The microtubule-associated protein tau forms disease-specific filamentous aggregates in several different neurodegenerative diseases. In order to understand how tau undergoes misfolding into a specific filament type and to control this process for drug development purposes, it is crucial to study in vitro tau aggregation methods and investigate the structures of the obtained filaments at the atomic level. Here, we used the tau fragment dGAE, which aggregates spontaneously, to seed the formation of full-length tau filaments. The structures of dGAE and full-length tau filaments were investigated by magic-angle spinning (MAS) solid-state NMR, showing that dGAE allows propagation of a chronic traumatic encephalopathy (CTE)-like fold to the full-length tau. The obtained filaments efficiently seeded tau aggregation in HEK293T cells. This work demonstrates that in vitro preparation of disease-specific types of full-length tau filaments is feasible.

• Klíčová slova
• NMR spectroscopy, dGAE fragment, filamentous aggregates, tau protein, tauopathies,
• MeSH
• chronická traumatická encefalopatie * metabolismus   patologie   MeSH
• HEK293 buňky MeSH
• lidé MeSH
• proteiny tau * metabolismus   chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• proteiny tau * MeSH

Tau protein is an intrinsically disordered protein that plays a key role in Alzheimer's disease (AD). In brains of AD patients, Tau occurs abnormally phosphorylated and aggregated in neurofibrillary tangles (NFTs). Together with Tau, 14-3-3 proteins - abundant cytosolic dimeric proteins - were found colocalized in the NFTs. However, so far, the molecular mechanism of the process leading to pathological changes in Tau structure as well as the direct involvement of 14-3-3 proteins are not well understood. Here, we aimed to reveal the effects of phosphorylation by protein kinase A (PKA) on Tau structural preferences and provide better insight into the interaction between Tau and 14-3-3 proteins. We also addressed the impact of monomerization-inducing phosphorylation of 14-3-3 at S58 on the binding to Tau protein. Using multidimensional nuclear magnetic resonance spectroscopy (NMR), chemical cross-linking analyzed by mass spectrometry (MS) and PAGE, we unveiled differences in their binding affinity, stoichiometry, and interfaces with single-residue resolution. We revealed that the interaction between 14-3-3 and Tau proteins is mediated not only via the 14-3-3 amphipathic binding grooves, but also via less specific interactions with 14-3-3 protein surface and, in the case of monomeric 14-3-3, also partially via the exposed dimeric interface. In addition, the hyperphosphorylation of Tau changes its affinity to 14-3-3 proteins. In conclusion, we propose quite complex interaction mode between the Tau and 14-3-3 proteins.

• Klíčová slova
• 14-3-3ζ protein, Cross-linking, Interaction, NMR, Phosphorylation, Tau protein,
• MeSH
• Alzheimerova nemoc metabolismus   MeSH
• fosforylace MeSH
• lidé MeSH
• molekulární modely MeSH
• multimerizace proteinu MeSH
• proteinkinasy závislé na cyklickém AMP metabolismus   MeSH
• proteiny 14-3-3 * metabolismus   chemie   MeSH
• proteiny tau * metabolismus   chemie   MeSH
• vazba proteinů * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• proteinkinasy závislé na cyklickém AMP MeSH
• proteiny 14-3-3 * MeSH
• proteiny tau * MeSH

Protein-protein interactions (PPIs) remain poorly explored targets for the treatment of Alzheimer's disease. The interaction of 14-3-3 proteins with Tau was shown to be linked to Tau pathology. This PPI is therefore seen as a potential target for Alzheimer's disease. When Tau is phosphorylated by PKA (Tau-PKA), several phosphorylation sites are generated, including two known 14-3-3 binding sites, surrounding the phosphorylated serines 214 and 324 of Tau. The crystal structures of 14-3-3 in complex with peptides surrounding these Tau phosphosites show that both these motifs are anchored in the amphipathic binding groove of 14-3-3. However, in the absence of structural data with the full-length Tau protein, the stoichiometry of the complex or the interface and affinity of the partners is still unclear. In this work, we addressed these points, using a broad range of biophysical techniques. The interaction of the long and disordered Tau-PKA protein with 14-3-3σ is restricted to two short sequences, containing phosphorylated serines, which bind in the amphipathic binding groove of 14-3-3σ. Phosphorylation of Tau is fundamental for the formation of this stable complex, and the affinity of the Tau-PKA/14-3-3σ interaction is in the 1-10 micromolar range. Each monomer of the 14-3-3σ dimer binds one of two different phosphorylated peptides of Tau-PKA, suggesting a 14-3-3/Tau-PKA stoichiometry of 2 : 1, confirmed by analytical ultracentrifugation. These results contribute to a better understanding of this PPI and provide useful insights for drug discovery projects aiming at the modulation of this interaction.

