title: Cu(II) and dopamine bind to α-synuclein and cause large conformational changes.
abstract: α-Synuclein (AS) is an intrinsically disordered protein that can misfold and aggregate to form Lewy bodies in dopaminergic neurons, a classic hallmark of Parkinson's disease. The binding of Cu(II) and dopamine to AS was evaluated by nanopore analysis with α-hemolysin. In the absence of Cu(II), wild-type AS (1 μM) readily translocated through the pore with a blockade current of--85 pA, but mostly bumping events were observed in the presence of 25 μM Cu(II). A binding site in the N-terminus was confirmed, because Cu(II) had no effect on the event profile of a peptide consisting of the C-terminal 96-140 residues. In the presence of dopamine (25 μM), the translocation events at--85 pA shifted to--80 pA, which also represents translocation events, because the event time decreases with increasing voltage. Events at--80 pA were also observed for the mutant A30P AS in the presence of dopamine. Event profiles for an N-terminal 1-60-residue peptide and a C-terminal 96-140-residue peptide were both altered in the presence of 25 μM dopamine. In contrast, dopamine had little effect on the CD spectrum of AS, and a single binding site with a Ka of 3.5 × 10(3) m(-1) was estimated by isothermal titration calorimetry. Thus, dopamine can interact with both the N-terminus and the C-terminus. Two-dimensional NMR spectroscopy of AS in the presence of dopamine showed that there were significant changes in the spectra in all regions of the protein. According to these findings, a model is presented in which dopamine induces folding between the N-terminus and C-terminus of AS. Partially folding conformations such as this may represent important intermediates in the misfolding of AS that leads to fibrillization.
authors.affiliation: Department of Biochemistry, University of Saskatchewan, Saskatoon, SK, Canada.
authors.forename: Omid
authors.initials: O
authors.lastname: Tavassoly
authors.forename: Sergiy
authors.initials: S
authors.lastname: Nokhrin
authors.forename: Oleg Y
authors.initials: OY
authors.lastname: Dmitriev
authors.forename: Jeremy S
authors.initials: JS
authors.lastname: Lee
mesh_terms: D001665:Binding Sites; D002151:Calorimetry; D003300:Copper / Q000378:metabolism*; D004298:Dopamine / Q000378:metabolism*; D011487:Protein Conformation; D021381:Protein Transport; D051844:alpha-Synuclein / Q000737:chemistry / Q000378:metabolism*
chemical_list: D051844:alpha-Synuclein; D003300:Copper; D004298:Dopamine
keywords: Parkinson's disease; dopamine; intrinsically disordered protein; nanopore analysis; α-synuclein