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  • 1.
    Ahl, Ing-Marie
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Cell Biology. Linköping University, Faculty of Health Sciences.
    Jonsson, Bengt-Harald
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Tibell, Lena
    Linköping University, Department of Science and Technology, Visual Information Technology and Applications (VITA). Linköping University, The Institute of Technology.
    Thermodynamic Characterization of the Interaction between the C-Terminal Domain of Extracellular Superoxide Dismutase and Heparin by Isothermal Titration Calorimetry2009In: BIOCHEMISTRY, ISSN 0006-2960, Vol. 48, no 41, p. 9932-9940Article in journal (Refereed)
    Abstract [en]

    Extracellular superoxide dismutase (ECSOD) interacts with heparin through its C-terminal domain. In this study we used isothermal titration calorimetry (ITC) to get detailed thermodynamic information about the interaction. We have shown that the interaction between ECSOD and intestinal mucosal heparin (M-w 6000-30000 Da) is exothermic and driven by enthalpy at physiological salt concentration. However, the contribution from entropy is favorable for binding or small isolated heparin fragments. By studying different size-defined heparin fragments, we also concluded that it hexasaccharide moiety is sufficient for strong binding to ECSOD. The binding involves proton transfer from the buffer to the ECSOD-heparin complex, and the results indicate that the number of ionic interactions made between ECSOD and heparin upon binding varies from three to five for heparin and an octasaccharide fragment, respectively. Surprisingly and despite the many charges found oil both the protein and the polysaccharide, our results indicate that the nonionic contribution to the binding is large. From the temperature dependence we have calculated the constant pressure heat capacity change (Delta C-p) of the interaction to -644 J K-1 mol(-1) and -306 J K-1 mol(-1) for heparin and all octasaccharide, respectively

  • 2.
    Ahl, Ing-Marie
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Cell Biology. Linköping University, Faculty of Health Sciences.
    Jonsson, Bengt-Harald
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Tibell, Lena A. E.
    Linköping University, Department of Science and Technology, Visual Information Technology and Applications (VITA). Linköping University, The Institute of Technology.
    Analysis of Effects of Mutations in the C-Terminal Domain of Extracellular Superoxide Dismutase by Isothermal Titration Calorimetry and Phage DisplayManuscript (preprint) (Other academic)
    Abstract [en]

    n/a

  • 3.
    Ahlner, Alexandra
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Carlsson, Mats
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Jonsson, Bengt-Harald
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    PINT: a software for integration of peak volumes and extraction of relaxation rates2013In: Journal of Biomolecular NMR, ISSN 0925-2738, E-ISSN 1573-5001, Vol. 56, no 3, p. 191-202Article in journal (Refereed)
    Abstract [en]

    We present the software Peak INTegration (PINT), designed to perform integration of peaks in NMR spectra. The program is very simple to run, yet powerful enough to handle complicated spectra. Peaks are integrated by fitting predefined line shapes to experimental data and the fitting can be customized to deal with, for instance, heavily overlapped peaks. The results can be inspected visually, which facilitates systematic optimization of the line shape fitting. Finally, integrated peak volumes can be used to extract parameters such as relaxation rates and information about low populated states. The utility of PINT is demonstrated by applications to the 59 residue SH3 domain of the yeast protein Abp1p and the 289 residue kinase domain of murine EphB2.

  • 4.
    Akke, Mikael
    et al.
    Department of Biophysical Chemistry, Lund University.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Conformational Dynamics by Relaxation Dispersion2012In: Encyclopedia of Biophysics / [ed] Gordon C. K. Roberts., Elsevier, 2012Chapter in book (Other academic)
    Abstract [en]

    The Encyclopedia of Biophysics is envisioned both as an easily accessible source of information and as an introductory guide to the scientific literature. It includes entries describing both Techniques and Systems. In the Techniques entries, each of the wide range of methods which fall under the heading of Biophysics are explained in detail, together with the value and the limitations of the information each provides. Techniques covered range from diffraction (X-ray, electron and neutron) through a wide range of spectroscopic methods (X-ray, optical, EPR, NMR) to imaging (from electron microscopy to live cell imaging and MRI), as well as computational and simulation approaches.In the Systems entries, biophysical approaches to specific biological systems or problems – from protein and nucleic acid structure to membranes, ion channels and receptors – are described. These sections, which place emphasis on the integration of the different techniques, therefore provide an inroad into Biophysics from a biological more than from a technique-oriented physical/chemical perspective. Thus the Encyclopedia is intended to provide a resource both for biophysicists interested in methods beyond those used in their immediate sub-discipline and for those readers who are approaching biophysics from either a physical or biological background.

