Spin-Label EPR on alpha-Synuclein Reveals Differences in the Membrane Binding Affinity of the Two Antiparallel Helices

Abstract

The putative function of the Parkinson's disease-related protein -Synuclein (S) is thought to involve membrane binding. Therefore, the interaction of S with membranes composed of zwitterionic (POPC) and anionic (POPG) lipids was investigated through the mobility of spin labels attached to the protein. Differently labelled variants of S were produced, containing a spin label at positions 9, 18 (both helix 1), 69, 90 (both helix 2), and 140 (C terminus). Protein binding to POPC/POPG vesicles for all but S140 resulted in two mobility components with correlation times of 0.5 and 3 ns, for POPG mole fractions >0.4. Monitoring these components as a function of the POPG mole fraction revealed that at low negative-charge densities helix 1 is more tightly bound than helix 2; this indicates a partially bound form of S. Thus, the interaction of S with membranes of low charge densities might be initiated at helix 1. The local binding information thus obtained gives a more differentiated picture of the affinity of S to membranes. These findings contribute to our understanding of the details and structural consequences of S-membrane interactions.