Multiparametric Fluorescence Detection of Early Stages in the Amyloid Protein Aggregation of Pyrene-labeled α-Synuclein

Thirunavukkuarasu, Shyamala; Jares‐Erijman, Elizabeth A.; Jovin, Thomas M.

doi:10.1016/j.jmb.2008.03.034

Cited by 73 publications

(85 citation statements)

References 42 publications

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“…Moreover, our data highlight the key role of the N-terminal part of the protein on early oligomeric interactions. In this regard, we observe that a number of small-molecule anti-amyloid compounds preferentially target the regions enclosing residues ϳVal 3 -Lys 23 and/or Leu 38 -Gly 51 (39,43). As discussed previously, our excimer profile suggests that ␤5 region is not proximal in oAS (Fig.…”

Section: Discussionsupporting

confidence: 75%

“…This ability of pyrene has been successfully employed to gather structural details of amyloid fibrils formed by a yeast prion (37) and toxic amyloid oligomers assembled from a non-disease related protein (38) as well as for monitoring ␣-synuclein amyloid formation (23). Inspired by these works, we relied on excimer signals arising from pyrene-labeled oAS to infer about intermolecular proximities among the monomers.…”

Section: Discussionmentioning

confidence: 99%

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Structural Insights into Amyloid Oligomers of the Parkinson Disease-related Protein α-Synuclein

Gallea

Celej

2014

Journal of Biological Chemistry

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Background: Soluble oligomers of ␣-synuclein, rather than the amyloid fibrils, are presumed to be more neurotoxic in Parkinson disease. Results: A site-specific fluorescence approach is used to unravel the internal architecture of ␣-synuclein oligomers. Conclusion: ␣-Synuclein oligomers are organized aggregates with a defined network of intermolecular contacts. Significance: This contact map can be used for developing molecular models, essential for mechanistic studies and drug design.

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Section: Discussionsupporting

confidence: 75%

Section: Discussionmentioning

confidence: 99%

Structural Insights into Amyloid Oligomers of the Parkinson Disease-related Protein α-Synuclein

Gallea

Celej

2014

Journal of Biological Chemistry

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“…The vanishing fluorescence anisotropy of this compound in aqueous solution fits well to the short rotational diffusion time θ ≈ 150 ps expected for molecules with the size of the used DPP compounds [37]. For the α-synuclein monomer and fibrils the orientational motion is much slower and in the order of θ = 1-2 ns and θ ≫ 10 ns respectively [38]. When sery313b is rigidly bound to the protein large values of r are expected.…”

Section: Absorption and Emission Propertiessupporting

confidence: 67%

Anle138b and related compounds are aggregation specific fluorescence markers and reveal high affinity binding to α-synuclein aggregates

Deeg

Reiner

Schmidt

et al. 2015

Biochimica et Biophysica Acta (BBA) - General Subjects

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Background: Special diphenyl-pyrazole compounds and in particular anle138b were found to reduce the progression of prion and Parkinson's disease in animal models. The therapeutic impact of these compounds was attributed to the modulation of α-synuclein and prion-protein aggregation related to these diseases. Methods: Photophysical and photochemical properties of the diphenyl-pyrazole compounds anle138b, anle186b and sery313b and their interaction with monomeric and aggregated α-synuclein were studied by fluorescence techniques. Results: The fluorescence emission of diphenyl-pyrazole is strongly increased upon incubation with α-synuclein fibrils, while no change in fluorescence emission is found when brought in contact with monomeric α-synuclein. This points to a distinct interaction between diphenyl-pyrazole and the fibrillar structure with a high binding affinity (K d = 190 ± 120 nM) for anle138b. Several α-synuclein proteins form a hydrophobic binding pocket for the diphenyl-pyrazole compound. A UV-induced dehalogenation reaction was observed for anle138b which is modulated by the hydrophobic environment of the fibrils. Conclusion: Fluorescence of the investigated diphenyl-pyrazole compounds strongly increases upon binding to fibrillar α-synuclein structures. Binding at high affinity occurs to hydrophobic pockets in the fibrils. General significance: The observed particular fluorescence properties of the diphenyl-pyrazole molecules open new possibilities for the investigation of the mode of action of these compounds in neurodegenerative diseases. The high binding affinity to aggregates and the strong increase in fluorescence upon binding make the compounds promising fluorescence markers for the analysis of aggregation-dependent epitopes.

