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  • 1.
    Almstedt, Karin
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Amyloid fibrils of human prion protein are spun and woven from morphologically disordered aggregates2009In: Prion, ISSN 1933-6896, Vol. 3, no 4, p. 224-235Article in journal (Refereed)
    Abstract [en]

    Propagation and infectivity of prions in human prionopathies are likely associated with conversion of the mainly α-helical human prion protein, HuPrP, into an aggregated form with amyloid-like properties. Previous reports on efficient conversion of recombinant HuPrP have used mild to harsh denaturing conditions to generate amyloid fibrils in vitro. Herein we report on the in vitro conversion of four forms of truncated HuPrP (sequences 90-231 and 121-231 with and without an N-terminal hexa histidine tag) into amyloid-like fibrils within a few hours by using a protocol (phosphate buffered saline solutions at neutral pH with intense agitation) close to physiological conditions. The conversion process monitored by thioflavin T, ThT, revealed a three stage process with lag, growth and equilibrium phases. Seeding with preformed fibrils shortened the lag phase demonstrating the classic nucleated polymerization mechanism for the reaction. Interestingly, comparing thioflavin T kinetics with solubility and turbidity kinetics it was found that the protein initially formed non-thioflavionophilic, morphologically disordered aggregates that over time matured into amyloid fibrils. By transmission electron microscopy and by fluorescence microscopy of aggregates stained with luminescent conjugated polythiophenes (LCPs); we demonstrated that HuPrP undergoes a conformational conversion where spun and woven fibrils protruded from morphologically disordered aggregates. The initial aggregation functioned as a kinetic trap that decelerated nucleation into a fibrillation competent nucleus, but at the same time without aggregation there was no onset of amyloid fibril formation. The agitation, which was necessary for fibril formation to be induced, transiently exposes the protein to the air-water interface suggests a hitherto largely unexplored denaturing environment for prion conversion.

  • 2.
    Bagheri, Maryam
    et al.
    Ilam University of Medical Science, Iran .
    Rezakhani, Arjang
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Turkina, Maria
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Health Sciences.
    Roghani, Mehrdad
    Shahed University, Iran .
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Mohseni, Simin
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Health Sciences.
    Amyloid Beta1-40-Induced Astrogliosis and the Effect of Genistein Treatment in Rat: A Three-Dimensional Confocal Morphometric and Proteomic Study2013In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, no 10Article in journal (Refereed)
    Abstract [en]

    Astrocytes are highly involved in regulation and homeostasis of the extracellular environment in the healthy brain. In pathological conditions, these cells play a major role in the inflammatory response seen in CNS tissues, which is called reactive astrogliosis and includes hypertrophy and proliferation of astrocytes. Here, we performed 3D confocal microscopy to evaluate the morphological response of reactive astrocytes positive for glial fibrillary acidic protein (GFAP) in rats, to the presence of Aβ1–40 in the rat brain before and after treatment with genistein. In 50 astrocytes per animal, we measured the volume and surface area for the nucleus, cell body, the entire cell, the tissue covered by single astrocytes and quantified the number and length of branches, the density of the astrocytes and the intensity of GFAP immunoreactivity. Injecting Aβ1–40 into the brain of rats caused astrogliosis indicated by increased values for all measured parameters. Mass spectrometric analysis of hippocampal tissue in Aβ1–40-injected brain showed decreased amounts of tubulins, enolases and myelin basic protein, and increased amounts of dihydropyrimidinase-related protein 2. In Aβ1–40-injected rats pretreated with genistein, GFAP intensity was decreased to the sham-operated group level, and Aβ1–40-induced astrogliosis was significantly ameliorated.

  • 3.
    Ellingsen, Pal Gunnar
    et al.
    Norwegian University of Science and Technology, Norway .
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Reitan, Nina Kristine
    Norwegian University of Science and Technology, Norway .
    Lindgren, Mikael
    Norwegian University of Science and Technology, Norway .
    Spectral correlation analysis of Amyloid beta plaque inhomogeneity from double staining experiments2013In: Journal of Biomedical Optics, ISSN 1083-3668, E-ISSN 1560-2281, Vol. 18, no 10Article in journal (Refereed)
    Abstract [en]

    A spectral correlation algorithm for the analysis of hyperspectral fluorescence images is proposed by Ellingsen et al. [J. Biomed. Opt. 18, 020501 (2013)]. Here, it is applied to the analysis of double-stained A beta amyloid plaques being related to the Alzheimers disease (AD). Sections of APP/PS1 AD mice model brains are double stained with luminescent-conjugated oligothiophenes, known to bind to amyloid protein deposits. Hyperspectral fluorescence images of the brain sections are recorded and by applying the correlation algorithm the spectral inhomogeneity of the double-stained samples is mapped in terms of radial distribution and spectral content. To further investigate the progression of A beta amyloid plaque formation, 19 AD mice of different ages up to 23 months are characterized, enabling a statistical analysis of the plaque heterogeneity. In accordance with recent findings by Nystrom et al. [ACS Chem. Biol. 8, 1128-1133 (2013)], the spectral distribution within A beta plaques is found to vary with age throughout the lifespan of the mouse. With the new correlation algorithm, it is possible to quantify the spectral abundance of the two stains depending on the relative distance from the plaque center and mouse age. Thus, we demonstrate the use of the correlation analysis approach in double-staining experiments and how it is possible to relate these to structural/spectral changes in biological samples. (C) The Authors. Published by SPIE under a Creative Commons Attribution 3.0 Unported License. Distribution or reproduction of this work in whole or in part requires full attribution of the original publication, including its DOI.

