Computational Analysis of Cholesterol Binding and Pore-Lining Regions in Alpha-Synuclein: Role in Mitochondrial Function

Alpha-synuclein (α-syn) protein is the major component of Lewy bodies, which are characteristic pathological trademarks for neurodegenerative diseases (e.g. Parkinson’s and Alzheimer’s diseases). It is primarily expressed in neural tissue, with smaller amounts found in heart, muscle and other tissues. The canonical form found in Homo sapiens (α-syn-1) contains 140 residues and interacts with neuronal mitochondria via an N-terminal 32 residue mitochondrial-targeting signal. All isoforms (there are 3) have multiple highly conserved lipid binding (KTKE(Q)G(Q) V) motifs, thought to mediate binding to phospholipid membranes. Two isoforms also contain an EF-hand-like (helixloop- helix) sequence found in a large family of calcium-binding proteins, as well as three copper binding sites. We investigate protein topology using computational analysis and find that each isoform contains a pore-lining region, two cholesterol-binding (CRAC/CARC) and three or four lipid binding motifs, with one cholesterol motif overlapping the pore-lining region. Two lipid-binding motifs also overlap the N-terminal mitochondrial-targeting region consistent with evidence that α-syn inserts into mitochondrial inner membrane. α-Syn-1 reportedly occurs physiologically as a helically folded tetramer that requires N-terminal acetylation. Thus, each α-syn-1 tetramer could contain 4 mitochondrial targeting regions, up to 4 pore-lining regions, 4 EF-hand domains, 8 bound cholesterol molecules and 16 lipid binding motifs with pore-lining regions merging to form a membrane channel. Cholesterol binding to CRAC motifs may in turn facilitate protein folding, Ca2+-channel formation, as well as mitochondrial membrane lipid-protein interactions, altering mitochondrial bioenergetics. Disruption of mitochondrial bioenergetics may be involved in the pathogenesis of Alzheimer’s disease and Parkinsonism.