title: Intracellular repair of oxidation-damaged α-synuclein fails to target C-terminal modification sites.
abstract: Cellular oxidative stress serves as a common denominator in many neurodegenerative disorders, including Parkinson's disease. Here we use in-cell NMR spectroscopy to study the fate of the oxidation-damaged Parkinson's disease protein alpha-synuclein (α-Syn) in non-neuronal and neuronal mammalian cells. Specifically, we deliver methionine-oxidized, isotope-enriched α-Syn into cultured cells and follow intracellular protein repair by endogenous enzymes at atomic resolution. We show that N-terminal α-Syn methionines Met1 and Met5 are processed in a stepwise manner, with Met5 being exclusively repaired before Met1. By contrast, C-terminal methionines Met116 and Met127 remain oxidized and are not targeted by cellular enzymes. In turn, persisting oxidative damage in the C-terminus of α-Syn diminishes phosphorylation of Tyr125 by Fyn kinase, which ablates the necessary priming event for Ser129 modification by CK1. These results establish that oxidative stress can lead to the accumulation of chemically and functionally altered α-Syn in cells.
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Andres
authors.initials: A
authors.lastname: Binolfi
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Antonio
authors.initials: A
authors.lastname: Limatola
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Silvia
authors.initials: S
authors.lastname: Verzini
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Jonas
authors.initials: J
authors.lastname: Kosten
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Francois-Xavier
authors.initials: FX
authors.lastname: Theillet
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Honor May
authors.initials: HM
authors.lastname: Rose
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Beata
authors.initials: B
authors.lastname: Bekei
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Marchel
authors.initials: M
authors.lastname: Stuiver
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Marleen
authors.initials: M
authors.lastname: van Rossum
authors.affiliation: Department of NMR-supported Structural Biology, In-Cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Robert-Roessle Strasse 10, Berlin 13125, Germany.
authors.forename: Philipp
authors.initials: P
authors.lastname: Selenko
mesh_terms: D020816:Amino Acid Motifs; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008715:Methionine / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010300:Parkinson Disease / Q000378:metabolism*; D010766:Phosphorylation; D012694:Serine / Q000378:metabolism; D051844:alpha-Synuclein / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D051844:alpha-Synuclein; D012694:Serine; D008715:Methionine