  • 4.
    Hammarström, Per
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Porcine prion protein amyloid2015In: Prion, ISSN 1933-6896, E-ISSN 1933-690X, Vol. 9, no 4, p. 266-277Article in journal (Refereed)
    Abstract [en]

    Mammalian prions are composed of misfolded aggregated prion protein (PrP) with amyloid-like features. Prions are zoonotic disease agents that infect a wide variety of mammalian species including humans. Mammals and by-products thereof which are frequently encountered in daily life are most important for human health. It is established that bovine prions (BSE) can infect humans while there is no such evidence for any other prion susceptible species in the human food chain (sheep, goat, elk, deer) and largely prion resistant species (pig) or susceptible and resistant pets (cat and dogs, respectively). PrPs from these species have been characterized using biochemistry, biophysics and neurobiology. Recently we studied PrPs from several mammals in vitro and found evidence for generic amyloidogenicity as well as cross-seeding fibril formation activity of all PrPs on the human PrP sequence regardless if the original species was resistant or susceptible to prion disease. Porcine PrP amyloidogenicity was among the studied. Experimentally inoculated pigs as well as transgenic mouse lines overexpressing porcine PrP have, in the past, been used to investigate the possibility of prion transmission in pigs. The pig is a species with extraordinarily wide use within human daily life with over a billion pigs harvested for human consumption each year. Here we discuss the possibility that the largely prion disease resistant pig can be a clinically silent carrier of replicating prions.

  • 5.
    Hammarström, Per
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Simon, Rozalyn
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, Faculty of Science & Engineering.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Konradsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Åslund, Andreas
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    A Fluorescent Pentameric Thiophene Derivative Detects in Vitro-Formed Prefibrillar Protein Aggregates2010In: BIOCHEMISTRY, ISSN 0006-2960, Vol. 49, no 32, p. 6838-6845Article in journal (Refereed)
    Abstract [en]

    Protein aggregation is associated with a wide range of diseases, and molecular probes that are able to detect a diversity of misfolded protein assemblies are of great importance. The identification of prefibrillar states preceding the formation of well-defined amyloid fibrils is of particular interest both because of their likely role in the mechanism of fibril formation and because of the growing awareness that these species are likely to play a critical role in the pathogenesis of protein deposition diseases. Herein, we explore the use of an anionic oligothiophene derivative, p-FTAA, for detection of prefibrillar protein aggregates during in vitro fibrillation of three different amyloidogenic proteins (insulin, lysozyme, and prion protein). p-FTAA generally detected prefibrillar protein aggregates that could not be detected by thioflavine T fluorescence and in addition showed high fluorescence when bound to mature fibrils. Second, the kinetics of protein aggregation or the formation of amyloid fibrils of insulin was not extensively influenced by the presence of various concentrations of p-FTAA. These results establish the use of p-FTAA as an additional tool for studying the process of protein aggregation.

  • 6.
    Herrmann, Uli S.
    et al.
    University of Zurich Hospital, Switzerland.
    Schuetz, Anne K.
    ETH, Switzerland.
    Shirani, Hamid
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Huang, Danzhi
    University of Zurich, Switzerland.
    Saban, Dino
    University of Zurich Hospital, Switzerland.
    Nuvolone, Mario
    University of Zurich Hospital, Switzerland.
    Li, Bei
    University of Zurich Hospital, Switzerland.
    Ballmer, Boris
    University of Zurich Hospital, Switzerland.
    Åslund, Andreas
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Mason, Jeffrey
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Rushing, Elisabeth
    University of Zurich Hospital, Switzerland.
    Budka, Herbert
    University of Zurich Hospital, Switzerland.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Boeckmann, Anja
    University of Lyon 1, France.
    Caflisch, Amedeo
    University of Zurich, Switzerland.
    Meier, Beat H.
    ETH, Switzerland.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Hornemann, Simone
    University of Zurich Hospital, Switzerland.
    Aguzzi, Adriano
    University of Zurich Hospital, Switzerland.
    Structure-based drug design identifies polythiophenes as antiprion compounds2015In: Science Translational Medicine, ISSN 1946-6234, E-ISSN 1946-6242, Vol. 7, no 299, p. 299ra123-Article in journal (Refereed)
    Abstract [en]

    Prions cause transmissible spongiform encephalopathies for which no treatment exists. Prions consist of PrPSc, a misfolded and aggregated form of the cellular prion protein (PrPC). We explore the antiprion properties of luminescent conjugated polythiophenes (LCPs) that bind and stabilize ordered protein aggregates. By administering a library of structurally diverse LCPs to the brains of prion-infected mice via osmotic minipumps, we found that antiprion activity required a minimum of five thiophene rings bearing regularly spaced carboxyl side groups. Solid-state nuclear magnetic resonance analyses and molecular dynamics simulations revealed that anionic side chains interacted with complementary, regularly spaced cationic amyloid residues of model prions. These findings allowed us to extract structural rules governing the interaction between LCPs and protein aggregates, which we then used to design a new set of LCPs with optimized binding. The new set of LCPs showed robust prophylactic and therapeutic potency in prion-infected mice, with the lead compound extending survival by greater than80% and showing activity against both mouse and hamster prions as well as efficacy upon intraperitoneal administration into mice. These results demonstrate the feasibility of targeted chemical design of compounds that may be useful for treating diseases of aberrant protein aggregation such as prion disease.

  • 7.
    Klingstedt, Therése
    et al.
    Linköping University, Faculty of Science & Engineering. Linköping University, Department of Physics, Chemistry and Biology, Chemistry.
    Shirani, Hamid
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Mahler, Jasmin
    University of Tubingen, Germany; German Centre Neurodegenerat Disease, Germany.
    Wegenast-Braun, Bettina M.
    University of Tubingen, Germany; German Centre Neurodegenerat Disease, Germany.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Goedert, Michel
    MRC, England.
    Jucker, Mathias
    University of Tubingen, Germany; German Centre Neurodegenerat Disease, Germany.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Distinct Spacing Between Anionic Groups: An Essential Chemical Determinant for Achieving Thiophene-Based Ligands to Distinguish Beta-Amyloid or Tau Polymorphic Aggregates2015In: Chemistry - A European Journal, ISSN 0947-6539, E-ISSN 1521-3765, Vol. 21, no 25, p. 9072-9082Article in journal (Refereed)
    Abstract [en]

    The accumulation of protein aggregates is associated with many devastating neurodegenerative diseases and the existence of distinct aggregated morphotypes has been suggested to explain the heterogeneous phenotype reported for these diseases. Thus, the development of molecular probes able to distinguish such morphotypes is essential. We report an anionic tetrameric oligothiophene compound that can be utilized for spectral assignment of different morphotypes of -amyloid or tau aggregates present in transgenic mice at distinct ages. The ability of the ligand to spectrally distinguish between the aggregated morphotypes was reduced when the spacing between the anionic substituents along the conjugated thiophene backbone was altered, which verified that specific molecular interactions between the ligand and the protein aggregate are necessary to detect aggregate polymorphism. Our findings provide the structural and functional basis for the development of new fluorescent ligands that can distinguish between different morphotypes of protein aggregates.

