An examination of the surface of the intrinsically disordered protein alpha synuclein

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

Seattle

Simposio; 33rd Annual Symposium of The Protein Society; 2019

The nature and size of the accessible surface area (SASA) of the polypeptide chain plays a pivotal role in protein folding and complex formation. To investigate SASA, we employ diazirine (DZN), a minute precursor of the extremely reactive methylene carbene (:CH2). Methylation signatures left on the polypeptide provide telltale clues on conformation and interactions. The extent of methylation (EM) metric derived straightforwardly from mass spectra (ESI-MS or MALDI-TOF) discriminates between native and alternate states. The archetypical IDP human alpha synuclein (AS) aggregates into amyloid fibrils, constituents of Lewy bodies, a hallmark of Parkinson disease. DZN labeling proves particularly fit to analyzing the conformational plasticity inherent to IDPs, an elusive goal by other classical biophysical methods. Unlike well-structured proteins where the methylation signal differs strikingly between native and unfolded states, AS in buffer or equilibrated in 6 M GdmCl display a similarly enhanced EM value, pointing to the high solvent exposure of AS under physiological conditions. Interestingly, compaction of monomeric AS due to calcium binding yields a somewhat decreased EM value. Remarkably, AS fibrils cause a larger fall in the EM value, a consequence of surface occlusion at an interface. Tryptic fragmentation of AS coupled to MALDI-TOF analysis is revealing methylation patterns at increased resolution. The peptide coverage achieved so far allows to construct a solvent-accessibility map of the different sequential domains of AS. This new information illuminates the role played by the constituent parts in the monomeric ensemble as well as the changes observed en route to the fibrillar aggregate.