Abstract
α-synuclein (αS) is an intrinsically disordered protein whose fibrillar aggregates are the major constituents of Lewy bodies in Parkinson's disease. Although the specific function of αS is still unclear, a general consensus is forming that it has a key role in regulating the process of neurotransmitter release, which is associated with the mediation of synaptic vesicle interactions and assembly. Here we report the analysis of wild-type αS and two mutational variants linked to familial Parkinson's disease to describe the structural basis of a molecular mechanism enabling αS to induce the clustering of synaptic vesicles. We provide support for this ‘double-anchor' mechanism by rationally designing and experimentally testing a further mutational variant of αS engineered to promote stronger interactions between synaptic vesicles. Our results characterize the nature of the active conformations of αS that mediate the clustering of synaptic vesicles, and indicate their relevance in both functional and pathological contexts.
Highlights
Because of its intrinsic ability to bind to a wide variety of biological membranes, the physiological state of membranebound aS is extremely difficult to characterize, as a variety of factors, including the presence of detergents[22] and chemical modification of the protein[27], can alter dramatically the structural properties of its bound state[15]
The mechanism, which was verified using both synthetic lipid vesicles and synaptic vesicles purified from rat brain, provides evidence that the specific level of affinity for membrane binding of the non-amyloid-b component (NAC) region of aS is a fundamental functional property enabling this protein to mediate the interaction between vesicles
Using solution-state and solid-state nuclear magnetic resonance (NMR) spectroscopy in combination with cryo-electron microscopy and stimulated emission depletion (STED) imaging, we have characterized the structural properties at the surface of synaptic-like vesicles of the familial aS mutants A30P31 and E46K32 and compared their behaviour with that of the wild-type protein[28]
Summary
Because of its intrinsic ability to bind to a wide variety of biological membranes, the physiological state of membranebound aS is extremely difficult to characterize, as a variety of factors, including the presence of detergents[22] and chemical modification of the protein[27], can alter dramatically the structural properties of its bound state[15]. Three major regions were identified to have distinct structural and dynamical properties that influence in different ways the nature of the membrane-bound state of aS28; these regions include an N-terminal a-helical segment, acting as the membrane-anchor, an unstructured C-terminal region, weakly associated with the membrane, and a central region, undergoing order–disorder transitions in the membrane-bound state and determining the affinity of aS for lipid bilayers of different composition[28] This structural variability indicates that it is of fundamental importance to investigate the binding of aS to lipid membranes under conditions that reproduce as closely as possible the physiological environment relevant to that of presynaptic vesicles[15]. The mechanism, which was verified using both synthetic lipid vesicles and synaptic vesicles purified from rat brain, provides evidence that the specific level of affinity for membrane binding of the non-amyloid-b component (NAC) region of aS is a fundamental functional property enabling this protein to mediate the interaction between vesicles
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