  • 8.
    Klingstedt, Therése
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Åslund, Andreas
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Simon, Rozalyn
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Johansson, Leif B. G.
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Mason, Jeffrey
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Protein Science. Linköping University, The Institute of Technology.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Protein Science. Linköping University, The Institute of Technology.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Synthesis of a library of oligothiophenes and their utilization as fluorescent ligands for spectral assignment of protein aggregates2011In: Organic and biomolecular chemistry, ISSN 1477-0520, E-ISSN 1477-0539, Vol. 9, no 24, p. 8356-8370Article in journal (Refereed)
    Abstract [en]

    Molecular probes for selective identification of protein aggregates are important to advance our understanding of the molecular pathogenesis underlying protein aggregation diseases. Here we report the chemical design of a library of anionic luminescent conjugated oligothiophenes (LCOs), which can be utilized as ligands for detection of protein aggregates. Certain molecular requirements were shown to be necessary for detecting (i) early non-thioflavinophilic protein assemblies of A beta 1-42 and insulin preceding the formation of amyloid fibrils and (ii) for obtaining distinct spectral signatures of the two main pathological hallmarks observed in human Alzheimers diease brain tissue (A beta plaques and neurofibrillary tangles). Our findings suggest that a superior anionic LCO-based ligand should have a backbone consisting of five to seven thiophene units and carboxyl groups extending the conjugated thiophene backbone. Such LCOs will be highly useful for studying the underlying molecular events of protein aggregation diseases and could also be utilized for the development of novel diagnostic tools for these diseases.

  • 9.
    Margalith, Ilan
    et al.
    University of Zurich Hospital, Switzerland .
    Suter, Carlo
    University of Zurich Hospital, Switzerland .
    Ballmer, Boris
    University of Zurich Hospital, Switzerland .
    Schwarz, Petra
    University of Zurich Hospital, Switzerland .
    Tiberi, Cinzia
    University of Zurich Hospital, Switzerland .
    Sonati, Tiziana
    University of Zurich Hospital, Switzerland .
    Falsig, Jeppe
    University of Zurich Hospital, Switzerland .
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Åslund, Andreas
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Yam, Alice
    Novartis Diagnost, USA .
    Whitters, Eric
    Novartis Diagnost, USA .
    Hornemann, Simone
    University of Zurich Hospital, Switzerland .
    Aguzzi, Adriano
    University of Zurich Hospital, Switzerland .
    Polythiophenes Inhibit Prion Propagation by Stabilizing Prion Protein (PrP) Aggregates2012In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 287, no 23, p. 18872-18887Article in journal (Refereed)
    Abstract [en]

    Luminescent conjugated polymers (LCPs) interact with ordered protein aggregates and sensitively detect amyloids of many different proteins, suggesting that they may possess antiprion properties. Here, we show that a variety of anionic, cationic, and zwitterionic LCPs reduced the infectivity of prion-containing brain homogenates and of prion-infected cerebellar organotypic cultured slices and decreased the amount of scrapie isoform of PrPC (PrPSc) oligomers that could be captured in an avidity assay. Paradoxically, treatment enhanced the resistance of PrPSc to proteolysis, triggered the compaction, and enhanced the resistance to proteolysis of recombinant mouse PrP(23-231) fibers. These results suggest that LCPs act as antiprion agents by transitioning PrP aggregates into structures with reduced frangibility. Moreover, ELISA on cerebellar organotypic cultured slices and in vitro conversion assays with mouse PrP(23-231) indicated that poly(thiophene-3-acetic acid) may additionally interfere with the generation of PrPSc by stabilizing the conformation of PrPC or of a transition intermediate. Therefore, LCPs represent a novel class of antiprion agents whose mode of action appears to rely on hyperstabilization, rather than destabilization, of PrPSc deposits.

  • 10.
    Maria Psonka-Antonczyk, Katarzyna
    et al.
    Norwegian University of Science and Technology.
    Duboisset, Julien
    Norwegian University of Science and Technology.
    Torger Stokke, Bjorn
    Norwegian University of Science and Technology.
    Zako, Tamotsu
    Riken Institute Phys and Chemistry Research.
    Kobayashi, Takahiro
    Riken Institute Phys and Chemistry Research.
    Maeda, Mizuo
    Riken Institute Phys and Chemistry Research.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Mason, Jeffrey
    Linköping University, Department of Physics, Chemistry and Biology, Protein Science. Linköping University, Faculty of Science & Engineering.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Lindgren, Mikael
    Norwegian University of Science and Technology.
    Nanoscopic and Photonic Ultrastructural Characterization of Two Distinct Insulin Amyloid States2012In: INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES, ISSN 1661-6596, Vol. 13, no 2, p. 1461-1480Article in journal (Refereed)
    Abstract [en]

    Two different conformational isoforms or amyloid strains of insulin with different cytotoxic capacity have been described previously. Herein these filamentous and fibrillar amyloid states of insulin were investigated using biophysical and spectroscopic techniques in combination with luminescent conjugated oligothiophenes (LCO). This new class of fluorescent probes has a well defined molecular structure with a distinct number of thiophene units that can adopt different dihedral angles depending on its binding site to an amyloid structure. Based on data from surface charge, hydrophobicity, fluorescence spectroscopy and imaging, along with atomic force microscopy (AFM), we deduce the ultrastructure and fluorescent properties of LCO stained insulin fibrils and filaments. Combined total internal reflection fluorescence microscopy (TIRFM) and AFM revealed rigid linear fibrous assemblies of fibrils whereas filaments showed a short curvilinear morphology which assemble into cloudy deposits. All studied LCOs bound to the filaments afforded more blue-shifted excitation and emission spectra in contrast to those corresponding to the fibril indicating a different LCO binding site, which was also supported by less efficient hydrophobic probe binding. Taken together, the multi-tool approach used here indicates the power of ultrastructure identification applying AFM together with LCO fluorescence interrogation, including TIRFM, to resolve structural differences between amyloid states.