• Klíčová slova
• 14-3-3 proteins, Alzheimer’s disease, NMR spectroscopy, Tau protein, analytical ultracentrifugation, protein-protein interactions,
• MeSH
• Alzheimerova nemoc genetika   metabolismus   patologie   MeSH
• exoribonukleasy chemie   genetika   metabolismus   MeSH
• fosforylace MeSH
• konformace proteinů MeSH
• lidé MeSH
• magnetická rezonanční spektroskopie MeSH
• multimerizace proteinu * MeSH
• mutace MeSH
• povrchová plasmonová rezonance MeSH
• proteinkinasy závislé na cyklickém AMP metabolismus   MeSH
• proteiny 14-3-3 chemie   genetika   metabolismus   MeSH
• proteiny tau chemie   genetika   metabolismus   MeSH
• serin chemie   metabolismus   MeSH
• vazba proteinů MeSH
• vazebná místa MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• exoribonukleasy MeSH
• proteinkinasy závislé na cyklickém AMP MeSH
• proteiny 14-3-3 MeSH
• proteiny tau MeSH
• serin MeSH
• SFN protein, human MeSH Prohlížeč

In Alzheimer's disease, tau protein undergoes post-translational modifications including hyperphosphorylation and truncation, which promotes two major conformational changes associated with progressive N-terminal folding. Along with the development of the disease, tau ubiquitination was previously shown to emerge in the early and intermediate stages of the disease, which is closely associated with early tau truncation at aspartic acid 421, but not with a subsequently truncated tau molecule at glutamic acid 391. In the same group of cases, using multiple immunolabeling and confocal microscopy, a possible relationship between the ubiquitin-targeting of tau and the progression of conformational changes adopted by the N-terminus of this molecule was further studied. A comparable number of neurofibrillary tangles was found displaying ubiquitin, an early conformation recognized by the Alz-50 antibody, and a phosphorylation. However, a more reduced number of neurofibrillary tangles were immunoreactive to Tau-66 antibody, a late tau conformational change marker. When double-labeling profiles of neurofibrillary tangles were assessed, ubiquitination was clearly demonstrated in tau molecules undergoing early N-terminal folding, but was barely observed in late conformational changes of the N-terminus adopted by tau. The same pattern of colocalization was visualized in neuritic pathology. Overall, these results indicate that a more intact conformation of the N-terminus of tau may facilitate tau ubiquitination, but this modification may not occur in a late truncated and more compressed folding of the N-terminus of the tau molecule.

• Klíčová slova
• Alzheimer’s disease, Neuritic pathology, Neurofibrillary tangles, Tau conformational changes, Ubiquitination,
• MeSH
• Alzheimerova nemoc metabolismus   patologie   MeSH
• konformace proteinů MeSH
• lidé středního věku MeSH
• lidé MeSH
• mozek metabolismus   patologie   MeSH
• neurofibrilární klubka chemie   patologie   MeSH
• proteiny tau chemie   metabolismus   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ubikvitinace fyziologie   MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• MAPT protein, human MeSH Prohlížeč
• proteiny tau MeSH

Alzheimer's disease (AD) is a neurodegenerative disease that is usually accompanied by aging, increasingly being the most common cause of dementia in the elderly. This disorder is characterized by the accumulation of beta amyloid plaques (Aβ) resulting from impaired amyloid precursor protein (APP) metabolism, together with the formation of neurofibrillary tangles and tau protein hyperphosphorylation. The exacerbated production of reactive oxygen species (ROS) triggers the process called oxidative stress, which increases neuronal cell abnormalities, most often followed by apoptosis, leading to cognitive dysfunction and dementia. In this context, the development of new therapies for the AD treatment is necessary. Antioxidants, for instance, are promising species for prevention and treatment because they are capable of disrupting the radical chain reaction, reducing the production of ROS. These species have also proven to be adjunctive to conventional treatments making them more effective. In this sense, several recently published works have focused their attention on oxidative stress and antioxidant species. Therefore, this review seeks to show the most relevant findings of these studies.