  • 5.
    Almstedt, Karin
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Lundqvist, Martin
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Carlsson, Jonas
    Linköping University, Department of Physics, Chemistry and Biology, Bioinformatics . Linköping University, The Institute of Technology.
    Karlsson, Martin
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Persson, Bengt
    Linköping University, Department of Physics, Chemistry and Biology, Bioinformatics . Linköping University, The Institute of Technology.
    Jonsson, Bengt-Harald
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Carlsson, Uno
    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.
    Unfolding a folding disease: folding, misfolding and aggregation of the marble brain syndrome-associated mutant H107Y of human carbonic anhydrase II2004In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 342, no 2, p. 619-633Article in journal (Refereed)
    Abstract [en]

    Most loss-of-function diseases are caused by aberrant folding of important proteins. These proteins often misfold due to mutations. The disease marble brain syndrome (MBS), known also as carbonic anhydrase II deficiency syndrome (CADS), can manifest in carriers of point mutations in the human carbonic anhydrase II (HCA II) gene. One mutation associated with MBS entails the His107Tyr substitution. Here, we demonstrate that this mutation is a remarkably destabilizing folding mutation. The loss-of-function is clearly a folding defect, since the mutant shows 64% of CO2 hydration activity compared to that of the wild-type at low temperature where the mutant is folded. On the contrary, its stability towards thermal and guanidine hydrochloride (GuHCl) denaturation is highly compromised. Using activity assays, CD, fluorescence, NMR, cross-linking, aggregation measurements and molecular modeling, we have mapped the properties of this remarkable mutant. Loss of enzymatic activity had a midpoint temperature of denaturation (Tm) of 16 °C for the mutant compared to 55 °C for the wild-type protein. GuHCl-denaturation (at 4 °C) showed that the native state of the mutant was destabilized by 9.2 kcal/mol. The mutant unfolds through at least two equilibrium intermediates; one novel intermediate that we have termed the molten globule light state and, after further denaturation, the classical molten globule state is populated. Under physiological conditions (neutral pH; 37 °C), the His107Tyr mutant will populate the molten globule light state, likely due to novel interactions between Tyr107 and the surroundings of the critical residue Ser29 that destabilize the native conformation. This intermediate binds the hydrophobic dye 8-anilino-1-naphthalene sulfonic acid (ANS) but not as strong as the molten globule state, and near-UV CD reveals the presence of significant tertiary structure. Notably, this intermediate is not as prone to aggregation as the classical molten globule. As a proof of concept for an intervention strategy with small molecules, we showed that binding of the CA inhibitor acetazolamide increases the stability of the native state of the mutant by 2.9 kcal/mol in accordance with its strong affinity. Acetazolamide shifts the Tm to 34 °C that protects from misfolding and will enable a substantial fraction of the enzyme pool to survive physiological conditions.

  • 6.
    Andersson, Theresa
    et al.
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
    Lundqvist, Martin
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Dolphin, Gunnar T.
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
    Enander, Karin
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics . Linköping University, The Institute of Technology.
    Jonsson, Bengt-Harald
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Nilsson, Jonas W.
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry . Linköping University, The Institute of Technology.
    Baltzer, Lars
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
    The binding of human Carbonic Anhydrase II by functionalized folded polypeptide receptors2005In: Chemistry and Biology, ISSN 1074-5521, E-ISSN 1879-1301, Vol. 12, no 11, p. 1245-1252Article in journal (Refereed)
    Abstract [en]

    Several receptors for human carbonic anhydrase II (HCAII) have been prepared by covalently attaching benzenesulfonamide carboxylates via aliphatic aminocarboxylic acid spacers of variable length to the side chain of a lysine residue in a designed 42 residue helix-loop-helix motif. The sulfonamide group binds to the active site zinc ion of human carbonic anhydrase II located in a 15 Å deep cleft. The dissociation constants of the receptor-HCAII complexes were found to be in the range from low micromolar to better than 20 nM, with the lowest affinities found for spacers with less than five methylene groups and the highest affinity found for the spacer with seven methylene groups. The results suggest that the binding is a cooperative event in which both the sulfonamide residue and the helix-loop-helix motif contribute to the overall affinity.

