warfare agents that cripple the nervous system and cause death through paralysis. The enzyme has both catalytic and peripheral sites to which inhibitors may bind.

Acetylcholinesterase complexes with the natural product inhibitors dihydrotanshinone I and territrem B: binding site assignment from inhibitor competition and validation through crystal structure determination.

Acetylcholinesterase (AChE) is a critical enzyme that regulates neurotransmission by degrading the neurotransmitter acetylcholine in synapses of the nervous system. It is an important target for both therapeutic drugs that treat Alzheimer's disease and organophosphate (OP) chemical warfare agents that cripple the nervous system and cause death through paralysis. We are exploring a strategy to design compounds that bind tightly at or near a peripheral or P-site near the mouth of the AChE active site gorge and exclude OPs from the active site while interfering minimally with the passage of acetylcholine.

The natural product dihydrotanshinone I provides a prototype for uncharged inhibitors that bind specifically to the acetylcholinesterase peripheral site with nanomolar affinity.

Cholinergic synaptic transmission often requires extremely rapid hydrolysis of acetylcholine by acetylcholinesterase (AChE). AChE is inactivated by organophosphates (OPs) in chemical warfare nerve agents. The resulting accumulation of acetylcholine disrupts cholinergic synaptic transmission and can lead to death.

The Alzheimer's amyloid-β(1-42) peptide forms off-pathway oligomers and fibrils that are distinguished structurally by intermolecular organization.

Increasing evidence suggests that soluble aggregates of amyloid-β (Aβ) initiate the neurotoxicity that eventually leads to dementia in Alzheimer's disease. Knowledge on soluble aggregate structures will enhance our understanding of the relationship between structures and toxicities. Our group has reported a stable and homogeneous preparation of Aβ(1-42) oligomers that has been characterized by various biophysical techniques.

Hydrolysis of low concentrations of the acetylthiocholine analogs acetyl(homo)thiocholine and acetyl(nor)thiocholine by acetylcholinesterase may be limited by selective gating at the enzyme peripheral site.

Hydrolysis of acetylcholine by acetylcholinesterase (AChE) is extremely rapid, with a second-order hydrolysis rate constant k(E) (often denoted k(cat)/K(M)) that approaches 10(8) M(-1) s(-1). AChE contains a deep active site gorge with two sites of ligand binding, an acylation site (or A-site) containing the catalytic triad at the base of the gorge and a peripheral site (or P-site) near the gorge entrance. The P-site is known to contribute to catalytic efficiency with acetylthiocholine (AcSCh) by transiently trapping the substrate in a low affinity complex on its way to the A-site, where a short-lived acyl enzyme intermediate is produced.