Quantitative solubilization and analysis of insoluble paired helical filaments from alzheimer disease

In this study, we evaluate the ability of several solvents to solubilize insoluble paired helical filaments (PHF) of Alzheimer disease. Specifically, we use protein extraction and reduction in the volume of insoluble material as quantitative assays to establish solvents of PHF. Using sequential cate...

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Bibliographic Details
Published in:Brain research 1996-04, Vol.717 (1), p.99-108
Main Authors: Smith, Mark A., Siedlak, Sandra L., Richey, Peggy L., Nagaraj, Ramanakoppa H., Elhammer, Ake, Perry, George
Format: Article
Language:English
Subjects:
Adolescent
Adult
Aged
Aged, 80 and over
Alkalies
Alzheimer disease
Alzheimer Disease - metabolism
Biological and medical sciences
Cell Fractionation
Cross-Linking Reagents - analysis
Cross-Linking Reagents - metabolism
Crosslink
Degenerative and inherited degenerative diseases of the nervous system. Leukodystrophies. Prion diseases
Densitometry
Endopeptidases
Formates
Glycation
Glycation End Products, Advanced - analysis
Guanidine
Guanidines
Hemostatics
Humans
Immunoblotting
Medical sciences
Microscopy, Electron
Middle Aged
Neurofibrillary pathology
Neurofilament Proteins - analysis
Neurofilament Proteins - metabolism
Neurofilament Proteins - ultrastructure
Neurology
Paired helical filament
Phosphorylation
Protein Denaturation
Sodium Dodecyl Sulfate
Solubility
Urea
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Summary:In this study, we evaluate the ability of several solvents to solubilize insoluble paired helical filaments (PHF) of Alzheimer disease. Specifically, we use protein extraction and reduction in the volume of insoluble material as quantitative assays to establish solvents of PHF. Using sequential categories of protein solvent to analyze insoluble PHF, only alkali or exhaustive proteolysis are effective in completely solubilizing PHF, while a variety of denaturants are ineffective. Alkali does not affect the phosphorylation state of PHF and complete dephosphorylation of PHF with hydrofluoric acid does not affect PHF solubility. These findings suggest that the ‘hyperphos-sphorylation’ of PHF proteins is not responsible for PHF insolubility. However the in vitro glycation of τ generates PHF that are insoluble in SDS and soluble in alkali. These findings suggest that protein crosslinks, including advanced glycation endproduct-derived crosslinks which were recently described in Alzheimer disease, play a major role in effecting PHF insolubility in vivo.
ISSN:0006-8993
1872-6240
DOI:10.1016/0006-8993(95)01473-X