  • 7.
    Andrésen, Cecilia
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Protein Structure and Interaction in Health and Disease2011Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    This thesis focuses on protein structure, dynamics and interaction and their relation to human disease. In particular, the biophysical and structural properties of both well-ordered and partially disordered proteins are studied using a range of biophysical techniques such as circular dichroism spectroscopy, fluorescence spectroscopy, mass spectrometry and nuclear magnetic resonance spectroscopy. Pseudomonas aeruginosa is a human pathogen due to its multidrug resistance (MDR) caused by overexpression of efflux pump systems. This thesis describes how MDR mutations within the MexR repressor of the MexAB-OprM system reduce the DNA affinity by altering its stability with maintained structure. The oncogenic protein c-Myc is involved in many essential biological functions such as cell proliferation, differentiation and apoptosis and is also highly associated with several forms of human cancers, and where the N-terminal domain is regulated by a plethora of protein interactions. In this thesis the intrinsically disordered N-terminal part of c-Myc and its interactions with the proteins Bin1 and TBP are described. Myc binds Bin1 with maintained disorder in a multivalent manner, which may explain why the onco-protein can interact with such a wide range of binding partners. A similarly dynamic interaction is observed for Myc with the TATA-binding protein (TBP). The essential human multidomain glutaredoxin Grx3 is associated with several biological functions such as redox signaling, proliferation and signal transduction. We have solved the structure and analyzed the dynamic properties in the ps-ns and ms time scale for the two N-terminal domains, providing a platform for further analysis of the Grx3 protein and its interactions. Taken together, this thesis emphasizes the importance of joint structural, biophysical and dynamic studies to better understand protein function in health and disease.

    List of papers
    1. Critical biophysical properties in the Pseudomonas aeruginosa efflux gene regulator MexR are targeted by mutations conferring multidrug resistance
    Open this publication in new window or tab >>Critical biophysical properties in the Pseudomonas aeruginosa efflux gene regulator MexR are targeted by mutations conferring multidrug resistance
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    2010 (English)In: Protein Science, ISSN 0961-8368, E-ISSN 1469-896X, Vol. 19, no 4, p. 680-692Article in journal (Refereed) Published
    Abstract [en]

    The self-assembling MexA-MexB-OprM efflux pump system, encoded by the mexO operon, contributes to facile resistance of Pseudomonas aeruginosa by actively extruding multiple antimicrobials. MexR negatively regulates the mexO operon, comprising two adjacent MexR binding sites, and is as such highly targeted by mutations that confer multidrug resistance (MDR). To understand how MDR mutations impair MexR function, we studied MexR-wt as well as a selected set of MDR single mutants distant from the proposed DNA-binding helix. Although DNA affinity and MexA-MexB-OprM repression were both drastically impaired in the selected MexR-MDR mutants, MexR-wt bound its two binding sites in the mexO with high affinity as a dimer. In the MexR-MDR mutants, secondary structure content and oligomerization properties were very similar to MexR-wt despite their lack of DNA binding. Despite this, the MexR-MDR mutants showed highly varying stabilities compared with MexR-wt, suggesting disturbed critical interdomain contacts, because mutations in the DNA-binding domains affected the stability of the dimer region and vice versa. Furthermore, significant ANS binding to MexR-wt in both free and DNA-bound states, together with increased ANS binding in all studied mutants, suggest that a hydrophobic cavity in the dimer region already shown to be involved in regulatory binding is enlarged by MDR mutations. Taken together, we propose that the biophysical MexR properties that are targeted by MDR mutations stability, domain interactions, and internal hydrophobic surfaces are also critical for the regulation of MexR DNA binding.

