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1.
The two characteristic neuropathological lesions of Alzheimer's disease are the neurofibrillary tangles and the senile plaques. Neurofibrillary tangles are made of abnormal filaments (PHF) accumulating in neurons and mainly composed of a modified form of the microtubule-associated protein tau (PHF-tau). Senile plaques are composed of a cluster of dystrophic neurites surrounding an extracellular deposit of amyloid fibers made of a 42 amino-acid peptide (beta-amyloid peptide). The abnormal filaments contain the complete sequences of the different tau isoforms. The PHF-tau proteins can be distinguished from the normal tau proteins by the presence of several phosphorylated sites. One of these sites is phosphorylated by a calcium-calmodulin-dependent kinase. The relationship between PHF-tau and the cytoskeletal pathology in Alzheimer's disease is further discussed.  相似文献   

2.
The most characteristic cellular change in Alzheimer's disease is the accumulation of aberrant filaments, the paired helical filaments (PHF), in the affected neurons. There is growing evidence from a number of laboratories that dementia correlates better with the accumulation of PHF than of the extracellular amyloid, the second major lesion of Alzheimer's disease. PHF are both morphologically and biochemically unlike any of the normal neurofibrils. The major polypeptides in isolated PHF are microtubule-associated protein tau. Tau in PHF is phosphorylated differently from tau in microtubules. This abnormal phosphorylation of tau in PHF occurs at several sites. The accumulation of abnormally phosphorylated tau in the affected neurons in Alzheimer's disease brain precedes both the formation and the ubiquitination of the neurofibrillary tangles. In Alzheimer's disease brain, tubulin is assembly competent, but the in vitro assembly of microtubules is not observed. In vitro, the phosphate groups in PHF are less accessible than those of tau to alkaline phosphatase. The in vitro dephosphorylated PHF polypeptides stimulate microtubule assembly from bovine tubulin. It is hypothesized that a defect in the protein phosphorylation/dephosphorylation system is one of the earliest events in the cytoskeletal pathology in Alzheimer's disease. Production of nonfunctional tau by its phosphorylation and its polymerization into PHF most probably contributes to a microtubule assembly defect, and consequently, to a compromise in both axoplasmic flow and neuronal function. Index Entries: Alzheimer's disease; mechanisms of neuronal degeneration; neurofibrillary changes; paired helical filaments: biochemistry; microtubule-associated protein tau; abnormal phosphorylation; ubiquitination; microtubule assembly; axoplasmic flow; protein phosphorylation/dephosphorylation.  相似文献   

3.
Paired helical filaments (PHF) are unusual neuronal fibers which accumulate progressively in the brain in Alzheimer's disease (AD). The insolubility of PHF in various kinds of solvents enabled us to obtain highly purified PHF, but prevented the application of conventional analytical methods to identify their components. Here we report that antibodies against purified PHF recognize tau protein, a brain-specific microtubule-associated protein, suggesting that a portion of PHF is tau protein.  相似文献   

4.
《The Journal of cell biology》1988,107(6):2703-2716
The paired helical filaments (PHFs) of Alzheimer's disease were purified by a strategy in which the neurons and amyloid plaque cores of protein (APCP) were initially isolated. This was achieved by several steps of isocratic sucrose centrifugations of increasing molarity and a discontinuous isotonic Percoll density gradient. After collagenase elimination of contaminating blood vessels, lysis of neurons was produced by SDS treatment. The released PHF cytoskeletons were separated from contaminating APCP and lipofuscin by sucrose density gradient. A final step consisted in the chemical purification of highly enriched PHFs and APCP components via a formic acid to guanidine hydrochloride transition. PHFs and APCPs were fractionated by size exclusion HPLC and further characterized and quantitated by automatic amino acid analysis. We also present some of the morphological and immunochemical characteristics of PHF polypeptides and APCP. Our studies indicate that apart from differences in localization and morphology, PHF and APCP significantly differ in (a) chemical structure (peptide and amino acid composition); (b) epitope specificity (antiubiquitin, antitau, antineurofilament); (c) physicochemical properties (structural conformation in guanidine hydrochloride); and (d) thioflavine T fluorescence emission. These parameters strongly suggest important differences in the composition and, probably, in the etiopathology of PHF and APCP of Alzheimer's disease.  相似文献   

5.
Several kinds of senile plaque found in 6 brains (4 from patients with Alzheimer's disease and 2 from patients with senile dementia) were examined in serial sections by light electron microscopy. The results obtained were as follows. All the senile plaques contained at least some amyloid fibrils, and these seemed to be produced at the basement membranes of capillary endothelial cells and projected into the surrounding parenchyma. Even when the senile plaques themselves appeared to lack amyloid fibrils by light microscopy, at least one degenerable capillary containing amyloid fibrils was demonstrable when serial sections were examined ultrastructurally. The findings described above suggest that the amyloid fibrils which form the cores of the several kinds of senile plaque, seem to be produced at the basement membrane of the endothelial cell. It is speculated that the capillary degeneration with the formation of amyloid fibrils may be primary change in the genesis of senile plaques.  相似文献   

