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| 4. |
- Zhang, Jun, Dr. 1987-, et al.
(författare)
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Intramolecular Proton and Charge Transfer of Pyrene-based trans-Stilbene Salicylic Acids Applied to Detection of Aggregated Proteins.
- 2018
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Ingår i: ChemPhysChem. - Weinheim, Germany : Wiley-VCH Verlag. - 1439-4235 .- 1439-7641. ; 19:22, s. 3001-3009
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Tidskriftsartikel (refereegranskat)abstract
- Two analogues to the fluorescent amyloid probe 2,5-bis(4'-hydroxy-3'-carboxy-styryl)benzene (X-34) were synthesized based on the trans-stilbene pyrene scaffold (Py1SA and Py2SA). The compounds show strikingly different emission spectra when bound to preformed Aβ1-42 fibrils. This remarkable emission difference is retained when bound to amyloid fibrils of four distinct proteins, suggesting a common binding configuration for each molecule. Density functional theory calculations show that Py1SA is twisted, while Py2SA is more planar. Still, an analysis of the highest occupied molecular orbitals (HOMOs) and lowest unoccupied molecular orbitals (LUMOs) of the two compounds indicates that the degree of electronic coupling between the pyrene and salicylic acid (SA) moieties is larger in Py1SA than in Py2SA. Excited state intramolecular proton transfer (ESIPT) coupled-charge transfer (ICT) was observed for the anionic form in polar solvents. We conclude that ICT properties of trans-stilbene derivatives can be utilized for amyloid probe design with large changes in emission spectra and decay times from analogous chemical structures depending on the detailed physical nature of the binding site.less thanbr /greater than (© 2018 Wiley-VCH Verlag GmbH and Co. KGaA, Weinheim.)
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| 5. |
- Zhang, Jun, Dr. 1987-
(författare)
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Synthesis and characterization of fluorescent stilbene-based probes targeting amyloid fibrils
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Alzheimer’s disease (AD) is characterized by two main protein aggregate hallmarks in the brain: extracellular deposition of the amyloid-β (Aβ) in senile plaques and intracellular neurofibrillary tangles (NFTs) consisting of hyperphosphorylated tau protein. The past decade has seen great progress in the development of imaging probes for the non-invasive detection of Aβ and tau aggregates. Here positron emission tomography (PET), single-photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI), are highly promising technologies for clinical diagnostics. However, as a research tool, optical imaging is superior because it is real-time, sensitive, inexpensive, not radioactive and that it in particular affords high-resolution studies both in vitro and in vivo. Fluorescent probes are especially useful for designing novel binding scaffolds for structure investigations of protein aggregates. This thesis describes design, synthesis and evaluation of a series of fluorescent probes for detection of amyloid fibrils, especially Aβ or tau aggregates in vitro.Firstly, trans-stilbenoid vinylbenzene-1,2-diol with benzene, naphthalene, anthracene, and pyrene are investigated with respect to their photophysical properties free in solution and when bound to amyloid fibrils, including time-resolved fluorescence measurements. It is noted that the extended conjugated systems retained the amyloid targeting properties of the probes and both the anthracene and pyrene moieties extensively enhanced the fluorescence intensity and prolonged lifetimes.Secondly, the synthesis of two molecules, Py1SA and Py2SA, based on pyrene linked to salicylic acid via a trans-stilbene C = C bond is presented. The compounds show strikingly different emission spectra when bound to preformed Aβ1-42 fibrils as well as to fibrils from four other distinct proteins. Additionally, excited state intramolecular proton transfer (ESIPT) coupled-charge transfer (ICT) is observed for the anionic form of the probes in polar solvents. This is likely the reason for the spectral differences of the probes when bound to amyloid fibrils.Moreover, the synthesis of a further development of the Congo red analogue X-34 [2,5-bis(4’-hydroxy-3’-carboxy-styryl) benzene] by rational design and synthesis is described. Full photophysical characterization was performed, including recording absorbance and fluorescence spectra, Stokes shift, quantum yield and fluorescence lifetimes. All ligands displayed high affinity towards recombinant amyloid fibrils of Aβ1-42 and tau as well as selectivity towards the corresponding disease-associated protein aggregates in human post mortem AD tissue.Lastly, the synthesis of a set of 2,1,3-benzothiadiazole (BTD)-based ligands with different conjugated spacers and variable patterns of OH substitutions of bis-styryl-BTD prototypes were developed. Aβ binding affinities (Aβ1-42 and Aβ1-40 fibrils) and the specificity towards Aβ plaques of all ligands were determined. These findings extend the structure to activity relationships of BTD-based ligands for Aβ fibril binding.Throughout the studies in this dissertation, new interesting properties of small molecule fluorescence probes have been discovered and analyzed. This knowledge should facilitate the development of noninvasive probes for early detection of Alzheimer's disease and to distinguish different Aβ fibril polymorphs.