    Place, publisher, year, edition, pages
    Cold Spring Harbor Laboratory Press, 2010
    Keywords
    DNA-binding protein, stability, efflux gene regulator, multidrug resistance, MarR family, Biacore, analytical ultracentrifugation, circular dichroism, fluorescence, real-time PCR
    National Category
    Engineering and Technology
    Identifiers
    urn:nbn:se:liu:diva-54849 (URN)10.1002/pro.343 (DOI)000276274900006 ()
    Available from: 2010-04-16 Created: 2010-04-16 Last updated: 2017-12-12Bibliographically approved
    2. Transient structure and intrinsic disorder in the c-Myc transactivation domain and its effects on ligand binding
    Open this publication in new window or tab >>Transient structure and intrinsic disorder in the c-Myc transactivation domain and its effects on ligand binding
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    The crucial role of c-Myc as an oncoprotein and as a key regulator of cell growth makes it essential to understand the molecular basis of c-Myc function. The transactivation domain of c-Myc coordinates a wealth of protein interactions involved in transformation, differentiation and apoptosis. We have characterized in detail the intrinsically disordered properties of c-Myc-1-88, where hierarchical phosphorylation of T58 and S62 regulates activation and destruction of the c-Myc protein. By NMR chemical shift analysis, relaxation measurements and NOE analysis, we show that both the MBI region (residues 45-65) and residues 22-33 are transiently structured regions, conserved also in other members of the Myc family. Binding of Bin1-SH3 to c-Myc-1-88 as assayed by NMR and SPR revealed primary binding to the S62 region, but also a dynamically disordered and multivalent complex in which intrinsic disorder of c-Myc-1-88 was retained while releasing transient intramolecular interactions. Our findings describe a novel mode of regulatory recognition of c-Myc that is in agreement with the increasingly recognized capability of intrinsically disordered regions to efficiently mediate transient interactions with a wide range of targets, with important implications towards understanding the unique multifaceted biological functions of c-Myc.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-70832 (URN)
    Available from: 2011-09-20 Created: 2011-09-20 Last updated: 2011-09-20Bibliographically approved
    3. Molecular characterization of the interaction between the disordered c-Myc transactivation domain and the TATA-binding protein (TBP)
    Open this publication in new window or tab >>Molecular characterization of the interaction between the disordered c-Myc transactivation domain and the TATA-binding protein (TBP)
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    The proto-oncogene c-myc affects the occurrence, expansion, and evolution of numerous aggressive human cancers, and is often associated with the late-stage and/or poor prognostic disease. Regulation of target gene activity by c-Myc occurs through protein interactions with the c-Myc transactivation domain (TAD) which, in addition to binding the TATA-binding protein (TBP) also recruits a wide variety of co-activators and suppressor proteins. Here, we present a molecular model, based on NMR, X-ray crystallography and SPR measurements, which describes how the c-Myc TAD binds to TBP. Our model contributes to the understanding of how c-Myc can regulate individual genes as well as entire gene programs.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-70833 (URN)
    Available from: 2011-09-20 Created: 2011-09-20 Last updated: 2011-09-20Bibliographically approved
    4. Structural and dynamic analysis of human glutaredoxin 3
    Open this publication in new window or tab >>Structural and dynamic analysis of human glutaredoxin 3
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Human glutaredoxin (Grx3) is an essential protein associated with biological functions including embryonic development and immune response, and is involved in human disease such as lung, colon cancer and cardiovascular disorder. Grx3 can harbour a [2Fe-2S]2+ cluster and is most likely involved in oxidative stress response. Grx3 consists of an N-terminal thioredoxin-like domain and two additional monothiol glutaredoxin domains, and is thus classified as a multidomain monothiol glutaredoxin. The Grx3 thioredoxin domain lacks both a characteristic active-site and catalytic activity, but is still essential in the yeast homologue and presumably functions together with its monothiol glutaredoxin domains. We have characterised the structures of the two Nterminal domains in Grx3, which have thioredoxin and glutaredoxin folds. We have analysed their dynamic and structural interdependence by analysing NMR relaxation data together with chemical shift changes between isolated and covalently linked domains. We find that although the two domains show interdomain mobility around a semi-flexible linker, there are indications for a preferred interaction surface between the two domains. Millisecond internal dynamics in a suggested ligand binding site in the isolated thioredoxin domain is dampened in the domain pair, suggesting that the two domains mutually affect each other on a profound level. Our results present a platform for further detailed studies of multidomain thioredoxin-glutaredoxin containing proteins, and their function in human cells.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-70834 (URN)
    Available from: 2011-09-20 Created: 2011-09-20 Last updated: 2011-09-20Bibliographically approved
    5. A Novel Monothiol Glutaredoxin (Grx4) from Escherichia coli Can Serve as a Substrate for Thioredoxin Reductase
    Open this publication in new window or tab >>A Novel Monothiol Glutaredoxin (Grx4) from Escherichia coli Can Serve as a Substrate for Thioredoxin Reductase
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    2005 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 280, no 26, p. 24544-24552Article in journal (Refereed) Published
    Abstract [en]

