Full-length amyloid beta peptides (Aβ 1-40/42 ) form neuritic amyloid plaques in Alzheimer's disease (AD) patients and are implicated in AD pathology. However, recent transgenic animal models cast doubt on their direct role in AD pathology. Nonamyloidogenic truncated amyloid-beta fragments and Aβ ) are also found in amyloid plaques of AD and in the preamyloid lesions of Down syndrome, a model system for early-onset AD study. Very little is known about the structure and activity of these smaller peptides, although they could be the primary AD and Down syndrome pathological agents. Using complementary techniques of molecular dynamics simulations, atomic force microscopy, channel conductance measurements, calcium imaging, neuritic degeneration, and cell death assays, we show that nonamyloidogenic Aβ 9-42 and Aβ 17-42 peptides form ion channels with loosely attached subunits and elicit single-channel conductances. The subunits appear mobile, suggesting insertion of small oligomers, followed by dynamic channel assembly and dissociation. These channels allow calcium uptake in amyloid precursor protein-deficient cells. The channel mediated calcium uptake induces neurite degeneration in human cortical neurons. Channel conductance, calcium uptake, and neurite degeneration are selectively inhibited by zinc, a blocker of amyloid ion channel activity. Thus, truncated Aβ fragments could account for undefined roles played by full length Aβs and provide a unique mechanism of AD and Down syndrome pathologies. The toxicity of nonamyloidogenic peptides via an ion channel mechanism necessitates a reevaluation of the current therapeutic approaches targeting the nonamyloidogenic pathway as avenue for AD treatment.atomic force microscopy | molecular dynamics | cell calcium imaging | neurite degeneration and cell death assays | single-channel conductance A myloid-beta peptides (Aβ 1-40/42 ) produced by β-and γ-secretase processing of amyloid precursor protein (APP) in the amyloidogenic pathway are involved in Alzheimer's disease (AD) pathology. Aβ 1-40/42 peptides form β-sheet-rich ordered aggregates and soluble oligomers. Small oligomers are emerging as the predominant toxic species (1-3); the toxicity is believed to be a result of the loss of ionic homeostasis, presumably via ion channels formed in cellular membranes (4, 5). EM images of Aβ oligomers show doughnut-like morphologies (6). Atomic force microscopic (AFM) images of Aβ peptides reconstituted in lipid bilayers show heteromeric (rectangular to hexagonal) ion channel-like structures with a ∼2.0-nm central pore and 8-to 12-nm outer diameters (7,8). Electrophysiological studies show heterodisperse cationselective single-channel conductances (7)(8)(9)(10)(11)(12)(13)(14) that are consistent with features of other amyloid ion channels (6-8).On the other hand, when APP is cleaved by γ-and α-secretases, it forms the nonamyloidogenic pathway generating ∼2.6-kDa fragments (Aβ 17-40/42 ) known as the p3 peptides (15). Cleavage by γ and BACE between Tyr10 and Glu11 generates another...

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More than two dozen clinical syndromes known as amyloid diseases are characterized by the buildup of extended insoluble fibrillar deposits in tissues. These amorphous Congo red staining deposits known as amyloids exhibit a characteristic green birefringence and cross-β structure. Substantial evidence implicates oligomeric intermediates of amyloids as toxic species in the pathogenesis of these chronic disease states. A growing body of data has suggested that these toxic species form ion channels in cellular membranes causing disruption of calcium homeostasis, membrane depolarization, energy drainage, and in some cases apoptosis. Amyloid peptide channels exhibit a number of common biological properties including the universal U-shape β-strand-turn-β-strand structure, irreversible and spontaneous insertion into membranes, production of large heterogeneous single-channel conductances, relatively poor ion selectivity, inhibition by Congo red, and channel blockade by zinc. Recent evidence has suggested that increased amounts of amyloids are not only toxic to its host target cells but also possess antimicrobial activity. Furthermore, at least one human antimicrobial peptide, protegrin-1, which kills microbes by a channel-forming mechanism, has been shown to possess the ability to form extended amyloid fibrils very similar to those of classic disease-forming amyloids. In this paper, we will review the reported antimicrobial properties of amyloids and the implications of these discoveries for our understanding of amyloid structure and function.

Emerging evidence supports the ion channel mechanism for Alzheimer's disease pathophysiology wherein small β-amyloid (Aβ) oligomers insert into the cell membrane, forming toxic ion channels and destabilizing the cellular ionic homeostasis. Solid-state NMR-based data of amyloid oligomers in solution indicate that they consist of a double-layered β-sheets where each monomer folds into β-strand-turn-β-strand and the monomers are stacked atop each other. In the membrane, Aβ peptides are proposed to be β-type structures. Experimental structural data available from atomic force microscopy (AFM) imaging of Aβ oligomers in membranes reveal heterogeneous channel morphologies. Previously, we modeled the channels in a non-tilted organization, parallel with the cross-membrane normal. Here, we modeled a β-barrel-like organization. β-Barrels are common in transmembrane toxin pores, typically consisting of a monomeric chain forming a pore, organized in a single-layered β-sheet with antiparallel β-strands and a right-handed twist. Our explicit solvent molecular dynamics simulations of a range of channel sizes and polymorphic turns and comparisons of these with AFM image dimensions support a β-barrel channel organization. Different from the transmembrane β-barrels where the monomers are folded into a circular β-sheet with antiparallel β-strands stabilized by the connecting loops, these Aβ barrels consist of multimeric chains forming double β-sheets with parallel β-strands, where the strands of each monomer are connected by a turn. Although the Aβ barrels adopt the right-handed β-sheet twist, the barrels still break into heterogeneous, loosely attached subunits, in good agreement with AFM images and previous modeling. The subunits appear mobile, allowing unregulated, hence toxic, ion flux.