• Klíčová slova
• Alzheimer’s disease, antioxidants, cellular respiration, free radicals, oxidative stress,
• MeSH
• Alzheimerova nemoc farmakoterapie   metabolismus   MeSH
• amyloidní beta-protein chemie   metabolismus   MeSH
• antioxidancia farmakologie   terapeutické užití   MeSH
• fosforylace MeSH
• klinické zkoušky jako téma MeSH
• lidé MeSH
• oxidační stres účinky léků   MeSH
• proteiny tau chemie   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• amyloidní beta-protein MeSH
• antioxidancia MeSH
• proteiny tau MeSH

During the last two decades, neuropathological examination of the brain has evolved both technically and scientifically. The increasing use of immunohistochemistry to detect protein aggregates paralleled a better understanding of neuroanatomical progression of protein deposition. As a consequence, an international effort was achieved to standardize hyperphosphorylated-Tau (phospho-TAU), ßAmyloid (Aß), alpha syncuclein (alpha-syn), phosphorylated transactive response DNA-binding protein 43 (phospho-TDP43) and vascular pathology detection. Meanwhile harmonized staging systems emerged in order to increase inter rater reproducibility. Therefore, a refined definition of Alzheimer's disease was recommended., a clearer picture of the neuropathological lesions diversity emerged secondarily to the systematic assessment of concomitant pathology highlighting finally a low rate of pure AD pathology. This brings new challenges to laboratory medicine in the field of cerebrospinal fluid (CSF) markers of Alzheimer's disease: how to further validate total Tau, phospho-TAU, Aß40 and Aß42 and new marker level cut-offs while autopsy rates are declining?

• MeSH
• Alzheimerova nemoc mozkomíšní mok   diagnóza   patologie   MeSH
• amyloidní beta-protein mozkomíšní mok   normy   MeSH
• biologické markery mozkomíšní mok   chemie   MeSH
• fosforylace MeSH
• lidé MeSH
• mozek patologie   MeSH
• progrese nemoci MeSH
• proteiny tau mozkomíšní mok   chemie   normy   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• amyloidní beta-protein MeSH
• biologické markery MeSH
• proteiny tau MeSH

The stability and dynamics of cytoskeleton in brain nerve cells are regulated by microtubule associated proteins (MAPs), tau and MAP2. Both proteins are intrinsically disordered and involved in multiple molecular interactions important for normal physiology and pathology of chronic neurodegenerative diseases. Nuclear magnetic resonance and cryo-electron microscopy recently revealed propensities of MAPs to form transient local structures and long-range contacts in the free state, and conformations adopted in complexes with microtubules and filamentous actin, as well as in pathological aggregates. In this paper, we compare the longest, 441-residue brain isoform of tau (tau40), and a 467-residue isoform of MAP2, known as MAP2c. For both molecules, we present transient structural motifs revealed by conformational analysis of experimental data obtained for free soluble forms of the proteins. We show that many of the short sequence motifs that exhibit transient structural features are linked to functional properties, manifested by specific interactions. The transient structural motifs can be therefore classified as molecular recognition elements of tau40 and MAP2c. Their interactions are further regulated by post-translational modifications, in particular phosphorylation. The structure-function analysis also explains differences between biological activities of tau40 and MAP2c.

• Klíčová slova
• intrinsically disordered protein, microtubule associated protein, nuclear magnetic resonance, phosphorylation, tau,
• MeSH
• lidé MeSH
• mikrotubuly chemie   metabolismus   MeSH
• proteiny asociované s mikrotubuly chemie   metabolismus   MeSH
• proteiny tau chemie   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• proteiny asociované s mikrotubuly MeSH
• proteiny tau MeSH

The presence of pre-existing natural antibodies against Alzheimer's disease (AD) pathological proteins might interfere with immune responses to therapeutic vaccination with these proteins. We aimed to compare levels of antibodies in CSF and serum: We observed higher reactivity of natural tau-reactive antibodies towards phosphorylated bovine tau protein than to human recombinant (non-phosphorylated) tau protein. Males with MCI-AD had higher amounts of these antibodies than corresponding controls. Concentrations of antibodies were lower in females with the MCI-AD than in control females. These findings may have implications for tau vaccination trials.