  • 11.
    Mishra, Rajesh
    et al.
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Sörgjerd, Karin
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Nyström, Sofie
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Nordigården, Amanda
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Biomedicine and Surgery, Division of cell biology.
    Chiu, Yu-Jui
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology.
    Hammarström, Per
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Lysozyme Amyloidogenesis Is Accelerated by Specific Nicking and Fragmentation but Decelerated by Intact Protein Binding and Conversion2007In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 366, no 3, p. 1029-1044Article in journal (Refereed)
    Abstract [en]

    We have revisited the well-studied heat and acidic amyloid fibril formation pathway (pH 1.6, 65 °C) of hen egg-white lysozyme (HEWL) to map the barriers of the misfolding and amyloidogenesis pathways. A comprehensive kinetic mechanism is presented where all steps involving protein hydrolysis, fragmentation, assembly and conversion into amyloid fibrils are accounted for. Amyloid fibril formation of lysozyme has multiple kinetic barriers. First, HEWL unfolds within minutes, followed by irreversible steps of partial acid hydrolysis affording a large amount of nicked HEWL, the 49-101 amyloidogenic fragment and a variety of other species over 5-40 h. Fragmentation forming the 49-101 fragment is a requirement for efficient amyloid fibril formation, indicating that it forms the rate-determining nucleus. Nicked full-length HEWL is recruited efficiently into amyloid fibrils in the fibril growth phase or using mature fibrils as seeds, which abolished the lag phase completely. Mature amyloid fibrils of HEWL are composed mainly of nicked HEWL in the early equilibrium phase but go through a "fibril shaving" process, affording fibrils composed of the 49-101 fragment and 53-101 fragment during more extensive maturation (incubation for longer than ten days). Seeding of the amyloid fibril formation process using sonicated mature amyloid fibrils accelerates the fibril formation process efficiently, however, addition of intact full-length lysozyme at the end of the lag phase slows the rate of amyloidogenesis. The intact full-length protein, in contrast to nicked lysozyme, slows fibril formation due to its slow conversion into the amyloid fold, probably due to inclusion of the non-amyloidogenic 1-48/102-129 portion of HEWL in the fibrils, which can function as a "molecular bumper" stalling further growth. © 2006 Elsevier Ltd. All rights reserved.

  • 12.
    Nilsson, Peter
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Åslund, Andreas
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry.
    Berg, Ina
    Nyström, Sofie
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Konradsson, Peter
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry.
    Herland, Anna
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biomolecular and Organic Electronics.
    Inganäs, Olle
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biomolecular and Organic Electronics.
    Stabo-Eeg, Frantz
    Lindgren, Mikael
    Westermark, Gunilla
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Biomedicine and Surgery, Division of cell biology.
    Lannfelt, Lars
    Nilsson, Lars N G
    Hammarström, Per
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Imaging distinct conformational states of amyloid-β fibrils in Alzheimer's disease using novel luminescent probes2007In: ACS Chemical Biology, ISSN 1554-8929, Vol. 2, no 8, p. 553-560Article in journal (Refereed)
    Abstract [en]

    Using luminescent conjugated polyelectrolyte probes (LCPs), we demonstrate the possibility to distinguish amyloid-β 1-42 peptide (Aβ1-42) fibril conformations, by analyzing in vitro generated amyloid fibrils of Aβ1-42 formed under quiescent and agitated conditions. LCPs were then shown to resolve such conformational heterogeneity of amyloid deposits in vivo. A diversity of amyloid deposits depending upon morphology and anatomic location was illustrated with LCPs in frozen ex vivo brain sections from a transgenic mouse model (tg-APPswe) of Alzheimer's disease. Comparative LCP fluorescence showed that compact-core plaques of amyloid β precursor protein transgenic mice were composed of rigid dense amyloid. A more abundant form of amyloid plaque displayed morphology of a compact center with a protruding diffuse exterior. Surprisingly, the compact center of these plaques showed disordered conformations of the fibrils, and the exterior was composed of rigid amyloid protruding from the disordered center. This type of plaque appears to grow from more loosely assembled regions toward solidified amyloid tentacles. This work demonstrates how application of LCPs can prove helpful to monitor aggregate structure of in vivo formed amyloid deposits such as architecture, maturity, and origin.

  • 13.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Early events in disease associated protein misfolding2009Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The scope of this thesis is to unravel some of the mysteries concerning events takingplace early in the amyloid cascade. In vitro studies of early misfolded states ofamyloidogenic proteins are important since the use of recombinant proteins allow us to monitor slight changes in environmental conditions as well as in amino acid composition and thereby illuminate the problem at near atomic resolution.

    Human prion protein (HuPrP) (associated with e.g. Creutzfeldt-Jakob disease) andthe Aβ1-42 peptide (associated with Alzheimer’s disease) recombinantly expressed in Escherichia coli have been used as model systems for these studies.

    A new protocol for amyloid fibril formation of human prion protein under native conditions was developed. This revealed an unusual pathway of conformational conversion from early formed disordered aggregates that later matured into amyloidfibrils.

    The polymorphism 129M/V in HuPrP has a large impact on susceptibility both to sporadic and infectious prion diseases. Some features of this polymorphism havebeen elucidated, employing a mutational study in position 129 (M, A, L, V, P, M, W,E, and K). These investigations have rendered new knowledge about the impact ofsize, charge and β-carbon branching in position 129 upon early intermolecular interactions and the effects of fibril seeding.

    Investigations of the interactions between different assembly forms of HuPrP and components of the innate immune system revealed that both native, oligomeric and fibrillar forms of HuPrP activate both the classical and alternative pathways of the Complement System. Most efficient activation is achieved upon binding of oligomeric HuPrP to the complement component C1q.

