Abnormal tau-immunoreactive filaments are a hallmark of tauopathies, including 
Alzheimer's disease (AD). A higher phosphorylation ("hyperphosphorylation") 
state of tau protein may represent a critical event. To determine the potential 
role of tau hyperphosphorylation in these disorders, mutated tau proteins were 
produced where serine/threonine residues known to be highly phosphorylated in 
tau filaments isolated from AD patients were substituted for glutamate to 
simulate a paired helical filament (PHF)-like tau hyperphosphorylation. We 
demonstrate that, like hyperphosphorylation, glutamate substitutions induce 
compact structure elements and SDS-resistant conformational domains in tau 
protein. Hyperphosphorylation-mimicking glutamate-mutated tau proteins display a 
complete functional loss in its ability to promote microtubule nucleation which 
can partially be overcome by addition of the osmolyte trimethylamine N-oxide 
(TMAO), which is similar to phosphorylated tau. In addition, glutamate-mutated 
tau proteins fail to interact with the dominant brain protein phosphatase 2A 
isoform ABalphaC, and exhibit a reduced ability to assemble into filaments. 
Interestingly, wild-type tau and phosphorylation-mimicking tau similarly bind to 
microtubules when added alone, but the mutated tau is almost completely 
displaced from the microtubule surface by equimolar concentrations of wild-type 
tau. The data indicate that glutamate-mutated tau proteins provide a useful 
model for analyzing the functional consequences of tau hyperphosphorylation. 
They suggest that several mechanisms contribute to the abnormal tau accumulation 
observed during tauopathies, in particular a selective displacement of 
hyperphosphorylated tau from microtubules, a functional loss in promoting 
microtubule nucleation, and a failure to interact with phosphatases.