Chemical Crosslinking of Isotopically Labeled α-Synuclein Reveals Structural Changes and the Interface of Oligomeric Species

ABSTRACT: Due to its high pharmaceutical relevance, α-synuclein is one of the best studied intrinsically disordered proteins to date. Its structural plasticity reaches from α-helical when bound to membranes through disordered in solution to β-sheet when forming amyloid fibrils. Recently the possible toxicity of membrane bound α-synuclein oligomers has elicited increased interest in the understanding of its membrane bound state. Both the high structural variability in the bound state and the requirement of membranes to elicit structural conversion of α-synuclein into is oligomeric membrane bound state have impeded the determination of a high-resolution oligomeric membrane-bound structure. The use of chemical crosslinking mass spectrometry (XL-MS) in this context leads to a very large set of peptide spectrum matches (PSMs), which do not permit the elucidation of a single, well defined structure based on XL-MS alone. The pattern of PSMs obtained does, however, represent possible links within the ensemble of membrane bound α-synuclein states. Here, we use fully 15N isotopically labeled α-synuclein in 1:1 mixtures with the natural abundance (14N) protein in order to allow distinction between intermolecular and intramolecular crosslinks in α-synuclein oligomers. Information obtained on the main crosslinking regions as well as changes in the observed PSM patterns is then used in conjunction with data obtained from nuclear magnetic resonance (NMR) measurements to develop high-resolution structural models of α-synuclein. We expect that the approach taken here will also prove useful in other highly flexible oligomeric systems.

INSTRUMENT(S): Orbitrap Fusion Lumos, Q Exactive HF-X

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Markus Hartl  

LAB HEAD: Robert Konrat

PROVIDER: PXD027349 | Pride | 2023-04-07

REPOSITORIES: Pride