6.
Alzheimer's disease (AD) is characterized by the deposition of amyloid plaques in the parenchyma and vasculature of the brain. Although previous analytical studies have provided much information about the composition and structure of synthetic amyloid-beta fibrils, there is, surprisingly, a dearth of data on intact amyloid plaques from AD brain. Therefore, to elucidate the structure and detailed composition of isolated amyloid plaque cores, we utilized a high-resolution, nondestructive technique, Raman microscopy. The data are of very high quality and contain detailed information about protein composition and conformation, about post-translational modification, and about the chemistry of metal binding sites. Remarkably, spectra obtained for senile plaque (SP) cores isolated from AD brain are essentially identical both within and among brains. The Raman data show for the first time that the SP cores are composed largely of amyloid-beta and confirm inferences from X-ray studies that the structure is beta-sheet with the additional possibility that this may be present as a parallel beta-helix. Raman bands characteristic of methionine sulfoxide show that extensive methionine oxidation has occurred in the intact plaques. The Raman spectra also demonstrate that Zn(II) and Cu(II) are coordinated to histidine residues in the SP cores, at the side chains' N(tau) and N(pi) atoms, respectively. Treatment of the senile plaques with the chelator ethylenediaminetetraacetate reverses Cu binding to SP histidines and leads to a broadening of amide features, indicating a "loosening" of the beta-structure. Our results indicate that Abeta in vivo is a metalloprotein, and the loosening of the structure following chelation treatment suggests a possible means for the solubilization of amyloid deposits. The results also reveal a direct chemical basis for oxidative damage caused by amyloid-beta protein in AD.  相似文献   

7.
The major constituent of Alzheimer's disease paired helical filaments (PHF) core is intrinsically disordered protein (IDP) tau. In spite of a considerable effort, insoluble character of PHF together with inherent physical properties of IDP tau have precluded so far reconstruction of PHF 3D structure by X-ray crystallography or NMR spectroscopy. Here we present first crystallographic study of PHF core C-terminus. Using monoclonal antibody MN423 specific to the tertiary structure of the PHF core, the in vivo PHF structure was imprinted into recombinant core PHF tau. Crystallization of the complex led to determination of the structure of the core PHF tau protein fragment 386TDHGAE391 at 1.65A resolution. Structural analysis suggests important role of the core PHF C-terminus for PHF assembly. It is reasonable to expect that this approach will help to reveal the structural principles underlying the tau protein assembly into PHF and possibly will facilitate rationale drug design for inhibition of Alzheimer neurofibrillary changes.  相似文献   

8.
9.
The purpose of the present study was to develop a purification and solubilization method, compatible with current amino acid sequencing techniques, for paired helical filaments (PHFs) derived from patients with Alzheimer's disease. We have developed a mild procedure that subjects conventionally isolated PHFs to Tris/borate/sodium dodecyl sulfate/2-mercaptoethanol electrophoresis and results in the separation of the relatively insoluble PHF structures from both copurifying contaminating proteins and solubilized PHF-associated proteins. At the end of 4.5 h of electrophoresis, the purified insoluble fraction had an amino acid composition that was invariant during subsequent electrophoresis. Electron microscopy revealed an intact PHF structure before and after electrophoresis but no evidence of any other structures in the insoluble fraction, a result consistent with the removal of PHF-associated proteins from the filament structure. Isolated insoluble filament structures displayed an enhanced immunoreactivity with antibodies raised against purified PHFs in other laboratories, when compared with the fraction not subjected to electrophoresis in enzyme-linked immunosorbent assays. Solubilization of the relatively insoluble PHFs was accomplished by extending the time of electrophoresis beyond the 4.5 h required for purification. Additional electrophoresis for 34.5 h solubilized 88% of the purified, relatively insoluble PHFs. This resulted in the identification of four major protein bands between Mr values of approximately 50,000 and 70,000 on sodium dodecyl sulfate-polyacrylamide electrophoresis gel analysis, with a predominant band with an Mr of approximately 66,000. A slow fragmentation of the PHF ultrastructure occurred during this time, as judged by electron microscopy. This purification technique will permit the isolation of consistently reproducible protein fragments from solubilized PHFs, which may be used for subsequent sequence analysis.  相似文献   

10.
Abnormal tau-containing filaments in neurodegenerative diseases   总被引:1,自引:0,他引:1  
It has been known for some time that the neurofibrillary pathology in Alzheimer's disease consists of so-called paired helical and straight filaments made up of the microtubule-associated protein tau. The degree of dementia observed in the disease correlates better with the extent of neurofibrillary pathology than with the Abeta amyloid deposits, the other characteristic defining pathological fibrous deposit in Alzheimer's disease. However, no familial cases of Alzheimer's disease have been genetically linked to the tau protein locus. Recently a group of frontotemporal dementias with parkinsonism linked to chromosome 17 has been shown to be caused by mutations in the tau gene. Some are missense mutations giving altered tau proteins, whereas others affect the splicing of the pre-mRNA and change the balance between different tau isoforms. Histologically these diseases are all characterised by various kinds of filamentous tau protein deposits, mostly in the complete absence of Abeta deposits. The abnormal tau filaments show different morphologies, depending on the nature of the tau mutation. These diseases show that tau mutations can be a prime cause of inherited dementing illness and may throw some light on the pathological process in the much larger number of sporadic cases of Alzheimer's disease.  相似文献   

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