One-pot total chemical synthesis of human α-synuclein

Fauvet, Bruno; Butterfield, Sara M.; Fuks, Jonas M.; Brik, Ashraf; Lashuel, Hilal A.

doi:10.1039/c3cc45353g

Cited by 42 publications

(34 citation statements)

References 23 publications

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“…Because c-Abl phosphorylates aSyn at Y125 as well as the primary site Y39, 30 we first opted for a three-fragment one-pot chemical protein semisynthesis strategy adapted from a previously described method, 35 to generate pY39 aSyn, with two ligation sites at A30 and A56 (Fig. S2A).…”

Section: Resultsmentioning

confidence: 99%

Semisynthetic and in Vitro Phosphorylation of Alpha-Synuclein at Y39 Promotes Functional Partly Helical Membrane-Bound States Resembling Those Induced by PD Mutations

et al. 2016

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Alpha-synuclein is a presynaptic protein of poorly understood function that is linked to both genetic and sporadic forms of Parkinson’s disease. We have proposed that alpha-synuclein may function specifically at synaptic vesicles docked at the plasma membrane, and that the broken-helix state of the protein, comprising two antiparallel membrane-bound helices connected by a non-helical linker, may target the protein to such docked vesicles by spanning between the vesicle and the plasma membrane. Here we demonstrate that phosphorylation of alpha-synuclein at tyrosine 39, carried out by c-Abl in vivo, may facilitate interconversion of synuclein from the vesicle-bound extended-helix state to the broken-helix state. Specifically, in the presence of lipid vesicles Y39 phosphorylation leads to decreased binding of a region corresponding to helix-2 of the broken-helix state, potentially freeing this region of the protein to interact with other membrane surfaces. This effect is largely recapitulated by the phosphomimetic mutation Y39E and expression of this mutant in yeast results in decreased membrane localization. Intriguingly, the effects of Y39 phosphorylation on membrane binding closely resemble those of the recently reported disease linked mutation G51D. These findings suggest that Y39 phosphorylation could modulate functional aspects of alpha-synuclein, and perhaps influence pathological aggregation of the protein as well.

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Section: Resultsmentioning

confidence: 99%

Semisynthetic and in Vitro Phosphorylation of Alpha-Synuclein at Y39 Promotes Functional Partly Helical Membrane-Bound States Resembling Those Induced by PD Mutations

et al. 2016

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“…Adapted from Ref. 131. D, immunocytochemistry (left) and Western blot (right) show that pY125 is a very labile modification, highlighting that the semisynthetic pY125 ␣-syn standard can be very useful for developing protocols to stabilize this modification in biological samples.…”

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confidence: 99%

Alpha-synuclein Post-translational Modifications as Potential Biomarkers for Parkinson Disease and Other Synucleinopathies

Schmid

Fauvet

Moniatte

et al. 2013

Molecular & Cellular Proteomics

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The development of novel therapies against neurodegenerative disorders requires the ability to detect their early, presymptomatic manifestations in order to enable treatment before irreversible cellular damage occurs. Precocious signs indicative of neurodegeneration include characteristic changes in certain protein levels, which can be used as diagnostic biomarkers when they can be detected in fluids such as blood plasma or cerebrospinal fluid. In the case of synucleinopathies, cerebrospinal alpha-synuclein (␣-syn) has attracted great interest as a potential biomarker; however, there is ongoing debate regarding the association between cerebrospinal ␣-syn levels and neurodegeneration in Parkinson disease and synucleinopathies. Post-translational modifications (PTMs) have emerged as important determinants of ␣-syn's physiological and pathological functions. Several PTMs are enriched within Lewy bodies and exist at higher levels in ␣-synucleinopathy brains, suggesting that certain modified forms of ␣-syn might be more relevant biomarkers than the total ␣-syn levels. However, the quantification of PTMs in bodily fluids poses several challenges. This review describes the limitations of current immunoassay-based ␣-syn quantification methods and highlights how these limitations can be overcome using novel mass-spectrometrybased assays. In addition, we describe how advances in chemical synthesis, which have enabled the preparation of ␣-syn proteins that are site-specifically modified at single or multiple residues, can facilitate the development of more accurate assays for detecting and quantifying ␣-syn PTMs in health and disease. Molecular & Cellular Proteomics

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“…This group can be cleaved afterward using methoxyamine·HCl at pH 4 (Monbaliu and Katritzky, 2012). An additional benefit of the Thz conversion is the suitability to perform the transformation within the ligation buffer following a one-pot approach (Bang and Kent, 2004;Fauvet et al, 2013). Therefore, to use Thz in the chemical synthesis strategy followed by NCL ligation conditions must be optimized and described above.…”

Section: Structurementioning

confidence: 99%

Challenges and Perspectives in Chemical Synthesis of Highly Hydrophobic Peptides

Mueller

Baumruck

Zhdanova

et al. 2020

Front. Bioeng. Biotechnol.

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Solid phase peptide synthesis (SPPS) provides the possibility to chemically synthesize peptides and proteins. Applying the method on hydrophilic structures is usually without major drawbacks but faces extreme complications when it comes to "difficult sequences." These includes the vitally important, ubiquitously present and structurally demanding membrane proteins and their functional parts, such as ion channels, G-protein receptors, and other pore-forming structures. Standard synthetic and ligation protocols are not enough for a successful synthesis of these challenging sequences. In this review we highlight, summarize and evaluate the possibilities for synthetic production of "difficult sequences" by SPPS, native chemical ligation (NCL) and follow-up protocols.

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One-pot total chemical synthesis of human α-synuclein

Cited by 42 publications

References 23 publications

Semisynthetic and in Vitro Phosphorylation of Alpha-Synuclein at Y39 Promotes Functional Partly Helical Membrane-Bound States Resembling Those Induced by PD Mutations

Semisynthetic and in Vitro Phosphorylation of Alpha-Synuclein at Y39 Promotes Functional Partly Helical Membrane-Bound States Resembling Those Induced by PD Mutations

Alpha-synuclein Post-translational Modifications as Potential Biomarkers for Parkinson Disease and Other Synucleinopathies

Challenges and Perspectives in Chemical Synthesis of Highly Hydrophobic Peptides

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