Date of Award
Doctor of Philosophy (PhD)
Biochemistry & Molecular Biophysics
Tau is an intrinsically disordered protein found mainly in neurons, composed of four main domains, the N-terminal domain, the proline rich region, the microtubule binding region, and the C-terminal domain. Tau regulates the dynamic instability of microtubules by facilitating the polymerization of tubulin. Tau is also a key component of the aggregates found in several neurodegenerative diseases collectively known as tauopathies, the most prevalent of which is Alzheimer’s disease. In this disease pathology, normally soluble tau aggregates to form insoluble neurofibrillary tangles. The presence of aggregated tau spreads from one region of the brain to another suggesting that tau propagates from cell to cell. Understanding the mechanisms by which tau interacts with microtubules and the aggregation pathway leading to the spread of disease pathology is vital for diagnosis and treatment of tauopathies. In this work, we primarily use single molecule fluorescence techniques to study three mechanisms of tau. We studied the initiation of aggregation of tau in the presence of polyphosphates, a biologically relevant polyanionic molecule. We find that the proline rich region contains multiple binding sites and contributes to the three mechanisms by which polyphosphates can initiation aggregation; a change in conformation towards a more aggregation-prone conformation, charge-screening to increase local concentrations of tau, and intermolecular crosslinking. We also studied the propagation of monomer tau between neuronal cells. We find that the proline rich region of tau has a unique ability to diffuse passively through a lipid membrane, and that tau has higher affinity to lipid membranes with more ordered structure, higher cholesterol content mimicking lipid rafts. Lastly, we investigated the effects of the R5L mutation on tau’s binding to tubulin. We find that the mutation found on the N-terminal domain of the protein doesn’t affect binding affinity to microtubules or tubulin in high molarity buffers but has a higher affinity for tau in the 3R isoform of tau in a low salt buffer. Together, this provides us with an insight into the importance of the domain dependence of tau in its functions and dysfunctions, as well as an insight into tau’s preference for binding partners.
Wickramasinghe, Sanjula, "Single Molecule Approaches To Studying Tau Function And Dysfunction" (2022). Publicly Accessible Penn Dissertations. 5471.