Figure 5.

Increasing alpha-synuclein concentration shifts alpha-synuclein-bound complexes into a different state, but this does not happen with phospho-synuclein.A₁, decreasing alpha-synuclein (aSyn) protein concentration from 57 to 1 μM with 300 bp DNA concentration fixed at either 0.1 nM (left) or 6 nM (right) produces a change in the apparent length of the complex from a higher value shift (running >1517 bp DNA, red arrow) to a lower value shift (running ∼600 bp, green arrow). Unshifted (unbound) DNA marked by the blue arrow. A₂, Western blot showing aSyn protein loaded into each lane. A₃, DNA gel (green) and aSyn Western blot protein (red) localization from the same experiment. A₄, group data showing shifted fraction as a function of aSyn concentration for the two different fixed DNA concentrations. The low DNA concentration curve is left-shifted (aSyn EC50: low DNA conc. = 0.273 μM, R² = 0.619; high DNA conc.=5.846 μM, R² = 0.977, three-parameter dose–response curve; N = 3–4 gels, x-axis aSyn concentration on log scale). B₁, decreasing serine-129 phosphorylated alpha-synuclein (pSyn) protein concentration from 57 to 1 μM with 300 bp DNA concentration fixed at either 0.1 nM (left) or 6 nM (right) produces only a complex with an apparent length of ∼500 bp (green arrow) at the higher pSyn concentrations. Unshifted (unbound) DNA marked by the blue arrow. B₂, Western blot showing pSyn protein loaded into each lane. B₃, DNA gel (green) and pSyn Western blot (red) localization from the same experiment. B₄, group data showing a similar shifted fraction as a function of pSyn concentration for the two different fixed DNA concentrations (pSyn EC50: low DNA conc.=27.34 μM, R² = 0.779; high DNA conc.=32.49 μM, R² = 0.981, four-parameter dose–response curve; N = 3 gels, x-axis pSyn concentration on log scale).