Nanoparticles and ubiquitination as modulators of Tau aggregation and condensation.

Neurodegenerative diseases (NDs) are a heterogeneous group of neurolog-ical disorders characterised by a progressive loss of neurons in central nervous sys-tem (CNS) or peripheral nervous system (PNS). NDs are caused by misfolded pro-tein aggregates in specific areas of the brain. The collapse of the structure and func-tion of neural networks, results in the breakdown of the core communicative cir-cuitry, culminating in impaired memory, cognition, behaviour, sensory, and/or mo-toric functions. The prevalence of these diseases rises dramatically with age and is expected to surge with the increasing life expectancy in most countries, in the next decades. Among neurogenerative disorders, Alzheimer's disease (AD) and Parkinson’s dis-ease are the most common forms of dementia in older adults and the two relevant related proteins are Tau and α-synuclein, respectively. Both are amyloidogenic and natively unfolded proteins, which convert from monomeric to aggregated states in pathological conditions. The formation of Neurofibrillary tangles (NFTs) of Tau underlies the onset of the AD, as well as the aggregation of α-synuclein is associated with PD. The possibility to redirect aggregation pathway is essential to understand and pre-vent the doomed fate of these proteins. In this thesis I focused my attention on Tau protein, I investigated the molecular mechanisms of aggregation, and I exploited different ways for preventing or target-ing the aggregation pathway. A body of evidence shows that post-translational modifications (PTMs) can play important role in both physiological and pathological behaviour of Tau protein. Therefore, I focused my attention on the investigation of the impact of ubiquitina-tion of Tau on the conversion into toxic species. For this purpose, I optimized a novel method to insert ubiquitin protein onto a specific position of Tau, via dehy-droalanine chemistry. After optimizing the synthetic step, I investigated the aggre-gation propensity of the ubiquitinated Tau and interestingly I observed that this modification interferes with the fibrils formation. Moreover, I investigated the influence of nanomaterials, in particular gold nanopar-ticles, on Tau properties. The application on nanomaterials in routine life is growing day by day, due to their unique physical properties. In this work, I used ultrasmall gold nanoparticles (usGNPs), because the core size gives them extraordinary properties like fluores-cence and biocompatibility. So, I combined the nanomaterials with Tau studies, in two different directions. First of all, I tried to understand if nanomaterials can pre-vent or at least slow down Tau aggregation. As well as for ubiquitination studies, I monitored Tau aggregation in the presence of different usGNPs concentrations, and I discovered that usGNPs are able to reduce Tau aggregation, and, at a certain amount, they can even arrest the aggregates formation. On the other hand, liquid-liquid phase separation of protein into condensed drop-lets, is a common mechanism for eukaryotic cells to carry out physiological reac-tions but mounting evidence suggest that this compartmentalization can be the spark that triggers amyloid protein aggregation. From this premise, I tried to char-acterize the system Tau-usGNPs, and actually I found that usGNPs are able to per-turb Tau droplets formation, but more interestingly I realized that they are good markers candidates to monitor Tau aggregation event in LLPS formation. Finally, I investigated the influence on Tau aggregation mediated by an Italian ex-presso coffee mixture of trigonelline, theobromine, genistein, and caffeine. In fact, it is well established that coffee and coffee derived phenolic compounds have a role in neuroprotection against oxidative-stress and neuro-inflammation thanks to the ability to cross the Blood Brain Barrier.