| 1. |
- Boutajangout, Allal, et al.
(författare)
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Affibody-Mediated Sequestration of Amyloid beta Demonstrates Preventive Efficacy in a Transgenic Alzheimer's Disease Mouse Model
- 2019
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Ingår i: Frontiers in Aging Neuroscience. - : Frontiers Media S.A.. - 1663-4365. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- Different strategies for treatment and prevention of Alzheimer's disease (AD) are currently under investigation, including passive immunization with anti-amyloid beta (anti-A beta) monoclonal antibodies (mAbs). Here, we investigate the therapeutic potential of a novel type of A beta-targeting agent based on an affibody molecule with fundamentally different properties to mAbs. We generated a therapeutic candidate, denoted Z(SYM73)-albumin-binding domain (ABD; 16.8 kDa), by genetic linkage of the dimeric Z(SYM73) affibody for sequestering of monomeric A beta-peptides and an ABD for extension of its in vivo half-life. Amyloid precursor protein (APP)/PS1 transgenic AD mice were administered with Z(SYM73)-ABD, followed by behavioral examination and immunohistochemistry. Results demonstrated rescued cognitive functions and significantly lower amyloid burden in the treated animals compared to controls. No toxicological symptoms or immunology-related side-effects were observed. To our knowledge, this is the first reported in vivo investigation of a systemically delivered scaffold protein against monomeric A beta, demonstrating a therapeutic potential for prevention of AD.
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| 2. |
- Härd, Torleif
(författare)
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A beta-Hairpin-Binding Protein for Three Different Disease-Related Amyloidogenic Proteins
- 2015
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Ingår i: ChemBioChem. - : Wiley. - 1439-4227 .- 1439-7633. ; 16, s. 411-414
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Tidskriftsartikel (refereegranskat)abstract
- Amyloidogenic proteins share a propensity to convert to the beta-structure-rich amyloid state that is associated with the progression of several protein-misfolding disorders. Here we show that a single engineered beta-hairpin-binding protein, the beta-wrapin AS10, binds monomers of three different amyloidogenic proteins, that is, amyloid-beta peptide, alpha-synuclein, and islet amyloid polypeptide, with sub-micromolar affinity. AS10 binding inhibits the aggregation and toxicity of all three proteins. The results demonstrate common conformational preferences and related binding sites in a subset of the amyloidogenic proteins. These commonalities enable the generation of multispecific monomer-binding agents.
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| 3. |
- Härd, Torleif
(författare)
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Contact between the β1 and β2 Segments of α-Synuclein that Inhibits Amyloid Formation
- 2015
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Ingår i: Angewandte Chemie International Edition. - : Wiley. - 1433-7851 .- 1521-3773. ; 54, s. 8837-8840
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Tidskriftsartikel (refereegranskat)abstract
- Conversion of the intrinsically disordered protein α-synuclein (α-syn) into amyloid aggregates is a key process in Parkinson’s disease. The sequence region 35–59 contains β-strand segments β1 and β2 of α-syn amyloid fibril models and most disease-related mutations. β1 and β2 frequently engage in transient interactions in monomeric α-syn. The consequences of β1–β2 contacts are evaluated by disulfide engineering, biophysical techniques, and cell viability assays. The double-cysteine mutant α-synCC, with a disulfide linking β1 and β2, is aggregation-incompetent and inhibits aggregation and toxicity of wild-type α-syn. We show that α-syn delays the aggregation of amyloid-β peptide and islet amyloid polypeptide involved in Alzheimer’s disease and type 2 diabetes, an effect enhanced in the α-synCC mutant. Tertiary interactions in the β1–β2 region of α-syn interfere with the nucleation of amyloid formation, suggesting promotion of such interactions as a potential therapeutic approach.
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| 4. |
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| 5. |
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| 6. |
- Lendel, Christofer, et al.