    We have developed a system for recombinant expression of human A,1-42. The monomeric peptides are assembled into various sized soluble oligomers (trimer, hexamer, nonamer, dodecamer). The oligomeric forms were stable in 8 M urea, 6 MGuHCl and SDS suggesting that these were covalently cross-linked. Some mechanistic features in the assembly process have been investigated and we have shown that cupric ions facilitates formation of stable oligomers in our system.

    List of papers
    1. Amyloid fibrils of human prion protein are spun and woven from morphologically disordered aggregates
    Open this publication in new window or tab >>Amyloid fibrils of human prion protein are spun and woven from morphologically disordered aggregates
    2009 (English)In: Prion, ISSN 1933-6896, Vol. 3, no 4, p. 224-235Article in journal (Refereed) Published
    Abstract [en]

    Propagation and infectivity of prions in human prionopathies are likely associated with conversion of the mainly α-helical human prion protein, HuPrP, into an aggregated form with amyloid-like properties. Previous reports on efficient conversion of recombinant HuPrP have used mild to harsh denaturing conditions to generate amyloid fibrils in vitro. Herein we report on the in vitro conversion of four forms of truncated HuPrP (sequences 90-231 and 121-231 with and without an N-terminal hexa histidine tag) into amyloid-like fibrils within a few hours by using a protocol (phosphate buffered saline solutions at neutral pH with intense agitation) close to physiological conditions. The conversion process monitored by thioflavin T, ThT, revealed a three stage process with lag, growth and equilibrium phases. Seeding with preformed fibrils shortened the lag phase demonstrating the classic nucleated polymerization mechanism for the reaction. Interestingly, comparing thioflavin T kinetics with solubility and turbidity kinetics it was found that the protein initially formed non-thioflavionophilic, morphologically disordered aggregates that over time matured into amyloid fibrils. By transmission electron microscopy and by fluorescence microscopy of aggregates stained with luminescent conjugated polythiophenes (LCPs); we demonstrated that HuPrP undergoes a conformational conversion where spun and woven fibrils protruded from morphologically disordered aggregates. The initial aggregation functioned as a kinetic trap that decelerated nucleation into a fibrillation competent nucleus, but at the same time without aggregation there was no onset of amyloid fibril formation. The agitation, which was necessary for fibril formation to be induced, transiently exposes the protein to the air-water interface suggests a hitherto largely unexplored denaturing environment for prion conversion.

    Place, publisher, year, edition, pages
    Austin: Landes Bioscience Journals, 2009
    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-21064 (URN)10.4161/pri.3.4.10112 (DOI)000280061100009 ()
    Available from: 2009-09-28 Created: 2009-09-28 Last updated: 2018-04-25
    2. Multiple substitutions of methionine 129 in human prion protein reveal its importance in the amyloid fibrillation pathway
    Open this publication in new window or tab >>Multiple substitutions of methionine 129 in human prion protein reveal its importance in the amyloid fibrillation pathway
    2012 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 287, no 31, p. 25975-25984Article in journal (Refereed) Published
    Abstract [en]

    The role of the polymorphism Met or Val in position 129 in the human prion protein is well documented regarding disease susceptibility and clinical manifestations. However, little is known about the molecular background to this phenomenon. We investigated herein the conformational stability, amyloid fibrillation kinetics, and seeding propensity of different 129 mutants, located in β-strand 1 of PrP (Met129 (WT), M129A, M129V, M129L, M129W, M129P, M129E, M129K, and M129C) in HuPrP(90–231). The mutations M129V, M129L, M129K, and M129C did not affect stability (midpoints of thermal denaturation, Tm = 65–66 °C), whereas the mutants M129A and M129E and the largest side chain M129W were destabilized by 3–4 °C. The most destabilizing substitution was M129P, which lowered the Tm by 7.2 °C. All mutants, except for M129C, formed amyloid-like fibrils within hours during fibril formation under near physiological conditions. Fibril-forming mutants showed a sigmoidal kinetic profile and showed shorter lag times during seeding with preformed amyloid fibrils implicating a nucleated polymerization reaction. In the spontaneous reactions, the lag time of fibril formation was rather uniform for the mutants M129A, M129V, and M129L resembling the wild type. When the substituted amino acid had a distinct feature discriminating it from the wild type, such as size (M129W), charge (M129E, M129K), or rotational constraint (M129P), the fibrillation was impeded. M129C did not form ThT/Congo red-positive fibrils, and non-reducing SDS-PAGE of M129C during fibrillation conditions at different time points revealed covalent dimer formation already 15 min after fibrillation reaction initiation. Position 129 appears to be a key site for dictating PrP receptiveness toward recruitment into the amyloid state.

    Place, publisher, year, edition, pages
    American Society for Biochemistry and Molecular Biology, 2012
    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-53174 (URN)10.1074/jbc.M112.372136 (DOI)000306916300025 ()
    Note

    funding agencies|EU-FP7 Health Programme Project LUPAS||Swedish Research Council||Knut and Alice Wallenberg Foundation||Swedish Foundation for Strategic Research||Linkoping University Center for Neuroscience||

    Available from: 2010-01-18 Created: 2010-01-18 Last updated: 2018-04-25
    3. Native, amyloid fibrils and β-oligomers of the C-terminal domain of human prion protein display differential activation of complement and bind C1q, factor H and C4b-binding protein directly
    Open this publication in new window or tab >>Native, amyloid fibrils and β-oligomers of the C-terminal domain of human prion protein display differential activation of complement and bind C1q, factor H and C4b-binding protein directly
    2008 (English)In: Molecular Immunology, ISSN 0161-5890, E-ISSN 1872-9142, Vol. 45, no 11, p. 3213-3221Article in journal (Refereed) Published
    Abstract [en]

    Prion protein (PrP) is an endogenous protein involved in the pathogenesis of bovine spongiform encephalopathy and Creutzfeldt–Jakob disease. Murine PrP has been reported to bind C1q and activate the classical pathway of complement in a copper-dependent manner. Here we show that various conformational isoforms (native, amyloid fibrils, and β-oligomers) of recombinant human PrP (90–231 and 121–231) bind C1q and activate complement. PrP binds both the globular head and collagenous stalk domains of C1q. Native, β-oligomeric and amyloid fibrils of PrP all activate the classical and alternative pathways of complement to different extent. However, they do not trigger the lectin pathway. Of the tested PrP conformational isoforms we find that β-oligomers bind C1q and activate complement most strongly. Membrane attack complex formation initiated by PrP is subdued in comparison to deposition of early complement components. This is most likely attributed to the interaction between human PrP and complement inhibitors factor H and C4b-binding protein. Accordingly, PrP-triggered complement activation in the terminal pathway was increased in serum lacking C4b-binding protein. Taken together the present study indicates that complement activation may be an important factor in human prion diseases, suggesting that complement induced activities may prove relevant therapeutic targets.

