Interaction of α-synuclein with vesicles that mimic mitochondrial membranes

Abstract

α-Synuclein, an intrinsically-disordered protein associated with Parkinson’s disease, interacts with mitochondria, but the details of this interaction are unknown. We probed the interaction of α-synuclein and its A30P variant with lipid vesicles by using fluorescence anisotropy and 19F nuclear magnetic resonance. Both proteins interact strongly with large unilamellar vesicles whose composition is similar to that of the inner mitochondrial membrane, which contains cardiolipin. However, the proteins have no affinity for vesicles mimicking the outer mitochondrial membrane, which lacks cardiolipin. The 19F data show that the interaction involves α-synuclein’s N-terminal region. These data indicate that the middle of the N-terminal region, which contains the KAKEGVVAAAE repeats, is involved in binding, probably via electrostatic interactions between the lysines and cardiolipin. We also found that the strength of α-synuclein binding depends on the nature of the cardiolipin acyl side chains. Eliminating one double bond increases affinity, while complete saturation dramatically decreases affinity. Increasing the temperature increases the binding of wild-type, but not the A30P variant. The data are interpreted in terms of the properties of the protein, cardiolipin demixing within the vesicles upon binding of α-synuclein, and packing density. The results advance our understanding of α-synuclein’s interaction with mitochondrial membranes.

Number and Brightness analysis of alpha-synuclein oligomerization and the associated mitochondrial morphology alterations in live cells

“…They showed that αsyn overexpression causes mitochondrial fragmentation, but they could not verify the conformation that αsyn acquired while damaging mitochondria. Furthermore, they previously showed by FRET that there is a specific interaction between αsyn and mitochondrial membranes [36], as reported also using an in vitro approach [42]. …”

“…They hypothesized that the variations in mitochondria morphology are due to the direct interaction of αsyn oligomers with mitochondrial membranes. This idea is further supported by the evidence of αsyn interaction with mitochondria [36] and localization of αsyn at membranes mimicking mitochondrial membranes [42] or within mitochondria [61]. Moreover, it was proposed that αsyn can inhibit mitochondrial fusion processes, again because of the protein special interactions with mitochondrial membranes [43] or that mitochondrial fission is increased through an increased Drp1 translocation to mitochondria due to αsyn overexpression [62].…”

Adsorption of α-Synuclein to Supported Lipid Bilayers: Positioning and Role of Electrostatics

“…Mitochondria have recently attracted interest as potential actors in the pathogenesis of PD. 31,35 Therefore, these types of lipids were chosen for this study. The average charge density is expected to be the same for the studied anionic bilayers with 30% PS or 15% CL (see Discussion for further details).…”

19F-Modified Proteins and 19F-Containing Ligands as Tools in Solution NMR Studies of Protein Interactions

“…In follow-up studies by 19 F NMR and fluorescence, interactions with large unilamellar vesicles, whose composition is similar to that of the inner and outer mitochondrial membrane, were probed. From these studies, the strength of α-synuclein binding was found to depend on the presence of cardiolipin, a component of the inner mitochondrial membrane (Zigoneanu, Yang, Krois, Haque, & Pielak, 2012). 19 F NMR has also been used to examine the emersion depth, secondary structure, and topology of the first transmembrane segment of the membrane protein diacyl glycerol kinase (DAGK).…”