Role of cysteine-291 and cysteine-322 in the polymerization of human tau into Alzheimer-like filaments.
Filamentous tau pathology is central to a large number of dementing disorders, including Alzheimer's disease in which polymerized tau is hyperphosphorylated. Previous studies on heparin-dependent tau polymerization, using recombinant tau isoforms lacking Cys-291, suggest that tau dimerization via Cys-322 is critical for initiation of assembly of soluble tau into filaments. We report heparin-dependent in vitro polymerization of human recombinant tau (1-383 isoform), containing both Cys-291 and Cys-322, into paired helical filaments as characterized by electron microscopy. Tau polymerization, under physiological tau concentrations in the presence of dithiothreitol (DTT), was followed by a Thioflavine S fluorescence assay. To understand the molecular basis for heparin-induced tau polymerization, we expressed and purified C291A, C322A, and C291A/C322A tau mutants. The DTT requirement for tau polymerization was abolished using either the C291A or C322A tau mutant and polymerization was not observed with the C291A/C322A tau double mutant. Analysis by sodium dodecyl sulfate gel electrophoresis showed that, unlike wild type tau, a significant amount of the C291A mutant and the C322A mutant is present as a disulfide bonded dimer. Taken together these results suggest that, in isoforms containing both Cys-291 and Cys-322, a dimeric tau with an intermolecular disulfide bond through either Cys-291 or Cys-322 is presumably acting as a seed for initiation of tau polymerization.
Bhattacharya
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Pharmacia Corporation, Kalamazoo, Michigan 49007, USA.
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D000544:Alzheimer Disease / Q000378:metabolism*; D001704:Biopolymers / Q000378:metabolism*; D003545:Cysteine / Q000378:metabolism*; D004229:Dithiothreitol / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008854:Microscopy, Electron; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D016875:tau Proteins / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure
D001704:Biopolymers; D020033:Protein Isoforms; D016875:tau Proteins; D003545:Cysteine; D004229:Dithiothreitol