    Glutaredoxins are ubiquitous proteins that catalyze the reduction of disulfides via reduced glutathione (GSH). Escherichia coli has three glutaredoxins (Grx1, Grx2, and Grx3), all containing the classic dithiol active site CPYC. We report the cloning, expression, and characterization of a novel monothiol E. coli glutaredoxin, which we name glutaredoxin 4 (Grx4). The protein consists of 115 amino acids (12.7 kDa), has a monothiol (CGFS) potential active site and shows high sequence homology to the other monothiol glutaredoxins and especially to yeast Grx5. Experiments with gene knock-out techniques showed that the reading frame encoding Grx4 was essential. Grx4 was inactive as a GSH-disulfide oxidoreductase in a standard glutaredoxin assay with GSH and hydroxyethyl disulfide in a complete system with NADPH and glutathione reductase. An engineered CGFC active site mutant did not gain activity either. Grx4 in reduced form contained three thiols, and treatment with oxidized GSH resulted in glutathionylation and formation of a disulfide. Remarkably, this disulfide of Grx4 was a direct substrate for NADPH and E. coli thioredoxin reductase, whereas the mixed disulfide was reduced by Grx1. Reduced Grx4 showed the potential to transfer electrons to oxidized E. coli Grx1 and Grx3. Grx4 is highly abundant (750–2000 ng/mg of total soluble protein), as determined by a specific enzyme-link immunosorbent assay, and most likely regulated by guanosine 3′,5′-tetraphosphate upon entry to stationary phase. Grx4 was highly elevated upon iron depletion, suggesting an iron-related function for the protein.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-41980 (URN)10.1074/jbc.M500678200 (DOI)59458 (Local ID)59458 (Archive number)59458 (OAI)
    Available from: 2009-10-10 Created: 2009-10-10 Last updated: 2017-12-13Bibliographically approved
  • 8.
    Andrésen, Cecilia
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Anandapadamanaban, Madhanagopal
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Helander, Sara
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Fladvad, Malin
    Department of Medical Biochemistry and Biophysics, Karolinska Institutet, S171 77 Stockholm.
    Andersson, Karl
    Biacore AB, GE Healthcare Europe GmbH, Björkgatan 30, SE-571 25 Uppsala, Sweden.
    Kanmert, Daniel
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Säfsten, Pär
    Biacore AB, GE Healthcare Europe GmbH, Björkgatan 30, SE-571 25 Uppsala, Sweden.
    Moche, Martin
    Department of Medical Biochemistry and Biophysics, Karolinska Institutet, S171 77 Stockholm.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Molecular characterization of the interaction between the disordered c-Myc transactivation domain and the TATA-binding protein (TBP)Manuscript (preprint) (Other academic)
    Abstract [en]

    The proto-oncogene c-myc affects the occurrence, expansion, and evolution of numerous aggressive human cancers, and is often associated with the late-stage and/or poor prognostic disease. Regulation of target gene activity by c-Myc occurs through protein interactions with the c-Myc transactivation domain (TAD) which, in addition to binding the TATA-binding protein (TBP) also recruits a wide variety of co-activators and suppressor proteins. Here, we present a molecular model, based on NMR, X-ray crystallography and SPR measurements, which describes how the c-Myc TAD binds to TBP. Our model contributes to the understanding of how c-Myc can regulate individual genes as well as entire gene programs.