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- Zhang, Jun, Dr. 1987-, et al.
(författare)
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trans-Stilbenoids with Extended Fluorescence Lifetimes for the Characterization of Amyloid Fibrils
- 2017
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Ingår i: ACS Omega. - : American Chemical Society (ACS). - 2470-1343. ; 2:8, s. 4693-4704
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Tidskriftsartikel (refereegranskat)abstract
- It was previously reported that two naphthyl-based trans-stilbene probes, (E)-4-(2-(naphthalen-1-yl)vinyl)benzene-1,2-diol (1) and (E)-4-(2-(naphthalen-2-yl)vinyl)benzene-1,2-diol (3), can bind to both native transthyretin (TTR) and misfolded protofibrillar TTR at physiological concentrations, displaying distinct emission maxima bound to the different conformational states (>100 nm difference). To further explore this amyloid probe scaffold to obtain extended fluorescence lifetimes, two new analogues with expanded aromatic ring systems (anthracene and pyrene), (E)-4-(2-(anthracen-2-yl)vinyl)benzene-1,2-diol (4) and (E)-4-(2-(pyren-2-yl)vinyl)benzene-1,2-diol (5), were synthesized employing the palladium-catalyzed Mizoroki–Heck reaction. (E)-4-Styrylbenzene-1,2-diol (2), 3, 4, and 5 were investigated with respect to their photophysical properties in methanol and when bound to insulin, lysozyme, and Aβ1-42 fibrils, including time-resolved fluorescence measurements. In conclusion, 4 and 5 can bind to both native and fibrillar TTR, becoming highly fluorescent. Compounds 2–5 bind specifically to insulin, lysozyme, and Aβ1-42 fibrils with an apparent fluorescence intensity increase and moderate binding affinities. The average fluorescence lifetimes of the probes bound to Aβ1-42 fibrils are 1.3 ns (2), 1.5 ns (3), 5.7 ns (4), and 29.8 ns (5). In summary, the variable aromatic moieties of the para-positioned trans-stilbenoid vinyl-benzene-1,2-diol with benzene, naphthalene, anthracene, and pyrene showed that the extended conjugated systems retained the amyloid targeting properties of the probes. Furthermore, both the anthracene and pyrene moieties extensively enhanced the fluorescence intensity and prolonged lifetimes. These attractive probe properties should improve amyloid detection and characterization by fluorescence-based techniques.
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| 7. |
- Andersson, D., et al.
