Native metastable prefibrillar oligomers are the most neurotoxic species among amyloid aggregates
| Autor: | Tamara C. Petrucci, Claudio Frank, Gianfranco Macchia, Maria Cristina Gaudiano, Fiorella Malchiodi-Albedi, Marco Diociaiuti, Pietro Chistolini, Serena Camerini, Silvia Paradisi |
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| Rok vydání: | 2014 |
| Předmět: |
Circular dichroism
Amyloid Photochemistry Size-exclusion chromatography Population Fibril Oligomer Hippocampus chemistry.chemical_compound Microscopy Electron Transmission Native state Neurotoxicity Animals Structural biophysics education Molecular Biology Cells Cultured education.field_of_study Amyloid oligomers Circular Dichroism Neurodegenerative diseases Brain Rats Electrophysiology Crystallography Monomer Cross-Linking Reagents chemistry Spectrometry Mass Matrix-Assisted Laser Desorption-Ionization Biophysics Chromatography Gel Molecular Medicine Calcium Dimerization |
| Zdroj: | Biochimica et Biophysica Acta (BBA) - Molecular Basis of Disease. 1842(9):1622-1629 |
| ISSN: | 0925-4439 |
| DOI: | 10.1016/j.bbadis.2014.06.006 |
| Popis: | Many proteins belonging to the amyloid family share the tendency to misfold and aggregate following common steps, and display similar neurotoxicity. In the aggregation pathway different kinds of species are formed, including several types of oligomers and eventually mature fibers. It is now suggested that the pathogenic aggregates are not the mature fibrils, but the intermediate, soluble oligomers. Many kinds of aggregates have been described to exist in a metastable state and in equilibrium with monomers. Up to now it is not clear whether a specific structure is at the basis of the neurotoxicity. Here we characterized, starting from the early aggregation stages, the oligomer populations formed by an amyloid protein, salmon calcitonin (sCT), chosen due to its very slow aggregation rate. To prepare different oligomer populations and characterize them by means of photoinduced cross-linking SDS-PAGE, Energy Filtered-Transmission Electron Microscopy (EF-TEM) and Circular Dichroism (CD) spectroscopy, we used Size Exclusion Chromatography (SEC), a technique that does not influence the aggregation process leaving the protein in the native state. Taking advantage of sCT low aggregation rate, we characterized the neurotoxic potential of the SEC-separated, non-crosslinked fractions in cultured primary hippocampal neurons, analyzing intracellular Ca2+ influx and apoptotic trend. We provide evidence that native, globular, metastable, prefibrillar oligomers (dimers, trimers and tetramers) were the toxic species and that low concentrations of these aggregates in the population was sufficient to render the sample neurotoxic. Monomers and other kind of aggregates, such as annular or linear protofibers and mature fibers, were totally biologically inactive. |
| Databáze: | OpenAIRE |
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