(författare)
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Combined Solution- and Magic Angle Spinning NMR Reveals Regions of Distinct Dynamics in Amyloid beta Protofibrils
- 2016
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Ingår i: Chemistryselect. - : Wiley. - 2365-6549. ; 1:18, s. 5850-5853
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Tidskriftsartikel (refereegranskat)abstract
- Solid-state magic angle spinning (MAS) NMR has emerged as an important tool for investigations of protein aggregates and amyloid fibrils, which are not accessible for solution NMR experiments. We recently presented a structural model for amyloid beta (A beta) protofibrils based on MAS-NMR data. The absence of resonances for the N-terminus of A beta in this dataset suggested that it might be disordered and more dynamic than the structural core. We here provide evidence for a distinct dynamic regime in the N-terminal part of the peptide and show that the structural characteristics of this region can be elucidated using C-13-detected solution NMR. The results shed more light on the structural properties of pre-fibrillar A beta species and demonstrate the potential of combining MAS and solution NMR experiments for the characterization of structure and dynamics of complex protein assemblies.
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| 7. |
- Lindberg, Hanna, et al.
(författare)
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A truncated and dimeric format of an Affibody library on bacteria enables FACS-mediated isolation of amyloid-beta aggregation inhibitors with subnanomolar affinity
- 2015
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Ingår i: Biotechnology Journal. - : Wiley. - 1860-6768 .- 1860-7314. ; 10:11, s. 1707-1718
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Tidskriftsartikel (refereegranskat)abstract
- The amyloid hypothesis suggests that accumulation of amyloid β (Aβ) peptides in the brain is involved in development of Alzheimer's disease. We previously generated a small dimeric affinity protein that inhibited Aβ aggregation by sequestering the aggregation prone parts of the peptide. The affinity protein is originally based on the Affibody scaffold, but is evolved to a distinct interaction mechanism involving complex structural rearrangement in both the Aβ peptide and the affinity proteins upon binding. The aim of this study was to decrease the size of the dimeric affinity protein and significantly improve its affinity for the Aβ peptide to increase its potential as a future therapeutic agent. We combined a rational design approach with combinatorial protein engineering to generate two different affinity maturation libraries. The libraries were displayed on staphylococcal cells and high-affinity Aβ-binding molecules were isolated using flow-cytometric sorting. The best performing candidate binds Aβ with a KD value of around 300 pM, corresponding to a 50-fold improvement in affinity relative to the first-generation binder. The new dimeric Affibody molecule was shown to capture Aβ1-42 peptides from spiked E. coli lysate. Altogether, our results demonstrate successful engineering of this complex binder for increased affinity to the Aβ peptide.
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| 8. |
- Pakharukova, Natalia, et al.
(författare)
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Structural basis for Myf and Psa fimbriae-mediated tropism of pathogenic strains of Yersinia for host tissues.
- 2016
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Ingår i: Molecular microbiology. - : Wiley. - 1365-2958 .- 0950-382X. ; 102:4, s. 593-610
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Tidskriftsartikel (refereegranskat)abstract
- Three pathogenic species of the genus Yersinia assemble adhesive fimbriae via the FGL-chaperone/usher pathway. Closely related Y. pestis and Y. pseudotuberculosis elaborate the pH6 antigen (Psa), which mediates bacterial attachment to alveolar cells of the lung. Y. enterocolitica, instead, assembles the homologous fimbriae Myf of unknown function. Here, we discovered that Myf, like Psa, specifically recognizes β1-3- or β1-4-linked galactose in glycosphingolipids, but completely lacks affinity for phosphatidylcholine, the main receptor for Psa in alveolar cells. The crystal structure of a subunit of Psa (PsaA) complexed with choline together with mutagenesis experiments revealed that PsaA has four phosphatidylcholine binding pockets that enable super-high-avidity binding of Psa-fibres to cell membranes. The pockets are arranged as six tyrosine residues, which are all missing in the MyfA subunit of Myf. Conversely, the crystal structure of the MyfA-galactose complex revealed that the galactose-binding site is more extended in MyfA, enabling tighter binding to lactosyl moieties. Our results suggest that during evolution, Psa has acquired a tyrosine-rich surface that enables it to bind to phosphatidylcholine and mediate adhesion of Y. pestis/pseudotuberculosis to alveolar cells, whereas Myf has specialized as a carbohydrate-binding adhesin, facilitating the attachment of Y. enterocolitica to intestinal cells.