(författare)
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Cofactor-induced refolding : Refolding of molten globule carbonic anhydrase induced by Zn(II) and Co(II)
- 2001
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Ingår i: Biochemistry. - : American Chemical Society (ACS). - 0006-2960 .- 1520-4995. ; 40:9, s. 2653-2661
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Tidskriftsartikel (refereegranskat)abstract
- The stability versus unfolding to the molten globule intermediate of bovine carbonic anhydrase II (BCA II) in guanidine hydrochloride (GuHCl) was found to depend on the metal ion cofactor [Zn(II) or Co(II)], and the apoenzyme was observed to be least stable. Therefore, it was possible to find a denaturant concentration (1.2 M GuHCl) at which refolding from the molten globule to the native state could be initiated merely by adding the metal ion to the apo molten globule. Thus, refolding could be performed without changing the concentration of the denaturant. The molten globule intermediate of BCA II could still bind the metal cofactor. Cofactor-effected refolding from the molten globule to the native state can be summarized as follows: (1) initially, the metal ion binds to the molten globule, (2) compaction of the metal-binding site region is then induced by the metal ion binding, (3) a functioning active center is formed, and (4) finally, the native tertiary structure is generated in the outer parts of the protein.
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| 8. |
- Bauer, Susanne
(författare)
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Cell type-specific translatome analysis of mouse models of three genetic neurodegenerative diseases
- 2023
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The burden neurodegenerative diseases place on patients, their loved ones, and the healthcare system is significant, and despite extensive research efforts, there is currently no cure. Since degenerative changes in the brain can begin years before symptoms appear, early intervention is critical. Additionally, neurodegenerative diseases target certain brain regions and neuron types early on. A more comprehensive understanding of the affected cells during the presymptomatic phase is therefore crucial for an effective and targeted intervention. Herein, we isolated, sequenced, and analyzed translatome samples from six neuronal cell types in knock-in mouse models of three monogenic neurodegenerative diseases at a presymptomatic stage: genetic Creutzfeldt-Jakob disease (gCJD), fatal familial insomnia (FFI), and Huntington’s disease (HD). To obtain the translatome samples, we used RiboTag to immunoprecipitate HA-tagged ribosomes with their translating mRNAs from targeted cell types. We analyzed six cell types across two brain regions: cerebral and cerebellar glutamatergic and GABAergic neurons, and cerebral parvalbumin (PV) and somatostatin (SST)-expressing neurons. In the first paper, we focused our analysis on the prion diseases, gCJD (E200K) and FFI (D178N). Here observed a similar response of SST+ neurons, a cell type not previously reported as affected, in both disease models. This was characterized by upregulation of ribosomeassociated genes, and downregulation of cytoskeleton and synapse-associated genes in FFI. Weighted gene co-expression network analysis of SST+ neurons pointed towards the downregulation of mTOR inhibition as a potential mechanism underlying the observed gene expression changes. In the second paper, we analyzed a 129S4-HdhQ200 knock-in mouse model of HD. Histological and behavioral assessment revealed pathological changes in the striatum and cerebellum at 9 months and a later, mild behavioral phenotype. Translatome analysis indicated a surprisingly strong response in reportedly resistant glutamatergic neurons of the cerebellum, marked by upregulation of cell cycle regulators Ccnd1 and chromobox protein genes. In the third paper, we aimed to compare disease-specific responses of PV+ neurons across the three disease models. This analysis revealed a milder response in HD compared to prion disease at comparable disease stages. Functional analysis further indicated PV+ neurons may respond differently in the investigated diseases, showing upregulation of immune response-associated pathways in gCJD, neurodegenerative-disease pathways in FFI, and autophagy in HD. Lastly, the generation of mouse models such as were used in papers I-III requires stable and predictable transgene expression without interfering with the expression of endogenous genes. In the fourth paper, we conducted a pilot study to compare three potential loci, Rpl6, Rpl7, and Eef1a1, as potential safe harbors for transgene integration. Preliminary results indicated that the Rpl6 locus may be best suited for our purposes. Furthermore, this work generated a novel dataset consisting of translatome profiles of six cell types in three neurodegenerative disease models. This provides gene expression data at a previously unavailable level of cellular resolution, especially in prion disease. We believe that this data will serve as a valuable resource for future research and help expand our understanding of the early molecular mechanisms in neurodegenerative disease beyond the scope of this thesis.
