Following the intracellular repair of oxidation-damaged α-synuclein by time-resolved in-cell NMR spectroscopy

Intracellular aggregation of the intrinsically disordered human protein α-synuclein (α-Syn) is causally linked to Parkinson's disease (PD) and a relationship between ageing, increased cellular oxidative stress leading to selective cell death in response to α-Syn aggregation is firmly established. In support of this notion, aggregated α-Syn harbors many forms of oxidative damage in PD patients' brains, including, most prominently, oxidation of its methione residues.

We studied the fate of methionine-oxidized α-Syn in healthy neuronal and non-neuronal cells by high-resolution in-cell NMR spectroscopy. Specifically, we use protein electroporation to deliver pre-damaged α-Syn into different mammalian cell types and follow its intracellular behavior by time-resolved in-cell NMR experiments.

We found that intracellular repair of oxidation-damaged α-Syn proceeds in a selective and strictly stepwise manner with N-terminal methionines (Met5 and Met1) being repaired, while at C-terminal region the intracellular repair fails the target and Met116 and Met127 remains oxidized. In turn, we showed that the persistence of oxidative damage at C-terminal region modulates the phosphorylation and de-phosphorylation behavior of neighboring residues. Phospho-modifications of these residues constitute pathological hallmark signatures of aggregated α-Syn in PD patients. Thus, our results establish that oxidative cellular stress leads to the accumulation of chemically and functionally altered α-Syn in cells.

Antonio Limatola graduated from the University of Naples "Federico II", Italy (2011), where he also obtained his European Phd degree in Medicinal Chemistry (2015), working with Prof. Dr. Alfonso Carotenuto.

During his PhD studentship, he was guest at the Leibniz institut (2014) and, recently, he has been a guest at the Max-Planck Laboratory in Rosario, Argentina (2015-2016), under the cordial hospitality of Dr. Andrès Binolfi. Currently, he is a postdoc at Molecular Pharmacology division (FMP) of Leibniz institut in Berlin, Germany, under the supervision of Dr. Philipp Selenko.

His expertise involves the use of NMR techniques as methods for fragment screening and to guide interactive optimisations of ligand-macromolecule interactions.