Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Single-particle characterization of A? oligomers in solution.

ABSTRACT: Determining the pathological role of amyloids in amyloid-associated diseases will require a method for characterizing the dynamic distributions in size and shape of amyloid oligomers with high resolution. Here, we explored the potential of resistive-pulse sensing through lipid bilayer-coated nanopores to measure the size of individual amyloid-? oligomers directly in solution and without chemical modification. This method classified individual amyloid-? aggregates as spherical oligomers, protofibrils, or mature fibers and made it possible to account for the large heterogeneity of amyloid-? aggregate sizes. The approach revealed the distribution of protofibrillar lengths (12- to 155 -mer) as well as the average cross-sectional area of protofibrils and fibers.

SUBMITTER: Yusko EC

PROVIDER: S-EPMC3418869 | biostudies-literature | 2012 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Single-particle characterization of Aβ oligomers in solution.

Yusko Erik C EC Prangkio Panchika P Sept David D Rollings Ryan C RC Li Jiali J Mayer Michael M

ACS nano 20120621 7

Determining the pathological role of amyloids in amyloid-associated diseases will require a method for characterizing the dynamic distributions in size and shape of amyloid oligomers with high resolution. Here, we explored the potential of resistive-pulse sensing through lipid bilayer-coated nanopores to measure the size of individual amyloid-β oligomers directly in solution and without chemical modification. This method classified individual amyloid-β aggregates as spherical oligomers, protofib ...[more]

PMID: 22686709

Similar Datasets

Direct characterization of amyloidogenic oligomers by single-molecule fluorescence.

Project description:A key issue in understanding the pathogenic conditions associated with the aberrant aggregation of misfolded proteins is the identification and characterization of species formed during the aggregation process. Probing the nature of such species has, however, proved to be extremely challenging to conventional techniques because of their transient and heterogeneous character. We describe here the application of a two-color single-molecule fluorescence technique to examine the assembly of oligomeric species formed during the aggregation of the SH3 domain of PI3 kinase. The single-molecule experiments show that the species formed at the stage of the reaction where aggregates have previously been found to be maximally cytotoxic are a heterogeneous ensemble of oligomers with a median size of 38 +/- 10 molecules. This number is remarkably similar to estimates from bulk measurements of the critical size of species observed to seed ordered fibril formation and of the most infective form of prion particles. Moreover, although the size distribution of the SH3 oligomers remains virtually constant as the time of aggregation increases, their stability increases substantially. These findings together provide direct evidence for a general mechanism of amyloid aggregation in which the stable cross-beta structure emerges via internal reorganization of disordered oligomers formed during the lag phase of the self-assembly reaction.

| S-EPMC2567173 | biostudies-literature

Force field-dependent solution properties of glycine oligomers.

Project description:Molecular simulations can be used to study disordered polypeptide systems and to generate hypotheses on the underlying structural and thermodynamic mechanisms that govern their function. As the number of disordered protein systems investigated with simulations increase, it is important to understand how particular force fields affect the structural properties of disordered polypeptides in solution. To this end, we performed a comparative structural analysis of Gly(3) and Gly(10) in aqueous solution from all atom, microsecond molecular dynamics (MD) simulations using the CHARMM 27 (C27), CHARMM 36 (C36), and Amber ff12SB force fields. For each force field, Gly(3) and Gly(10) were simulated for at least 300 ns and 1 ?s, respectively. Simulating oligoglycines of two different lengths allows us to evaluate how force field effects depend on polypeptide length. Using a variety of structural metrics (e.g., end-to-end distance, radius of gyration, dihedral angle distributions), we characterize the distribution of oligoglycine conformers for each force field and show that each sample conformation space differently, yielding considerably different structural tendencies of the same oligoglycine model in solution. Notably, we find that C36 samples more extended oligoglycine structures than both C27 and ff12SB.

| S-EPMC4450816 | biostudies-literature

Routine single particle CryoEM sample and grid characterization by tomography.

