An integrated strategy to correlate aggregation state, structure and toxicity of Aß 1–42 oligomers
| Autor: | Antonino Natalello, Raffaella Colombo, Federica Bisceglia, Laura Verga, Cristina Lanni, Ersilia De Lorenzi, Melania Maria Serafini |
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| Přispěvatelé: | Bisceglia, F, Natalello, A, Serafini, M, Colombo, R, Verga, L, Lanni, C, De Lorenzi, E |
| Rok vydání: | 2018 |
| Předmět: |
Protein Conformation
alpha-Helical 0301 basic medicine Cell Survival Amyloid beta Population Sample preparation FIS/07 - FISICA APPLICATA (A BENI CULTURALI AMBIENTALI BIOLOGIA E MEDICINA) Peptide A beta Fibril oligomer Phosphates Analytical Chemistry Turn (biochemistry) 03 medical and health sciences chemistry.chemical_compound Cell Line Tumor Spectroscopy Fourier Transform Infrared Aß 1–42 Humans Dimethyl Sulfoxide education Protein secondary structure chemistry.chemical_classification Aß oligomer education.field_of_study Amyloid beta-Peptides biology Fourier transform infrared spectroscopy Electrophoresis Capillary Alzheimer's disease Peptide Fragments Random coil Capillary electrophoresi 030104 developmental biology Monomer A beta 1-42 chemistry Solvents biology.protein Biophysics Protein Conformation beta-Strand Protein Multimerization |
| Zdroj: | Talanta. 188:17-26 |
| ISSN: | 0039-9140 |
| Popis: | Despite great efforts, it is not known which oligomeric population of amyloid beta (Aß) peptides is the main neurotoxic mediator in Alzheimer's disease. In vitro and in vivo experiments are challenging, mainly because of the high aggregation tendency of Aß (in particular of Aß 1–42 peptide), as well as because of the dynamic and non covalent nature of the prefibrillar aggregates. As a step forward in these studies, an analytical platform is here proposed for the identification and characterization of Aß 1–42 oligomeric populations resulting from three different sample preparation protocols. To preserve the transient nature of aggregates, capillary electrophoresis is employed for monitoring the oligomerization process in solution until fibril precipitation, which is probed by transmission electron microscopy. Based on characterization studies by ultrafiltration and SDS-PAGE/Western Blot, we find that low molecular weight oligomers build up over time and form bigger aggregates (> dodecamers) and that the kinetics strongly depends on sample preparations. The use of phosphate buffer results to be more aggregating, since trimers are the smallest species found in solution, whereas monomers and dimers are obtained by solubilizing Aß 1–42 in a basic mixture. For the first time, attenuated total reflection-Fourier transform infrared spectroscopy is used to assign secondary structure to the separated oligomers. Random coil and/or α-helix are most abundant in smaller species, whereas ß-sheet is the predominant conformation in bigger aggregates, which in turn are demonstrated to be responsible for Aß 1–42 toxicity. |
| Databáze: | OpenAIRE |
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