  • 9.
    Andrésen, Cecilia
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Helander, Sara
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Lemak, Alexander
    University of Toronto, Canada .
    Fares, Christophe
    University of Toronto, Canada .
    Csizmok, Veronika
    Hospital for Sick Children, Canada .
    Carlsson, Jonas
    Linköping University, Department of Physics, Chemistry and Biology, Bioinformatics. Linköping University, The Institute of Technology.
    Penn, Linda Z
    University of Toronto, Canada .
    Forman-Kay, Julie D
    Hospital Sick Children, Canada University of Toronto, Canada .
    Arrowsmith, Cheryl H
    University of Toronto, Canada.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Transient structure and dynamics in the disordered c-Myc transactivation domain affect Bin1 binding2012In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 40, no 13, p. 6353-6366Article in journal (Refereed)
    Abstract [en]

    The crucial role of Myc as an oncoprotein and as a key regulator of cell growth makes it essential to understand the molecular basis of Myc function. The N-terminal region of c-Myc coordinates a wealth of protein interactions involved in transformation, differentiation and apoptosis. We have characterized in detail the intrinsically disordered properties of Myc-1-88, where hierarchical phosphorylation of S62 and T58 regulates activation and destruction of the Myc protein. By nuclear magnetic resonance (NMR) chemical shift analysis, relaxation measurements and NOE analysis, we show that although Myc occupies a very heterogeneous conformational space, we find transiently structured regions in residues 22-33 and in the Myc homology box I (MBI; residues 45-65); both these regions are conserved in other members of the Myc family. Binding of Bin1 to Myc-1-88 as assayed by NMR and surface plasmon resonance (SPR) revealed primary binding to the S62 region in a dynamically disordered and multivalent complex, accompanied by population shifts leading to altered intramolecular conformational dynamics. These findings expand the increasingly recognized concept of intrinsically disordered regions mediating transient interactions to Myc, a key transcriptional regulator of major medical importance, and have important implications for further understanding its multifaceted role in gene regulation.

  • 10.
    Andrésen, Cecilia
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Helander, Sara
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Lemak, Alexander
    Department of Medical Biophysics, University of Toronto, 101 College Street, Toronto, Ontario M5G 1L7, Canada.
    Farès, Christophe
    Department of Medical Biophysics, University of Toronto, 101 College Street, Toronto, Ontario M5G 1L7, Canada.
    Csizmok, Veronika
    Molecular Structure and Function Program, Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.
    Carlsson, Jonas
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Penn, Linda Z.
    Department of Medical Biophysics, University of Toronto, 101 College Street, Toronto, Ontario M5G 1L7, Canada.
    Forman-Kay, Julie D.
    Molecular Structure and Function Program, Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.
    Arrowsmith, Cheryl H.
    Department of Medical Biophysics, University of Toronto, 101 College Street, Toronto, Ontario M5G 1L7, Canada.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Transient structure and intrinsic disorder in the c-Myc transactivation domain and its effects on ligand bindingManuscript (preprint) (Other academic)
    Abstract [en]

    The crucial role of c-Myc as an oncoprotein and as a key regulator of cell growth makes it essential to understand the molecular basis of c-Myc function. The transactivation domain of c-Myc coordinates a wealth of protein interactions involved in transformation, differentiation and apoptosis. We have characterized in detail the intrinsically disordered properties of c-Myc-1-88, where hierarchical phosphorylation of T58 and S62 regulates activation and destruction of the c-Myc protein. By NMR chemical shift analysis, relaxation measurements and NOE analysis, we show that both the MBI region (residues 45-65) and residues 22-33 are transiently structured regions, conserved also in other members of the Myc family. Binding of Bin1-SH3 to c-Myc-1-88 as assayed by NMR and SPR revealed primary binding to the S62 region, but also a dynamically disordered and multivalent complex in which intrinsic disorder of c-Myc-1-88 was retained while releasing transient intramolecular interactions. Our findings describe a novel mode of regulatory recognition of c-Myc that is in agreement with the increasingly recognized capability of intrinsically disordered regions to efficiently mediate transient interactions with a wide range of targets, with important implications towards understanding the unique multifaceted biological functions of c-Myc.

