Site-Specific Nitration Differentially Influences τ Assembly in Vitro
- 1 October 2005
- journal article
- research article
- Published by American Chemical Society (ACS) in Biochemistry
- Vol. 44 (42), 13997-14009
- https://doi.org/10.1021/bi051028w
Abstract
Previously, we reported that the microtubule-associated τ protein, the major constituent of neurofibrillary tangles (NFTs) in Alzheimer's brain, undergoes site-selective nitration by peroxynitrite (ONOO-) and that this event inhibits τ polymerization in vitro [Reynolds et al. (2005) Biochemistry 44, 1690−1700]. In the present study, we extend our analysis of τ nitration to include mutant τ proteins singly nitrated at each residue targeted by ONOO- in vitro (Tyr18, Tyr29, Tyr197, and Tyr394). Using our polymerization paradigm, we demonstrate that site-specific Tyr nitration differentially alters the rate and/or extent of τ assembly and generates robust changes in filament morphology. As determined by quantitative electron microscopy, select nitration of residues Tyr29 and Tyr197 increases the average length of synthetic τ filaments but does not alter the steady-state polymer mass. In contrast, site-specific nitration of residues Tyr18 and Tyr394 decreases the average length and/or number of synthetic filaments, resulting in a significant reduction in filamentous mass and an increase in τ critical concentration. Intriguingly, affinity measurements demonstrate that nitrative modifications do not preclude formation of the Alz-50 epitope, a pathological τ conformation detectable in authentic paired helical filaments (PHFτ). In fact, the Alz-50 antibody binds filaments assembled from nitrated mutant τ with higher avidity than wild-type filaments, even in instances where the overall filamentous mass is reduced. Taken together, our results suggest that site-specific nitration modulates the nucleation and/or elongation capacity of assembly-competent τ and that assumption of the Alz-50 conformation may be necessary, but not sufficient, to induce filament formation.Keywords
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