    Keywords
    C1q, C4b-binding protein, Complement activation, Factor H, Human prion protein, Transmissible spongiform encephalopathies
    National Category
    Engineering and Technology
    Identifiers
    urn:nbn:se:liu:diva-45977 (URN)10.1016/j.molimm.2008.02.023 (DOI)
    Available from: 2009-10-11 Created: 2009-10-11 Last updated: 2018-04-25
    4. Properties of defined recombinant oligomeric forms of Aβ1‐42
    Open this publication in new window or tab >>Properties of defined recombinant oligomeric forms of Aβ1‐42
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Oligomers of Aβ1-42 have been identified in human Alzheimer´s disease (AD) patients and in mouse models of AD. These species have attracted intense interest as possible neurological pathogens in AD. In our hands, expression of recombinant human Aβ1-42 in Escherichia coli followed by purification in the presence of cupric ions (CuCl2) afforded recovery of high quantities (>5 mg/L of culture) of well defined trimeric, hexameric, nonameric and dodecameric Aβ1-42. Strong denaturing conditions such as 6 M GuHCI, 8 M urea or boiling in 6.5 M urea supplemented with 2.5 % SDS all failed to separate the oligomers into smaller building blocks implicating that the oligomers are composed of covalently cross-linked Aβ1-42 monomers. Purification in the absence of cupric ions resulted in monomeric Aβ1-42. The Aβ1-42 oligomers were toxic and induced apoptosis when administered to neuroblastoma cells in culture. The described method producing oligomeric Aβ1-42 from a recombinant expression system paves the way for mechanistic studies, structural analysis, drug screening and opens up for vaccine development.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-53175 (URN)
    Available from: 2010-01-18 Created: 2010-01-18 Last updated: 2018-04-25
  • 14.
    Nyström, Sofie
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Generic amyloidogenicity of mammalian prion proteins from species susceptible and resistant to prions2015In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 5, no 10101Article in journal (Refereed)
    Abstract [en]

    Prion diseases are lethal, infectious diseases associated with prion protein (PrP) misfolding. A large number of mammals are susceptible to both sporadic and acquired prion diseases. Although PrP is highly conserved and ubiquitously expressed in all mammals, not all species exhibit prion disease. By employing full length recombinant PrP from five known prion susceptible species (human, cattle, cat, mouse and hamster) and two species considered to be prion resistant (pig and dog) the amyloidogenicity of these PrPs has been delineated. All the mammalian PrPs, even from resistant species, were swiftly converted from the native state to amyloid-like structure when subjected to a native condition conversion assay. The PrPs displayed amyloidotypic tinctorial and ultrastructural hallmarks. Self-seeded conversion of the PrPs displayed significantly decreased lag phases demonstrating that nucleation dependent polymerization is a dominating mechanism in the fibrillation process. Fibrils from A beta 1-40, A beta 1-42, Lysozyme, Insulin and Transthyretin did not accelerate conversion of HuPrP whereas fibrils from HuPrP90-231 and HuPrP121-231 as well as full length PrPs of all PrPs efficiently seeded conversion showing specificity of the assay requiring the C-terminal PrP sequence. Our findings have implications for PrP misfolding and could have ramifications in the context of prion resistant species and silent carriers.

  • 15.
    Nyström, Sofie
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Is the prevalent human prion protein 129M/V mutation a living fossil from a Paleolithic panzootic superprion pandemic?2014In: Prion, ISSN 1933-6896, E-ISSN 1933-690X, Vol. 8, no 1Article in journal (Refereed)
    Abstract [en]

    Prion diseases are consistently associated with prion protein (PrPC) misfolding rendering a cascade of auto-catalytic self-perpetuation of misfolded PrP in an afflicted individual. The molecular process is intriguingly similar to all known amyloid diseases both local and systemic. The prion disease is also infectious by the transfer of misfolded PrP from one individual to the next. Transmissibility is surprisingly efficient in prion diseases and given the rapid disease progression following initial symptoms the prionoses stand out from other amyloidoses, which all may be transmissible under certain circumstances. The nature of the infectious prion as well as the genotype of the host is important for transmissibility. For hitherto unexplained reasons the majority of Europeans carry a missense mutation on one or both alleles of the PrP gene (PRNP), and hence express a variant of PrP with a substitution for valine (V) instead of methionine (M) in position 129. In fact the 129M/V variant is very common in all populations except for the Japanese. Sporadic Creutzfeldt-Jakob disease is a disease rarely striking people below the age of 60, where homozygosity especially 129MM is a very strong risk factor. Paradoxically, the 129M/V polymorphism suggestive of heterozygote advantage is one of the most clear cut disease associated traits of the human population, yet prion disease is extraordinarily rare. The genetic basis for how this trait spread with such prevalence within human populations is still target to investigations and deserves attention. This short essay represents a somewhat provocative hypothetical notion of a possible ancient significance of this polymorphism.

