Population of Tau Embedded in Parallel β-Sheets Depends on Heparin Stoichiometry

The aggregation of the human tau protein is heavily implicated in the progression of Alzhiemer’s Disease. In this study we examined the aggregation kinetics of a truncated segment of the longest tau isoform, Tau 187, and induced aggregation with the polyanion hepain. We applied cw-EPR in order to quantitatively measure the population of tau embedded in parallel β-sheet structure and compared these measurements to traditional kinetic measurement techniques, Tht fluorescence and turbidity. Cw-EPR revealed rapid formation of parallel β-sheet structure early in the aggregation process well before fibrils are thought to appear. We observe a large population of spins first exhibits reduced mobility upon mixing with heparin, which is later followed by an increase in β-sheet structure consistent with proposed models of nucleation followed by structural rearrangement. Additionally, comparing cw-EPR with Tht fluorescence demonstrates Tht binding is non-linear with respect to parallel β-sheet structure and the precise binding structure of Tht remains unclear. The transfer of tau pathology from infected to healthy cells has been well established by the literature, however, little is known about the relationship between seed structure and seeding efficacy. Here we investigated the importance of β-sheet content as detected by cw-EPR on seeding efficacy, but were unable to establish a relationship due to the overriding effect of heparin. Previously our group has observed a conformational shift around the PHF6* hexapeptide from compact to extended state and we examined the stoichiometric effect of heparin on this extension to determine the potency of heparin for inducing changes in the tau system. Finally, dependence of tau aggregation kinetics on heparin stoichiometry was investigated. We discovered the extent of observable distance change depended on heparin stoichiometry, however, even dilute quantities of heparin are able to induce significant extension around the PHF6* hexapeptide. The amount of total fibril formation was also found to depend on the heparin stoichiometry with lower heparin concentrations generating less total fibril content. Our findings suggests even minute quantities of heparin exerts great influence on the tau system and future work will require ways of working around or limiting heparin’s influence in order to directly probe the underlying fundamentals of tau aggregation