title: Earliest events in α-synuclein fibrillation probed with the fluorescence of intrinsic tyrosines.
abstract: The fluorescence of the four tyrosines of α-synuclein (Syn) was used for probing the earliest events preceding the fibrillation of Syn, during the onset of the so-called lag-time of fibrillation. Steady-state fluorescence experiments revealed an increase in the fluorescence intensity (FI) for Syn solutions at pH values 3 and 2, in comparison with pH7, and fluorescence decays indicated that the FI increase did not result from suppression of excited-state proton transfer from the tyrosines to aspartates and glutamates, exposure of tyrosines to more hydrophobic environments, or reduction of homo-energy transfer. Instead, the FI increase was due to changes in the population of the tyrosine rotamers at low pH values. Stopped-flow experiments (pH-jumps) showed that the FI enhancement involves two processes: a fast (sub-7 ms) intramolecular (concentration-independent) process, which we assign to the protein collapse at low pH, and a slower intermolecular (concentration-dependent) process of protein dimerization/oligomerization, starting at 4-10s after acidification. To the best of our knowledge, this is the first work on the experimental detection of these earliest processes in the fibrillation of Syn.
authors.affiliation: Centro de Química Estrutural, Instituto Superior Técnico, UTL, Lisboa, Portugal. Electronic address: marco.saraiva@ist.utl.pt.
authors.forename: Marco A
authors.initials: MA
authors.lastname: Saraiva
authors.affiliation: Instituto de Tecnologia Química e Biológica, UNL, Oeiras, Portugal.
authors.forename: Carla D
authors.initials: CD
authors.lastname: Jorge
authors.affiliation: Instituto de Tecnologia Química e Biológica, UNL, Oeiras, Portugal.
authors.forename: Helena
authors.initials: H
authors.lastname: Santos
authors.affiliation: Centro de Química Estrutural, Instituto Superior Técnico, UTL, Lisboa, Portugal.
authors.forename: António L
authors.initials: AL
authors.lastname: Maçanita
mesh_terms: D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D057927:Hydrophobic and Hydrophilic Interactions; D055503:Protein Multimerization; D011522:Protons; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D013056:Spectrophotometry, Ultraviolet; D014443:Tyrosine / Q000737:chemistry*; D051844:alpha-Synuclein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D005456:Fluorescent Dyes; D011522:Protons; D011994:Recombinant Proteins; D051844:alpha-Synuclein; D014443:Tyrosine
keywords: Intrinsic fluorescence probe; Protein aggregation; Stopped-flow; pH-jump