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“…Protein was expressed and purified as previously described (50,51). For a 1-liter culture, isotopically ( 13 C/ 15 N) enriched WT ␣-syn was produced by growing Escherichia coli BL21(DE3)pLysS in M9 medium supplemented with [ 13 C]glucose (2 g) and 15 NH 4 Cl (1 g). Imiglucerase, purified recombinant GCase, was obtained from Genzyme Corp. and N370S GCase was a generous gift from Dr. Timothy Edmonds (Genzyme Corp.).…”

Section: Methodsmentioning

confidence: 99%

“…Because abnormally misfolded proteins and amyloid deposits are found in other neurodegenerative diseases such as Alzheimer disease and prion encephalopathies (11), considerable research has focused at gaining molecular insights into fibril formation (12)(13)(14)(15)(16)(17) and biomolecules that stimulate/inhibit this process (18 -21). For example, lipid-protein interactions are of particular interest because ␣-syn localizes near synaptic vesicles (2,18) and is thought to act as a chaperone in SNARE complex assembly, which is necessary for neurotransmitter release from presynaptic vesicles (23).…”

mentioning

confidence: 99%

α-Synuclein Interacts with Glucocerebrosidase Providing a Molecular Link between Parkinson and Gaucher Diseases

Yap

Gruschus

Velayati

et al. 2011

Journal of Biological Chemistry

165

143

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The presynaptic protein ␣-synuclein (␣-syn), particularly in its amyloid form, is widely recognized for its involvement in Parkinson disease (PD). Recent genetic studies reveal that mutations in the gene GBA are the most widespread genetic risk factor for parkinsonism identified to date. GBA encodes for glucocerebrosidase (GCase), the enzyme deficient in the lysosomal storage disorder, Gaucher disease (GD). In this work, we investigated the possibility of a physical linkage between ␣-syn and GCase, examining both wild type and the GD-related N370S mutant enzyme. Using fluorescence and nuclear magnetic resonance spectroscopy, we determined that ␣-syn and GCase interact selectively under lysosomal solution conditions (pH 5.5) and mapped the interaction site to the ␣-syn C-terminal residues, 118 -137. This ␣-syn-GCase complex does not form at pH 7.4 and is stabilized by electrostatics, with dissociation constants ranging from 1.2 to 22 M in the presence of 25 to 100 mM NaCl. Intriguingly, the N370S mutant form of GCase has a reduced affinity for ␣-syn, as does the inhibitor conduritol-␤-epoxidebound enzyme. Immunoprecipitation and immunofluorescence studies verified this interaction in human tissue and neuronal cell culture, respectively. Although our data do not preclude protein-protein interactions in other cellular milieux, we suggest that the ␣-syn-GCase association is favored in the lysosome, and that this noncovalent interaction provides the groundwork to explore molecular mechanisms linking PD with mutant GBA alleles.

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Multiparametric Fluorescence Detection of Early Stages in the Amyloid Protein Aggregation of Pyrene-labeled α-Synuclein

Cited by 73 publications

References 42 publications

Structural Insights into Amyloid Oligomers of the Parkinson Disease-related Protein α-Synuclein

Structural Insights into Amyloid Oligomers of the Parkinson Disease-related Protein α-Synuclein

Anle138b and related compounds are aggregation specific fluorescence markers and reveal high affinity binding to α-synuclein aggregates

α-Synuclein Interacts with Glucocerebrosidase Providing a Molecular Link between Parkinson and Gaucher Diseases

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