• Klíčová slova
• Alzheimer's disease, Antibody, Mild cognitive impairment, Post-translational modification, Sex, Tau protein,
• MeSH
• Alzheimerova nemoc krev   mozkomíšní mok   imunologie   MeSH
• autoprotilátky analýza   krev   mozkomíšní mok   MeSH
• demence krev   mozkomíšní mok   imunologie   MeSH
• druhová specificita MeSH
• fosfoproteiny chemie   imunologie   MeSH
• fosforylace MeSH
• kognitivní dysfunkce krev   mozkomíšní mok   imunologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• pohlavní dimorfismus * MeSH
• posttranslační úpravy proteinů MeSH
• proteiny tau chemie   imunologie   MeSH
• rekombinantní proteiny imunologie   MeSH
• reprodukovatelnost výsledků MeSH
• senioři MeSH
• skot MeSH
• specificita protilátek MeSH
• zvířata MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• skot MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• autoprotilátky MeSH
• fosfoproteiny MeSH
• MAPT protein, human MeSH Prohlížeč
• proteiny tau MeSH
• rekombinantní proteiny MeSH

Post-translational modifications, including phosphorylation, greatly impact the physiological function of proteins, especially those that are natively unfolded and implicated in many neurodegenerative diseases. However, structural and functional studies of such proteins require fully defined phosphorylation, including those that are not physiological. Thus, the kinases ERK2 and GSK-3β were immobilized to various superparamagnetic beads with carboxylic, aldehyde, Ni2+, or Co3+ functional groups, with a view to efficiently phosphorylate peptides and proteins in vitro. Full phosphorylation of specific synthetic peptides confirmed that beads were successfully loaded with kinases. Remarkably, enzymes covalently immobilized on carboxylated SeraMag beads remained active upon reuse, with residual activity after 10 uses 99.5 ± 0.34% for GSK-3β and 36.2 ± 2.01% for ERK2. The beads were also used to sequentially phosphorylate recombinant tau, which in vivo is a biomarker of Alzheimer's disease. Thus, a system consisting of two fully active kinases immobilized to magnetic beads is demonstrated for the first time. In comparison to soluble enzymes, the beads are easier to handle, reusable, and thus low-cost. Importantly, these beads are also convenient to remove from reactions to minimize contamination of phosphorylated products or to exchange with other kinases.

• MeSH
• enzymy imobilizované MeSH
• fosforylace * MeSH
• kinasa glykogensynthasy 3beta metabolismus   MeSH
• králíci MeSH
• lidé MeSH
• magnetismus MeSH
• mikrosféry MeSH
• mitogenem aktivovaná proteinkinasa 1 metabolismus   MeSH
• peptidy chemie   MeSH
• proteiny tau chemie   MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• enzymy imobilizované MeSH
• kinasa glykogensynthasy 3beta MeSH
• mitogenem aktivovaná proteinkinasa 1 MeSH
• peptidy MeSH
• proteiny tau MeSH

Although tau protein was long regarded as an intracellular protein with several functions inside the cell, new evidence has shown tau secretion into the extracellular space. The active secretion of tau could be a physiological response of neurons to increased intracellular amounts of tau during the progression of tau pathology. We looked for potential differences in the serum levels of toxic tau oligomers in regards to cognitive impairment of subjects. We detected tau oligomers in the serum of Alzheimer's disease (AD) patients, but they were also present to some extent in the serum of healthy older subjects where the levels positively correlated with aging (Spearman r = 0.26, p = 0.016). On the contrary, we found lower levels of tau oligomers in the serum of mild cognitive impairment (MCI) (p = 0.033) and MCI-AD (p = 0.006) patients. These results could suggest that clearance of extracellular tau proteins takes place, in part, in the periphery. In the case of MCI patients, the lower levels of tau oligomers could be the result of impaired clearance of tau protein from interstitium to blood and consequent accumulation of tau aggregates in the brain.

• Klíčová slova
• Alzheimer’s disease, oligomers, serum, tau protein,
• MeSH
• Alzheimerova nemoc krev   mozkomíšní mok   komplikace   MeSH
• amyloidní beta-protein krev   mozkomíšní mok   MeSH
• ELISA MeSH
• kognitivní dysfunkce krev   etiologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• peptidové fragmenty krev   MeSH
• proteiny tau krev   mozkomíšní mok   chemie   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• stárnutí krev   MeSH
• záznam o duševním stavu MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• amyloidní beta-protein MeSH
• peptidové fragmenty MeSH
• proteiny tau MeSH