Evidence for the Presence of Three Distinct Binding Sites for the Thioflavin T Class of Alzheimer's Disease PET Imaging Agents on β-Amyloid Peptide Fibrils

Lockhart, Andrew; Liang, Y. T.; Judd, Duncan B.; Merritt, Andy; Lowe, Peter N.; Morgenstern, Jennifer L.; Hong, Guizhu; Gee, Antony D.; Brown, John

doi:10.1074/jbc.m412056200

Cited by 210 publications

(309 citation statements)

References 33 publications

Supporting

Mentioning

278

Contrasting

Order By: Relevance

“…Apparently several α-synuclein units combined in the particular structure of the fibrillar aggregates are required to form one binding pocket. The dissociation constant in the 200 nM range and the number of α-synuclein monomers forming one binding site are small compared to results from investigations of thioflavin T. For the binding of thioflavin T to fibrils formed of the protein Aβ values of K d = 750 nM and B = 40.5 were determined [41]. The strong binding of anle138b to aggregated α-synuclein supports the key role of the interactions with misfolded α-synuclein in the therapeutic action.…”

Section: Quantitative Binding Experiments With Anle138bmentioning

confidence: 85%

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

View full text Add to dashboard Cite

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.

show abstract

Section: Quantitative Binding Experiments With Anle138bmentioning

confidence: 85%

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

View full text Add to dashboard Cite

show abstract

“…Aggregates of A␤ were prepared by allowing spontaneous aggregation in an A␤ solution at 25°C for at least 24 h, which was then divided into aliquots and stored at Ϸ20°C. The aggregation of A␤ was measured by assaying the fluorescence of TfT, which increases upon binding to A␤ aggregates [excitation filter of 435 nm (bandwidth, 5 nm) and emission filter of 486 nm (bandwidth, 10 nm)] (46,47).…”

Section: Preparation Of Hemementioning

confidence: 99%

Amyloid-β peptide binds with heme to form a peroxidase: Relationship to the cytopathologies of Alzheimer’s disease

Atamna¹,

Boyle²

2006

Proc. Natl. Acad. Sci. U.S.A.

267

279

View full text Add to dashboard Cite

Amyloid-␤ peptide (A␤) is the toxic agent in Alzheimer's disease (AD), although the mechanism causing the neurodegeneration is not known. We previously proposed a mechanism in which excessive A␤ binds to regulatory heme, triggering functional heme deficiency (HD), causing the key cytopathologies of AD. We demonstrated that HD triggers the release of oxidants (e.g., H 2O2) from mitochondria due to the loss of complex IV, which contains heme-a. Now we add more evidence that A␤ binding to regulatory heme in vivo is the mechanism by which A␤ causes HD. Heme binds to A␤, thus preventing A␤ aggregation by forming an A␤-heme complex in a cell-free system. We suggest that this complex depletes regulatory heme, which would explain the increase in heme synthesis and iron uptake we observe in human neuroblastoma cells. The A␤-heme complex is shown to be a peroxidase, which catalyzes the oxidation of serotonin and 3,4-dihydroxyphenylalanine by H 2O2. Curcumin, which lowers oxidative damage in the brain in a mouse model for AD, inhibits this peroxidase. The binding of A␤ to heme supports a unifying mechanism by which excessive A␤ induces HD, causes oxidative damage to macromolecules, and depletes specific neurotransmitters. The relevance of the binding of regulatory heme with excessive A␤ for mitochondrial dysfunction and neurotoxicity and other cytopathologies of AD is discussed.curcumin ͉ heme deficiency ͉ mitochondria ͉ regulatory heme ͉ serotonin

show abstract

“…By itself, resveratrol is a potent anti-oxidant which is thought to be responsible for the French Paradox [65]; relating to the low incidence of cardiovascular disease in a French population with high intake of saturated fats. In addition to its demonstrated capacity to inhibit (25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35) fibril formation in vitro [16], resveratrol is also capable of promoting intracellular uptake and degradation of through a proteosome-dependent mechanism [66] and protecting against -induced neurotoxicity [67,68], possibly through activation of the NAD-dependent histone deacetylase, SIRT1 [69]. In the present study using the ELISA-based assay, catechin showed no inhibitory activity during the early stages of oligomerization, whereas rosmarinic acid demonstrated some activity at the highest concentration tested.…”

Section: Discussionmentioning

confidence: 99%

“…While the identity of a number of oligomerization inhibitors have been reported, few address the stoichiometric relationship between inhibitor and peptide, and when calculated, suggest inhibitor to peptide ratios of no greater than one, or in some cases less than one [26][27][28][29][30][31][32]. Presuming that inhibitors must physically bind to peptide to prevent oligomerization [33,34], it is difficult to imagine a mechanism of action where inhibitor to peptide ratios are less than 1.…”

Section: Introductionmentioning

confidence: 99%

A Sensitive Aβ Oligomerization Assay for Identification of Small Molecule Inhibitors

Gonzales¹,

Orlando²

2009

TOBIOTJ

View full text Add to dashboard Cite

Amyloid deposits found in Alzheimer's disease result from aggregation of peptide which leads to loss of synaptic function, chronic microglial activation and cognitive impairment. Because of this, identification of small molecule inhibitors of aggregation as potential therapeutics is a topic of current interest. The majority of inhibitor screening approaches rely on in vitro assays that lack the necessary sensitivity to distinguish low-molecular weight oligomers from larger, more advanced-stage fibrillar structures. Differentiating between these two structures is of vital concern since recent studies indicate that small, early-stage oligomers are the most neurotoxic form of peptide aggregate. To address this limitation, we have explored the adaptability of a recently described ELISA-based assay for discovery of small molecule inhibitors of oligomerization. Results show that this assay is highly sensitive as it is able to quantify oligomers with as little as 80 nM input peptide. In addition, data were obtained re-confirming the function of curcumin as a potent inhibitor of aggregation (IC 50 = 2 μM) and defining its inhibitor:peptide functional stoichiometry. Further examination of other known anti-aggregation compounds showed that this assay is able to discriminate between inhibitors of early-stage, low-molecular weight oligomers and later-stage, high-molecular weight fibrillar structures. These findings indicate that this new ELISA-based assay is capable of identifying novel small molecule inhibitors that function during the initial stages of peptide assembly.

show abstract

Evidence for the Presence of Three Distinct Binding Sites for the Thioflavin T Class of Alzheimer's Disease PET Imaging Agents on β-Amyloid Peptide Fibrils

Cited by 210 publications

References 33 publications

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

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

Amyloid-β peptide binds with heme to form a peroxidase: Relationship to the cytopathologies of Alzheimer’s disease

A Sensitive Aβ Oligomerization Assay for Identification of Small Molecule Inhibitors

Contact Info

Product

Resources

About