Preparation of Alzheimer's-disease-relevant Tau protein in HEK293 mammalian cells
| Název česky | Příprava proteinu Tau relevantního pro Alzheimerovu chorobu v savčích HEK293 buňkách |
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| Autoři | |
| Rok publikování | 2024 |
| Druh | Konferenční abstrakty |
| Fakulta / Pracoviště MU | |
| Citace | |
| Popis | Tau protein is a microtubule-binding protein that is usually post-translationally modified (PTM). Hyperphosphorylation and abnormal PTMs of Tau have been identified as key determinants connecting Tau and Alzheimer’s disease. After such abnormal modification, Tau detaches from microtubules and aggregates into neurotoxic neurofibrillary tangles. However, the molecular mechanism and exact factors causing Tau aggregation are so far not well understood. Here, we aimed to prepare Tau protein in mammalian HEK293 cells to obtain Tau bearing natural PTMs. Tau protein was successfully expressed, purified from the cell lysate, and its purity and identity were verified using SDS-PAGE, Western blot, and mass spectrometry (MS). Currently, the presence of PTMs within Tau protein is inspected by advanced MS analysis. Next step will be to study the effects of the PTMs on Tau structural, interaction, and aggregation properties in comparison to un-modified Tau expressed in E. coli or in vitro phosphorylated Tau (already published in article by Crha R, Kozeleková A, Hofrová A, Iľkovičová L, Gašparik N, Kadeřávek P, Hritz J. Hiding in plain sight: Complex interaction patterns between Tau and 14-3-3? protein variants. Int J Biol Macromol. 2024, doi: 10.1016/j.ijbiomac.2024.130802). The majority of results were obtained within Industrial PhD programme in biotech company BioVendor. Project acknowledgement: This project has received funding from the European Union’s Horizon Europe program under the grant agreement No. 101087124. We acknowledge CEITEC Proteomics Core Facility of CIISB, Instruct-CZ Centre, supported by MEYS CR (LM2023042, e-INFRA CZ (ID:90254)). |
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