Abstract
Involvement of the intrinsically disordered protein alpha-synuclein (a-syn) in Parkinson's disease (PD) and other synucleinopathies is complicated and difficult to trace on cellular and molecular levels. Recently we established that a-syn can compromise mitochondrial function by affecting the outer membrane channel VDAC (Voltage Dependent Anion Channel). In planar lipid membranes, a-syn blocks the VDAC pore, thus reducing transport of ions and metabolites mediated by this channel. Our results suggest that a-syn is able to use VDAC as an entryway to mitochondria, where it can target specific enzymes. We proposed that binding of synuclein molecules to the lipid membrane is crucial for both a-syn blocking and translocating through VDAC. Here we report an inhibitory effect of a small membrane-binding peptide (MBP) on the a-syn interaction with VDAC. Additions of MBP in micromolar concentrations to the same side of the membrane as a-syn results in a progressive reduction of the frequency of VDAC blockage events; at 50 µM of MBP the a-syn-induced blockages of the channel are virtually abolished. Furthermore, the presence of MBP affects only the frequency of blockage events, but not their duration, suggesting a competition for membrane binding of MBP and a-syn. Similarly, MBP reduces a-syn blockage of another large beta-barrel - the a-hemolysin channel. Using the method of bilayer overtone analysis, we found that MBP, similar to a-syn, has a propensity for membrane binding, but with a much lower affinity of Kd ∼ 3 μM, compared with a-syn-membrane binding with Kd ∼ 30 nM. Our results demonstrate that it is possible to regulate a-syn blockage and translocation through VDAC by a rationally designed peptide, thus suggesting new venues in the search for small-molecule inhibitors of a-syn mitochondrial toxicity in PD and other synucleinopathies.
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