α-Synuclein as an intrinsically disordered monomer--fact or artefact?

Fibrillization of the protein α-synuclein (α-syn) is a hallmark of Parkinson's disease and other α-synucleinopathies. The well-established idea that α-syn is a natively disordered monomer prone to forming fibrils was recently challenged by data showing that the protein mostly exists in vitro and in vivo as helically folded tetramers that are resistant to fibrillization. These apparently conflicting findings may be reconciled by the idea that α-syn exists as a disordered monomer in equilibrium with variable amounts of dynamic oligomeric species. In this context, varying the approaches used for protein purification, such as the method used to lyse cells or the inclusion of denaturing agents, could dramatically perturb this equilibrium and hence alter the relative abundance of the disordered monomer. In the present study, we investigated how the current methods for α-syn purification affect the structure and oligomeric state of the protein, and we discuss the main pitfalls associated with the production of recombinant α-syn in Escherichia coli. We demonstrate that α-syn was expressed in E. coli as a disordered monomer independent of both the cell lysis method and the use of heating/acidification for protein purification. In addition, we provide convincing evidence that the disordered monomer exists in equilibrium with a dynamic dimer, which is not an artefact of the cross-linking protocol as previously suggested. Unlike the helically folded tetramer, α-syn dimer is prone to fibrillate and thus it may be an interesting target for anti-fibrillogenic molecules.