  • 16.
    Nyström, Sofie
    et al.
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
    Kågedal, Katarina
    Linköping University, Department of Neuroscience and Locomotion. Linköping University, Faculty of Health Sciences.
    Jonsson, Maria
    Astra Zeneca R & D, Södertälje, Sweden.
    Hedin, Linnea
    Astra Zeneca R & D, Södertälje, Sweden.
    Svensson, Samuel
    Astra Zeneca R & D, Södertälje, Sweden.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
    Properties of defined recombinant oligomeric forms of Aβ1‐42Manuscript (preprint) (Other academic)
    Abstract [en]

    Oligomers of Aβ1-42 have been identified in human Alzheimer´s disease (AD) patients and in mouse models of AD. These species have attracted intense interest as possible neurological pathogens in AD. In our hands, expression of recombinant human Aβ1-42 in Escherichia coli followed by purification in the presence of cupric ions (CuCl2) afforded recovery of high quantities (>5 mg/L of culture) of well defined trimeric, hexameric, nonameric and dodecameric Aβ1-42. Strong denaturing conditions such as 6 M GuHCI, 8 M urea or boiling in 6.5 M urea supplemented with 2.5 % SDS all failed to separate the oligomers into smaller building blocks implicating that the oligomers are composed of covalently cross-linked Aβ1-42 monomers. Purification in the absence of cupric ions resulted in monomeric Aβ1-42. The Aβ1-42 oligomers were toxic and induced apoptosis when administered to neuroblastoma cells in culture. The described method producing oligomeric Aβ1-42 from a recombinant expression system paves the way for mechanistic studies, structural analysis, drug screening and opens up for vaccine development.

  • 17.
    Nyström, Sofie
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Multiple substitutions of methionine 129 in human prion protein reveal its importance in the amyloid fibrillation pathway2012In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 287, no 31, p. 25975-25984Article in journal (Refereed)
    Abstract [en]

    The role of the polymorphism Met or Val in position 129 in the human prion protein is well documented regarding disease susceptibility and clinical manifestations. However, little is known about the molecular background to this phenomenon. We investigated herein the conformational stability, amyloid fibrillation kinetics, and seeding propensity of different 129 mutants, located in β-strand 1 of PrP (Met129 (WT), M129A, M129V, M129L, M129W, M129P, M129E, M129K, and M129C) in HuPrP(90–231). The mutations M129V, M129L, M129K, and M129C did not affect stability (midpoints of thermal denaturation, Tm = 65–66 °C), whereas the mutants M129A and M129E and the largest side chain M129W were destabilized by 3–4 °C. The most destabilizing substitution was M129P, which lowered the Tm by 7.2 °C. All mutants, except for M129C, formed amyloid-like fibrils within hours during fibril formation under near physiological conditions. Fibril-forming mutants showed a sigmoidal kinetic profile and showed shorter lag times during seeding with preformed amyloid fibrils implicating a nucleated polymerization reaction. In the spontaneous reactions, the lag time of fibril formation was rather uniform for the mutants M129A, M129V, and M129L resembling the wild type. When the substituted amino acid had a distinct feature discriminating it from the wild type, such as size (M129W), charge (M129E, M129K), or rotational constraint (M129P), the fibrillation was impeded. M129C did not form ThT/Congo red-positive fibrils, and non-reducing SDS-PAGE of M129C during fibrillation conditions at different time points revealed covalent dimer formation already 15 min after fibrillation reaction initiation. Position 129 appears to be a key site for dictating PrP receptiveness toward recruitment into the amyloid state.

  • 18.
    Nyström, Sofie
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Propagating Artificial Amyloid Strains of Recombinant Human Prion Protein with Mutations in Position 1292010In: Prion, ISSN 1933-6896, E-ISSN 1933-690X, Vol. 4, no 3, p. 124-124Article in journal (Other academic)
    Abstract [en]

    The influence of the polymorphism M129V in the human PrPgene is well documented. Most cases of sporadic CJD afflicthomozygous individuals. Differences in codon 129 genotypegive rise to differences in phenotype regarding plaque and clinicalsymptoms. Despite this, little is known about the molecularbackground to this phenomenon.

    To study this phenomenon in greater detail we employedrecombinant human prion protein. Using several artificial mutationsallowed us to study the influence of different amino acidproperties on the formation of amyloid prion protein. The variantsused were 129A, 129V, 129L, 129M, 129W, 129P, 129E and129K. Three mutants were chosen to vary the hydrophobicity,the tryptophan mutant was chosen due to its bulkiness and theproline for its constraint of the polypeptide backbone. 129E and129K may give information regarding the effect of charge in this position.

    The protein was expressed in Escherichia coli, purified andsubjected to agitation at 37°C at physiological pH and salt concentration(Almstedt et al. Prion 2009). All mutants formedcongophilic and Thioflavine T positive aggregates within hours.Fibrillar morphology was also confirmed using transmission electronmicroscopy.

    Seeding the mutant proteins with preformed fibrils of themutant itself or of wild type protein revealed differences in seedingefficiency for the different mutants. By monitoring the fibrilsresulting from the seeded fibrillation reactions using luminescentconjugated polymers, a templating effect was seen. This strainlikebehavior was followed through several generations of fibrils.The fragility of the seeding fibrils was taken under considerationand was analyzed using urea denaturation.

    Almstedt, Nyström S, Nilsson P, Hammarström P. Prion2009; 3:224-35.

  • 19.
    Nyström, Sofie
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Psonka-Antonczyk, Katarzyna M.
    Norwegian University of Science and Technology, Norway .
    Ellingsen, Pal Gunnar
    Norwegian University of Science and Technology, Norway .
    Johansson, Leif
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Reitan, Nina
    Norwegian University of Science and Technology, Norway .
    Handrick, Susann
    University of Medical Berlin, Germany .
    Prokop, Stefan
    University of Medical Berlin, Germany .
    Heppner, Frank L.
    University of Medical Berlin, Germany .
    Wegenast-Braun, Bettina M.
    German Centre Neurodegenerat Disease, Germany .
    Jucker, Mathias
    German Centre Neurodegenerat Disease, Germany .
    Lindgren, Mikael
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Torger Stokke, Bjorn
    Norwegian University of Science and Technology, Norway .
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Nilsson, Peter K R.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Evidence for Age-Dependent in Vivo Conformational Rearrangement within A beta Amyloid Deposits2013In: ACS Chemical Biology, ISSN 1554-8929, E-ISSN 1554-8937, Vol. 8, no 6, p. 1128-1133Article in journal (Refereed)
    Abstract [en]

    Deposition of aggregated A beta peptide in the brain is one of the major hallmarks of Alzheimers disease. Using a combination of two structurally different, but related, hypersensitive fluorescent amyloid markers, LCOs, reporting on separate ultrastructural elements, we show that conformational rearrangement occurs within A beta plaques of transgenic mouse models as the animals age. This important mechanistic insight should aid the design and evaluation of experiments currently using plaque load as readout.

