Abstract
Alpha-synuclein (α-syn) is a small presynaptic protein that is believed to play an important role in the pathogenesis of Parkinson's disease (PD). It localizes to presynaptic terminals where it partitions between a cytosolic soluble and a lipid-bound state. Recent evidence suggests that α-syn can also associate with mitochondrial membranes where it interacts with a unique anionic phospholipid cardiolipin (CL). Here, we examine the conformation of the flexible fragments of a monomeric α-syn bound to lipid vesicles composed of anionic 1,2-dioleoyl-sn-glycero-3-phosphate (DOPA) and 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) lipids, of tetraoleoyl CL (TOCL) and DOPC, and of fibrils. The dynamic properties of α-syn associated with DOPA:DOPC vesicles were the most favorable for conducting three-dimensional NMR experiments, and the 13C, 15N and amide 1H chemical shifts of the flexible and disordered C-terminus of α-syn could be assigned using three-dimensional through-bond magic angle spinning NMR spectroscopy. Although the C-terminus is more dynamically constrained in fibrils and in α-syn bound to TOCL:DOPC vesicles, a direct comparison of carbon chemical shifts detected using through bond two-dimensional spectroscopy indicates that the C-terminus is flexible and unstructured in all the three samples.
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Acknowledgements
We thank Ms. Paula Boubel for her assistance with collecting initial MAS NMR data, and Dr. Sameer Al-Abdul-Wahid of the University of Guelph NMR Centre for help with setting 3D NMR experiments. We are grateful to Prof. Alfonso De Simone (Imperial College London) for sharing chemical shift data for α-syn.
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Chemical shifts for assigned residues in the sample of α-syn bound to DOPA:DOPC liposomes have been deposited to BioMagResBank (http://www.bmrb.wisc.edu) under BMRB entry 50746.
Funding
This research was supported by Natural Science ang Engineering and Research Council of Canada (Discovery Grant RGPIN-2014-04547 to V.L.; Discovery Grant RGPIN-121541 to G.H.) and by the Parkinson Foundation of Canada (Grant 2018-00285 to V.L.) Canadian Institutes for Health Research Project Grant (PJT-159443-2018) to S.D.R, Canada Foundation for Innovation and Ontario Ministry of Economic Development and Innovation, and by the University of Guelph through their support of the Advanced Analysis Centre.
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Medeiros, J., Bamm, V.V., Jany, C. et al. Partial magic angle spinning NMR 1H, 13C, 15N resonance assignments of the flexible regions of a monomeric alpha-synuclein: conformation of C-terminus in the lipid-bound and amyloid fibril states. Biomol NMR Assign 15, 297–303 (2021). https://doi.org/10.1007/s12104-021-10020-z
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DOI: https://doi.org/10.1007/s12104-021-10020-z