Project description:Single particle cryo-electron microscopy (cryoEM) is often performed under the assumption that particles are not adsorbed to the air-water interfaces and in thin, vitreous ice. In this study, we performed fiducial-less tomography on over 50 different cryoEM grid/sample preparations to determine the particle distribution within the ice and the overall geometry of the ice in grid holes. Surprisingly, by studying particles in holes in 3D from over 1000 tomograms, we have determined that the vast majority of particles (approximately 90%) are adsorbed to an air-water interface. The implications of this observation are wide-ranging, with potential ramifications regarding protein denaturation, conformational change, and preferred orientation. We also show that fiducial-less cryo-electron tomography on single particle grids may be used to determine ice thickness, optimal single particle collection areas and strategies, particle heterogeneity, and de novo models for template picking and single particle alignment.

| S-EPMC5999397 | biostudies-literature

Characterization of lipid-based nanomedicines at the single-particle level.

Project description:Lipid-based nanomedicines (LBNMs), including liposomes, lipid nanoparticles (LNPs) and extracellular vesicles (EVs), are recognized as one of the most clinically acceptable nano-formulations. However, the bench-to-bedside translation efficiency is far from satisfactory, mainly due to the lack of in-depth understanding of their physical and biochemical attributes at the single-particle level. In this review, we first give a brief introduction of LBNMs, highlighting some milestones and related scientific and clinical achievements in the past several decades, as well as the grand challenges in the characterization of LBNMs. Next, we present an overview of each category of LBNMs as well as the core properties that largely dictate their biological characteristics and clinical performance, such as size distribution, particle concentration, morphology, drug encapsulation and surface properties. Then, the recent applications of several analytical techniques including electron microscopy, atomic force microscopy, fluorescence microscopy, Raman microscopy, nanoparticle tracking analysis, tunable resistive pulse sensing and flow cytometry on the single-particle characterization of LBNMs are thoroughly discussed. Particularly, the comparative advantages of the newly developed nano-flow cytometry that enables quantitative analysis of both the physical and biochemical characteristics of LBNMs smaller than 40 nm with high throughput and statistical robustness are emphasized. The overall aim of this review article is to illustrate the importance, challenges and achievements associated with single-particle characterization of LBNMs.

| S-EPMC11197724 | biostudies-literature

Characterization of AAV vector particle stability at the single-capsid level.

Project description:Virus families have evolved different strategies for genome uncoating, which are also followed by recombinant vectors. Vectors derived from adeno-associated viruses (AAV) are considered as leading delivery tools for in vivo gene transfer, and in particular gene therapy. Using a combination of atomic force microscopy (AFM), biochemical experiments, and physical modeling, we investigated here the physical properties and stability of AAV vector particles. We first compared the morphological properties of AAV vectors derived from two different serotypes (AAV8 and AAV9). Furthermore, we triggered ssDNA uncoating by incubating vector particles to increasing controlled temperatures. Our analyses, performed at the single-particle level, indicate that genome release can occur in vitro via two alternative pathways: either the capsid remains intact and ejects linearly the ssDNA molecule, or the capsid is ruptured, leaving ssDNA in a compact entangled conformation. The analysis of the length distributions of ejected genomes further revealed a two-step ejection behavior. We propose a kinetic model aimed at quantitatively describing the evolution of capsids and genomes along the different pathways, as a function of time and temperature. This model allows quantifying the relative stability of AAV8 and AAV9 particles.

| S-EPMC5928021 | biostudies-literature

Single-Molecule Characterization of the Interactions between Extracellular Chaperones and Toxic ?-Synuclein Oligomers.

Project description:The aberrant aggregation of ?-synuclein is associated with several human diseases, collectively termed the ?-synucleinopathies, which includes Parkinson's disease. The progression of these diseases is, in part, mediated by extracellular ?-synuclein oligomers that may exert effects through several mechanisms, including prion-like transfer, direct cytotoxicity, and pro-inflammatory actions. In this study, we show that two abundant extracellular chaperones, clusterin and ?2-macroglobulin, directly bind to exposed hydrophobic regions on the surface of ?-synuclein oligomers. Using single-molecule fluorescence techniques, we found that clusterin, unlike ?2-macroglobulin, exhibits differential binding to ?-synuclein oligomers that may be related to structural differences between two previously described forms of ?S oligomers. The binding of both chaperones reduces the ability of the oligomers to permeabilize lipid membranes and prevents an oligomer-induced increase in ROS production in cultured neuronal cells. Taken together, these data suggest a neuroprotective role for extracellular chaperones in suppressing the toxicity associated with ?-synuclein oligomers.

| S-EPMC6024880 | biostudies-other

A?42 pentamers/hexamers are the smallest detectable oligomers in solution.