  • 20.
    Nyström, Sofie
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Vahdat Shariat Panahi, Aida
    Linköping University, Department of Clinical and Experimental Medicine, Divison of Neurobiology. Linköping University, Faculty of Medicine and Health Sciences.
    Nilsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Westermark, Per
    d Department of Immunology , Genetics and Pathology, Uppsala University , Uppsala , Sweden.
    Westermark, Gunilla T.
    e Department of Medical Cell Biology , Uppsala University , Uppsala , Sweden.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Lundmark, Katarzyna
    Linköping University, Department of Clinical and Experimental Medicine, Divison of Neurobiology. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Center for Diagnostics, Clinical pathology.
    Seed-dependent templating of murine AA amyloidosis2017In: Amyloid: Journal of Protein Folding Disorders, ISSN 1350-6129, E-ISSN 1744-2818, Vol. 24, no sup1, p. 140-141Article in journal (Other academic)
    Abstract [en]

    n/a

  • 21.
    Sandberg, Alexander
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Purification and Fibrillation of Recombinant Human Amyloid-ß, Prion Protein, and Tau Under Native Conditions2018In: Amyloid Proteins: Methods and Protocols / [ed] Einar M. Sigurdsson, Miguel Calero and María Gasset, Humana Press, 2018, Vol. 1779, p. 147-166Chapter in book (Refereed)
    Abstract [en]

    Protein misfolding, aggregation, and amyloid formation is involved in a large number of diseases. Recombinantly expressed proteins to study the amyloid fibril formation process are important for mechanistic studies. We here report protocols for production, purification, and fibrillation of three different proteins commonly found in cerebral amyloid; Aß and Tau found in Alzheimers disease, Chronic traumatic brain injury, Corticobasal degeneration, and Progressive Supranuclear Palsy and human prion protein found in Creutzfeldt-Jakobs disease. The three protocols have in common that the protein is in a pH-neutral phosphate saline buffer during fibrillation to mimic their endogenous near physiological environment.

  • 22.
    Sjöberg, Andreas P.
    et al.
    Lund University, Department of Laboratory Medicine, Division of Medical Protein Chemistry, Wallenberg Laboratory, Malmö, Sweden.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Blom, Anna M.
    Lund University, Department of Laboratory Medicine, Division of Medical Protein Chemistry, Wallenberg Laboratory, Malmö, Sweden.
    Native, amyloid fibrils and β-oligomers of the C-terminal domain of human prion protein display differential activation of complement and bind C1q, factor H and C4b-binding protein directly2008In: Molecular Immunology, ISSN 0161-5890, E-ISSN 1872-9142, Vol. 45, no 11, p. 3213-3221Article in journal (Refereed)
    Abstract [en]

    Prion protein (PrP) is an endogenous protein involved in the pathogenesis of bovine spongiform encephalopathy and Creutzfeldt–Jakob disease. Murine PrP has been reported to bind C1q and activate the classical pathway of complement in a copper-dependent manner. Here we show that various conformational isoforms (native, amyloid fibrils, and β-oligomers) of recombinant human PrP (90–231 and 121–231) bind C1q and activate complement. PrP binds both the globular head and collagenous stalk domains of C1q. Native, β-oligomeric and amyloid fibrils of PrP all activate the classical and alternative pathways of complement to different extent. However, they do not trigger the lectin pathway. Of the tested PrP conformational isoforms we find that β-oligomers bind C1q and activate complement most strongly. Membrane attack complex formation initiated by PrP is subdued in comparison to deposition of early complement components. This is most likely attributed to the interaction between human PrP and complement inhibitors factor H and C4b-binding protein. Accordingly, PrP-triggered complement activation in the terminal pathway was increased in serum lacking C4b-binding protein. Taken together the present study indicates that complement activation may be an important factor in human prion diseases, suggesting that complement induced activities may prove relevant therapeutic targets.

  • 23.
    Zhang, Jun
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Surface Physics and Nano Science. Linköping University, Faculty of Science & Engineering. Linköping University, Department of Physics, Chemistry and Biology, Chemistry.
    Sandberg, Alexander
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Wu, Xiongyu
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Lindgren, Mikael
    Department of Physics, The Norwegian University of Science and Technology, Trondheim, Norway.
    Konradsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    trans-Stilbenoids with Extended Fluorescence Lifetimes for the Characterization of Amyloid Fibrils2017In: ACS Omega, ISSN 2470-1343, Vol. 2, no 8, p. 4693-4704Article in journal (Refereed)
    Abstract [en]

    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.

  • 24.
    Zhang, Jun
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Wang, Jun
    Linköping University, Department of Physics, Chemistry and Biology, Bioinformatics. Linköping University, Faculty of Science & Engineering.
    Sandberg, Alexander
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Wu, Xiongyu
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Nyström, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    LeVine, Harry III
    Sanders-Brown Center on Aging, University of Kentucky, KY 40536-0230, Lexington, USA..
    Konradsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Hammarström, Per
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Durbeej, Bo
    Linköping University, Department of Physics, Chemistry and Biology, Bioinformatics. Linköping University, Faculty of Science & Engineering.
    Lindgren, Mikael
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering. Department of Physics, Norwegian University of Science and Technology, 7491, Trondheim, Norway..
    Intramolecular Proton and Charge Transfer of Pyrene-based trans-Stilbene Salicylic Acids Applied to Detection of Aggregated Proteins.2018In: ChemPhysChem, ISSN 1439-4235, E-ISSN 1439-7641, Vol. 19, no 22, p. 3001-3009Article in journal (Refereed)
    Abstract [en]

    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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