The fusogenic tilted peptide (67-78) of alpha-synuclein is a cholesterolbinding domain

Parkinson's disease-associated alpha-synuclein is an amyloidogenic protein not only expressed in the cytoplasm of neurons, but also secreted in the extracellular space and internalized into glial cells through a lipid raft-dependent process. We previously showed that alpha-synuclein interacts with raft glycosphingolipids through a structural motif common to various viral and amyloidogenic proteins. Here we report that alpha-synuclein also interacts with cholesterol, as assessed by surface pressure measurements of cholesterol-containing monolayers. Using a panel of recombinant fragments and synthetic peptides, we identified two distinct cholesterol-binding domains in alpha-synuclein. One of these domains, which corresponds to the tilted peptide of alpha-synuclein (67-78), bound cholesterol with high affinity and was toxic for cultured astrocytes. Molecular modeling suggested that cholesterol binds to this peptide with a tilt angle of 46 degrees. alpha-synuclein also contains a cholesterol recognition consensus motif, which had a lower affinity for cholesterol and was devoid of toxicity. This motif is encased in the glycosphingolipid-binding domain (34-45) of alpha-synuclein. In raft-like model membranes containing both cholesterol and glycosphingolipids, the head groups of glycosphingolipids prevented the accessibility of cholesterol to exogenous ligands. Nevertheless, cholesterol appeared to 'signal' its presence by tuning glycosphingolipid conformation, thereby facilitating alpha-synuclein binding to raft-like membranes. We propose that the association of alpha-synuclein with lipid rafts involves both the binding of alpha-synuclein (34-45) to glycosphingolipids, and the interaction of the fusogenic tilted peptide (67-78) with cholesterol. Coincidentally, a similar mechanism is used by viruses (HIV-1, HTLV-I, Ebola) which display a tilted peptide and fuse with host cell membranes through a sphingolipid/cholesterol-dependent process. (C) 2011 Elsevier B.V. All rights reserved.