Project description:Amyloid ? (A?) oligomers may play a decisive role in Alzheimer's disease related neurodegeneration, but their structural properties are poorly understood. In this report, sedimentation velocity centrifugation, small angle neutron scattering (SANS) and molecular modelling were used to identify the small oligomeric species formed by the 42 amino acid residue long isoform of A? (A?42) in solution, characterized by a sedimentation coefficient of 2.56?S, and a radius of gyration between 2 and 4?nm. The measured sedimentation coefficient is in close agreement with the sedimentation coefficient calculated for A?42 hexamers using MD simulations at µM concentration. To the best of our knowledge this is the first report detailing the A?42 oligomeric species by SANS measurements. Our results demonstrate that the smallest detectable species in solution are penta- to hexamers. No evidences for the presence of dimers, trimers or tetramers were found, although the existence of those A?42 oligomers at measurable quantities had been reported frequently.

| S-EPMC5449387 | biostudies-literature

Peptidines: glycine-amidine-based oligomers for solution- and solid-phase synthesis.

Project description:Efforts to emulate biological oligomers have given rise to a host of useful technologies, ranging from solid-phase peptide and nucleic acid synthesis to various peptidomimetic platforms. Herein we introduce a novel class of peptide-like oligomers called "peptidines" wherein each carbonyl O-atom within poly-N-alkyl glycine oligomers is replaced with a functionalized N-atom. Compared to peptoids or peptides, the presence of this amidine N-substituent in peptidines effectively doubles the number of diversification sites per monomeric unit, and can decrease their overall conformational flexibility. We have developed iterative solution- and solid-phase protocols for the straightforward assembly of peptidines containing diverse backbone and amidine substituents, derived from readily available primary and secondary amines. We have also performed crystallographic and computational studies, which demonstrate a strong preference for the trans (E) amidine geometry. Given their straightforward synthetic preparation and high functional group density, peptidines have the potential to serve as useful tools for library generation, peptide mimicry, and the identification of biologically active small molecules.

| S-EPMC6006957 | biostudies-literature

HaloTag display enables quantitative single-particle characterization and functionalization of engineered extracellular vesicles.

Project description:Extracellular vesicles (EVs) play key roles in diverse biological processes, transport biomolecules between cells, and have been engineered for therapeutic applications. A useful EV bioengineering strategy is to express engineered proteins on the EV surface to confer targeting, bioactivity, and other properties. Measuring how incorporation varies across a population of EVs is important for characterizing such materials and understanding their function, yet it remains challenging to quantitatively characterize the absolute number of engineered proteins incorporated at single-EV resolution. To address these needs, we developed a HaloTag-based characterization platform in which dyes or other synthetic species can be covalently and stoichiometrically attached to engineered proteins on the EV surface. To evaluate this system, we employed several orthogonal quantification methods, including flow cytometry and fluorescence microscopy, and found that HaloTag-mediated quantification is generally robust across EV analysis methods. We compared HaloTag-labeling to antibody-labeling of EVs using single vesicle flow cytometry, enabling us to quantify the substantial degree to which antibody labeling can underestimate the absolute number of proteins present on an EV. Finally, we demonstrate use of HaloTag to compare between protein designs for EV bioengineering. Overall, the HaloTag system is a useful EV characterization tool which complements and expands existing methods.

| S-EPMC10557717 | biostudies-literature

Particle segmentation algorithm for flexible single particle reconstruction.

Project description:As single particle cryo-electron microscopy has evolved to a new era of atomic resolution, sample heterogeneity still imposes a major limit to the resolution of many macromolecular complexes, especially those with continuous conformational flexibility. Here, we describe a particle segmentation algorithm towards solving structures of molecules composed of several parts that are relatively flexible with each other. In this algorithm, the different parts of a target molecule are segmented from raw images according to their alignment information obtained from a preliminary 3D reconstruction and are subjected to single particle processing in an iterative manner. This algorithm was tested on both simulated and experimental data and showed improvement of 3D reconstruction resolution of each segmented part of the molecule than that of the entire molecule.

| S-EPMC5515998 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data