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| 9. |
- Elgland, Mathias, 1987-
(författare)
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Synthesis and application of β-configured [18/19F]FDGs : Novel prosthetic CuAAC click chemistry fluoroglycosylation tools for amyloid PET imaging and cancer theranostics
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Positron emission tomography (PET) is a non-invasive imaging method that renders three-dimensional images of tissue that selectively has taken up a radiolabelled organic compound, referred to as a radiotracer. This excellent technique provides clinicians with a tool to monitor disease progression and to evaluate how the patient respond to treatment. The by far most widely employed radiotracer in PET is called 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG), which is often referred to as the golden standard in PET. From a molecular perspective, [18F]FDG is an analogue of glucose where a hydroxyl group has been replaced with a radioactive fluorine atom (18F). It is well known that covalent attachment of carbohydrates (i.e., glycosylation) to biomolecules tend to improve their properties in the body, in terms of; improved pharmacokinetics, increased metabolic stability and faster clearance from blood and other non-specific tissue. It is therefore natural to pursuit the development of a [18F]fluoroglycosylation method where [18F]FDG is chemically conjugated to a ligand with high affinity for a given biological target (e.g., tumors or disease-associated protein aggregates).This thesis describes a novel [18F]fluoroglycosylation method that in a simple and general manner facilitate the conjugation of [18F]FDG to biological ligands using click chemistry. The utility of the developed [18F]fluoroglycosylation method is demonstrated by radiolabelling of curcumin, thus forming a tracer that may be employed for diagnosis of Alzheimer’s disease. Moreover, a set of oligothiophenes were fluoroglycosylated for potential diagnosis of Alzheimer’s disease but also for other much rarer protein misfolding diseases (e.g., Creutzfeldt-Jakob disease and systemic amyloidosis). In addition, the synthesis of a series of 19F-fluoroglycosylated porphyrins is described which exhibited promising properties not only to detect but also to treat melanoma cancer. Lastly, the synthesis of a set of 19F-fluorinated E-stilbenes, structurally based on the antioxidant resveratrol is presented. The E-stilbenes were evaluated for their capacity to spectrally distinguish between native and protofibrillar transthyretin in the pursuit of finding diagnostic markers for the rare but severe disease, transthyretin amyloidosis.
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| 10. |
- Frantz, S.E.A., et al.
(författare)
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Quantum efficiency and two-photon absorption cross-section of conjugated polyelectrolytes used for protein conformation measurements with applications on amyloid structures
- 2007
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Ingår i: Chemical Physics. - : Elsevier BV. - 0301-0104 .- 1873-4421. ; 336:2-3, s. 121-126
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Tidskriftsartikel (refereegranskat)abstract
- Amyloid diseases such as Alzheimer's and spongiform encephalopathies evolve from aggregation of proteins due to misfolding of the protein structure. Early disease handling require sophisticated but yet simple techniques to follow the complex properties of the aggregation process. Conjugated polyelectrolytes (CPEs) have shown promising capabilities acting as optical biological sensors, since they can specifically bind to polypeptides both in solution and in solid phase. The structural changes in biomolecules can be monitored by changes of the optical spectra of the CPEs, both in absorption and emission modes. Notably, the studied CPEs possess multi-photon excitation capability, making them potential for in vivo imaging using laser scanning microscopy. Aggregation of proteins depends on concentration, temperature and pH. The optical effect on the molecular probe in various environments must also be investigated if applied in these environments. Here we present the results of quantum efficiency and two-photon absorption cross-section of three CPEs: POMT, POWT and PTAA in three different pH buffer systems. The extinction coefficient and quantum efficiency were measured. POMT was found to have the highest quantum efficiency being approximately 0.10 at pH 2.0. The two-photon absorption cross-section was measured for POMT and POWT and was found to be more than 18-25 times and 7-11 times that of Fluorescein, respectively. We also show how POMT fluorescence can be used to distinguish conformational differences between amyloid fibrils formed from reduced and non-reduced insulin in spectrally resolved images recorded with a laser scanning microscope using both one- and two-photon excitation. © 2007 Elsevier B.V. All rights reserved.
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