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O-GlcNAc modification blocks the aggregation and toxicity of the protein α-synuclein associated with Parkinson's disease

Nature Chemistry volume 7, pages 913–920 (2015)Cite this article

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Abstract

Several aggregation-prone proteins associated with neurodegenerative diseases can be modified by O-linked N-acetyl-glucosamine (O-GlcNAc) in vivo. One of these proteins, α-synuclein, is a toxic aggregating protein associated with synucleinopathies, including Parkinson's disease. However, the effect of O-GlcNAcylation on α-synuclein is not clear. Here, we use synthetic protein chemistry to generate both unmodified α-synuclein and α-synuclein bearing a site-specific O-GlcNAc modification at the physiologically relevant threonine residue 72. We show that this single modification has a notable and substoichiometric inhibitory effect on α-synuclein aggregation, while not affecting the membrane binding or bending properties of α-synuclein. O-GlcNAcylation is also shown to affect the phosphorylation of α-synuclein in vitro and block the toxicity of α-synuclein that was exogenously added to cells in culture. These results suggest that increasing O-GlcNAcylation may slow the progression of synucleinopathies and further support a general function for O-GlcNAc in preventing protein aggregation.

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Figure 1: O-GlcNAc modification and α-synuclein.
Figure 2: Semisynthesis of α-synuclein.
Figure 3: O-GlcNAcylation blocks α-synuclein aggregation.
Figure 4: O-GlcNAcylation has no effect on α-synuclein membrane binding or bending.
Figure 5: O-GlcNAcylation blocks α-synuclein toxicity.
Figure 6: O-GlcNAcylated α-synuclein is largely excluded from the protein aggregates.

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Acknowledgements

B.W.Z. is a fellow of the National Science Foundation Graduate Research Fellowship Program (DGE-0937362). This research was supported by the Michael J. Fox Foundation (M.R.P.), the National Institutes of Health (R01GM063915 to R.L. and R01NS081678 to D.B.A.) and in part by the US National Cancer Institute of the US National Institutes of Health (CCSG P30CA014089). Some circular dichroism and dynamic light scattering measurement were performed at the USC Nano Biophysics Core Facility. TEM of protein fibres was performed at the USC Center for Electron Microscopy and Microanalysis.

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Authors and Affiliations

  1. Department of Chemistry, University of Southern California, Los Angeles, 90089, California, USA

    Nicholas P. Marotta, Yu Hsuan Lin, Yuka E. Lewis, Balyn W. Zaro & Matthew R. Pratt

  2. Department of Biochemistry and Molecular Biology and the Zilka Neurogenetic Institute, University of Southern California, Los Angeles, 90089, California, USA

    Mark R. Ambroso & Ralf Langen

  3. Department of Molecular and Computational Biology, University of Southern California, Los Angeles, 90089, California, USA

    Maxwell T. Roth, Don B. Arnold & Matthew R. Pratt

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Contributions

N.P.M., Y.H.L., Y.E.L., M.R.A., B.W.Z., M.R., D.B.A., R.L. and M.R.P. designed experiments and interpreted data. N.P.M. carried out the synthesis of the synuclein proteins, performed aggregation reactions and collected circular dichroism and dynamic light scattering spectroscopy data, ThT measurements and TEM images. Y.H.L. performed cell-death assays. Y.E.L. performed mutant synuclein experiments. M.R.A. performed membrane-binding experiments. B.W.Z. performed SDS–PAGE analysis. M.R. collected primary neurons. N.P.M., Y.H.L., Y.E.L. and M.R.P. prepared the manuscript.

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Correspondence to Matthew R. Pratt.

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Marotta, N., Lin, Y., Lewis, Y. et al. O-GlcNAc modification blocks the aggregation and toxicity of the protein α-synuclein associated with Parkinson's disease. Nature Chem 7, 913–920 (2015). https://doi.org/10.1038/nchem.2361

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