In Alzheimer's disease, calcium permeability through cellular membranes appears to underlie neuronal cell death. It is increasingly accepted that calcium permeability involves toxic ion channels. We modeled Alzheimer's disease ion channels of different sizes (12-mer to 36-mer) in the lipid bilayer using molecular dynamics simulations. Our Abeta channels consist of the solid-state NMR-based U-shaped beta-strand-turn-beta-strand motif. In the simulations we obtain ion-permeable channels whose subunit morphologies and shapes are consistent with electron microscopy/atomic force microscopy. In agreement with imaged channels, the simulations indicate that beta-sheet channels break into loosely associated mobile beta-sheet subunits. The preferred channel sizes (16- to 24-mer) are compatible with electron microscopy/atomic force microscopy-derived dimensions. Mobile subunits were also observed for beta-sheet channels formed by cytolytic PG-1 beta-hairpins. The emerging picture from our large-scale simulations is that toxic ion channels formed by beta-sheets spontaneously break into loosely interacting dynamic units that associate and dissociate leading to toxic ionic flux. This sharply contrasts intact conventional gated ion channels that consist of tightly interacting alpha-helices that robustly prevent ion leakage, rather than hydrogen-bonded beta-strands. The simulations suggest why conventional gated channels evolved to consist of interacting alpha-helices rather than hydrogen-bonded beta-strands that tend to break in fluidic bilayers. Nature designs folded channels but not misfolded toxic channels.

We performed multiple analyses of Tyrannosaurus rex (specimen MOR 1125) fibrous cortical and medullary tissues remaining after demineralization. The results indicate that collagen I, the main organic component of bone, has been preserved in low concentrations in these tissues. The findings were independently confirmed by mass spectrometry. We propose a possible chemical pathway that may contribute to this preservation. The presence of endogenous protein in dinosaur bone may validate hypotheses about evolutionary relationships, rates, and patterns of molecular change and degradation, as well as the chemical stability of molecules over time.

Alzheimer’s disease (AD) is a protein misfolding disease characterized by a build-up of β-amyloid (Aβ) peptide as senile plaques, uncontrolled neurodegeneration, and memory loss. AD pathology is linked to the destabilization of cellular ionic homeostasis and involves Aβ peptide-plasma membrane interactions. In principle, there are two possible ways through which disturbance of the ionic homeostasis can take place: directly, where the Aβ peptide either inserts into the membrane and creates ion-conductive pores or destabilizes the membrane organization; or, indirectly, where the Aβ peptide interacts with existing cell membrane receptors. To distinguish between these two possible types of Aβ-membrane interactions, we took advantage of the biochemical tenet that ligand-receptor interactions are stereospecific; L-amino acid peptides, but not their D-counterparts, bind to cell membrane receptors. However, with respect to the ion channel-mediated mechanism, like L-amino acids, D-amino acid peptides will also form ion channel-like structures. Using atomic force microscopy (AFM) we imaged the structures of both D- and L-enantiomers of the full length Aβ1-42 when reconstituted in lipid bilayers. AFM imaging shows that both L- and D-Aβ isomers form similar channel-like structures. Molecular dynamics (MD) simulations support the AFM imaged 3D structures. Earlier we have shown that D-Aβ1-42 channels conduct ions similarly to their L-counter parts. Taken together, our results support the direct mechanism of Aβ ion channel-mediated destabilization of ionic homeostasis rather than the indirect mechanism through Aβ interaction with membrane receptors.

Amyloid-β (Aβ) oligomers destabilize cellular ionic homeostasis, mediating Alzheimer's disease (AD). It is still unclear whether the mechanism (i) is mediated by cell surface receptors; (ii) is direct, with Aβ oligomers interacting with membrane lipids; or (iii) both mechanisms take place. Recent studies indicate that Aβ oligomers may act by either of the last two. Little is known about the oligomers’ structures and how they spontaneously insert into the membrane. Using explicit solvent molecular dynamics (MD) simulations, we show that fibril-like Aβ17-42 (p3) oligomer is capable of penetrating the membrane. Insertion is similar to that observed for protegrin-1 (PG-1), a cytolytic β-sheet-rich antimicrobial peptide (AMP). Both Aβ and PG-1 favor the amphipathic interface of the lipid bilayer in the early stage of interaction with the membrane. U-shaped Aβ oligomers are observed in solution and in the membrane, suggesting that the preformed seeds can be shared by amyloid fibrils in the growth phase and membrane toxicity. Here we provide sequential events in possible Aβ oligomer membrane-insertion pathways. We speculate that for the U-shaped motif, a trimer is the minimal oligomer size to insert effectively. We propose that monomers and dimers may insert in (apparently on-pathway) aggregation-intermediate β-hairpin state, and may (or may not) convert to a U-shape in the bilayer. Together with earlier observations, our results point to a non-specific, broadly heterogeneous landscape of membrane-inserting oligomer conformations, pathways, and membrane-mediated toxicity of β-rich oligomers.