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| 9. |
- Rahman, Mahafuzur, et al.
(författare)
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Binding of Human Proteins to Amyloid-beta Protofibrils
- 2015
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Ingår i: ACS Chemical Biology. - : American Chemical Society (ACS). - 1554-8929 .- 1554-8937. ; 10:3, s. 766-774
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Tidskriftsartikel (refereegranskat)abstract
- The progressive neurodegeneration in Alzheimers disease is believed to be linked to the presence of prefibrillar aggregates of the amyloid-beta (A beta) peptide in the brain. The exact role of these aggregates in the disease pathology is, however, still an open question. Any mechanism by which oligomeric A beta may cause damage to neuronal cells must, in one way or another, involve interactions with other molecules. Here, we identify proteins in human serum and cerebrospinal fluid that bind to stable protofibrils formed by an engineered variant of A beta 42 (A beta(42CC)). We find that the protofibrils attract a substantial number of protein binding partners. Many of the 101 identified proteins are involved in lipid transport and metabolism, the complement system, or in hemostasis. Binding of representative proteins from all of these groups with micromolar affinity was confirmed using surface plasmon resonance. In addition, binding of apolipoprotein E to the protofibrils with nanomolar affinity was demonstrated. We also find that aggregation of A beta enhances protein binding, as lower amounts of proteins bind monomeric A beta. Proteins that bind to A beta protofibrils might contribute to biological effects in which these aggregates are involved. Our results therefore suggest that an improved understanding of the mechanisms by which A beta causes cytotoxicity and neurodegeneration might be gained from studies carried out in biologically relevant matrices in which A beta-binding proteins are present.
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| 10. |
- Rahman, Mahafuzur, et al.
(författare)
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Protofibrillar and Fibrillar Amyloid-beta Binding Proteins in Cerebrospinal Fluid
- 2018
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Ingår i: Journal of Alzheimer's Disease. - 1387-2877 .- 1875-8908. ; 66:3, s. 1053-1064
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Tidskriftsartikel (refereegranskat)abstract
- Aggregation and deposition of misfolded amyloid-beta (A beta) peptide in the brain is central to Alzheimer's disease (AD). Oligomeric, protofibrillar, and fibrillar forms of A beta are believed to be neurotoxic and cause neurodegeneration in AD, but the toxicity mechanisms are not well understood and may involve A beta-interacting molecular partners. In a previous study, we identified potential A beta(42) protofibrillar-binding proteins in serum and cerebrospinal fluid (CSF) using an engineered version of A beta(42) (A beta 42CC) that forms protofibrils, but not fibrils. Here we studied binding of proteins to A beta(42) fibrils in AD and non-AD CSF and compared these with protofibrillar A beta 42CC-binding partners. A beta(42) fibrils sequestered 2.4-fold more proteins than A beta 42CC protofibrils. Proteins with selective binding to fibrillar aggregates with low nanomolar affinity were identified. We also found that protofibrillar and fibrillar A beta-binding proteins represent distinct functional categories. A beta 42CC protofibrils triggered interactions with proteins involved in catalytic activities, like transferases and oxidoreductases, while A beta(42) fibrils were more likely involved in binding to proteoglycans, growth factors and neuron-associated proteins, e.g., neurexin-1, -2, and -3. Interestingly, 10 brain-enriched proteins were identified among the fibril-binding proteins, while protofibril-extracted proteins had more general expression patterns. Both types of A beta aggregates bound several extracellular proteins. Additionally, we list a set of CSF proteins that might have potential to discriminate between AD and non-AD CSF samples. The results may be of relevance both for biomarker studies and for studies of A beta-related toxicity mechanisms.
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