Fibrillation of alpha synuclein probed by surface methylation

RIAL HAWILA, MR; GÓMEZ G.E; BINOLFI A; DELFINO J.M

VIRTUAL

Congreso; 20th Congress of the International Union for Pure and Applied Biophysics (IUPAB), 50th Annual Meeting of the Brazilian Society for Biochemistry and Molecular Biology (SBBq) and 45th Congress of the Brazilian Society of Biophysics (SBBf); 2021

The nature and size of the accessible surface area (SASA) of the polypeptide chain plays a key role in protein folding and complex formation. SASA can be probed with diazirine (DZN), a tiny precursor of the extremely reactive methylene carbene (:CH2). Sparse methylated sites left on the polypeptide provide revealing signs on the conformation. The metric extent of methylation (EM) derived from mass spectra discriminates between native and alternate states. Human alpha synuclein (AS) aggregates into oligomers and amyloid fibrils, constituents of Lewy bodies, a cytosolic hallmark of Parkinson disease. DZN labeling proves particularly fit to analyzing the conformational plasticity inherent to this intrinsically disordered protein. Unlike folded proteins where EM differs markedly between native and unfolded states, AS in buffer or in 6 M GdmCl displays a similarly enhanced value, revealing high solvent accessibility under normal physiological conditions. Strikingly, AS fibrils display an enhanced EM value compared to the monomer, as a consequence of the organization of a hydrophobic interface. In addition, location of methylated sites by multidimensional NMR (1H15N-HSQC) opens a vast panorama. The extent of :CH2 reaction across the surface of AS defines sequential intensity profiles. Differential labeling signs point to the involvement of the N-terminal domain in fibril formation. We anticipate that the combined picture derived from MS and NMR data will serve to elucidate the contribution to fibrillation of the different structural moieties of AS, allowing the appraisal of species belonging to the monomeric ensemble, the lesser-known oligomers, and the fibrillar aggregates. Key words: synuclein, methylation, diazirineFunding: UBACyT, CONICET and ANPCyT