  • 11.
    Andrésen, Cecilia
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Jalal, Shah
    Karolinska University Hospital.
    Aili, Daniel
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics . Linköping University, The Institute of Technology.
    Wang, Yi
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
    Islam, Sohidul
    Karolinska University Hospital.
    Jarl, Anngelica
    Linköping University, Department of Physics, Chemistry and Biology, Molecular genetics . Linköping University, The Institute of Technology.
    Liedberg, Bo
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics . Linköping University, The Institute of Technology.
    Wretlind, Bengt
    Karolinska University Hospital.
    Mårtensson, Lars-Göran
    Linköping University, Department of Physics, Chemistry and Biology, Biochemistry. Linköping University, The Institute of Technology.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Critical biophysical properties in the Pseudomonas aeruginosa efflux gene regulator MexR are targeted by mutations conferring multidrug resistance2010In: Protein Science, ISSN 0961-8368, E-ISSN 1469-896X, Vol. 19, no 4, p. 680-692Article in journal (Refereed)
    Abstract [en]

    The self-assembling MexA-MexB-OprM efflux pump system, encoded by the mexO operon, contributes to facile resistance of Pseudomonas aeruginosa by actively extruding multiple antimicrobials. MexR negatively regulates the mexO operon, comprising two adjacent MexR binding sites, and is as such highly targeted by mutations that confer multidrug resistance (MDR). To understand how MDR mutations impair MexR function, we studied MexR-wt as well as a selected set of MDR single mutants distant from the proposed DNA-binding helix. Although DNA affinity and MexA-MexB-OprM repression were both drastically impaired in the selected MexR-MDR mutants, MexR-wt bound its two binding sites in the mexO with high affinity as a dimer. In the MexR-MDR mutants, secondary structure content and oligomerization properties were very similar to MexR-wt despite their lack of DNA binding. Despite this, the MexR-MDR mutants showed highly varying stabilities compared with MexR-wt, suggesting disturbed critical interdomain contacts, because mutations in the DNA-binding domains affected the stability of the dimer region and vice versa. Furthermore, significant ANS binding to MexR-wt in both free and DNA-bound states, together with increased ANS binding in all studied mutants, suggest that a hydrophobic cavity in the dimer region already shown to be involved in regulatory binding is enlarged by MDR mutations. Taken together, we propose that the biophysical MexR properties that are targeted by MDR mutations stability, domain interactions, and internal hydrophobic surfaces are also critical for the regulation of MexR DNA binding.

  • 12.
    Andrésen, Cecilia
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Structural and dynamic analysis of human glutaredoxin 3Manuscript (preprint) (Other academic)
    Abstract [en]

    Human glutaredoxin (Grx3) is an essential protein associated with biological functions including embryonic development and immune response, and is involved in human disease such as lung, colon cancer and cardiovascular disorder. Grx3 can harbour a [2Fe-2S]2+ cluster and is most likely involved in oxidative stress response. Grx3 consists of an N-terminal thioredoxin-like domain and two additional monothiol glutaredoxin domains, and is thus classified as a multidomain monothiol glutaredoxin. The Grx3 thioredoxin domain lacks both a characteristic active-site and catalytic activity, but is still essential in the yeast homologue and presumably functions together with its monothiol glutaredoxin domains. We have characterised the structures of the two Nterminal domains in Grx3, which have thioredoxin and glutaredoxin folds. We have analysed their dynamic and structural interdependence by analysing NMR relaxation data together with chemical shift changes between isolated and covalently linked domains. We find that although the two domains show interdomain mobility around a semi-flexible linker, there are indications for a preferred interaction surface between the two domains. Millisecond internal dynamics in a suggested ligand binding site in the isolated thioredoxin domain is dampened in the domain pair, suggesting that the two domains mutually affect each other on a profound level. Our results present a platform for further detailed studies of multidomain thioredoxin-glutaredoxin containing proteins, and their function in human cells.

  • 13.
    Auer, Renate
    et al.
    University of Toronto.
    Neudecker, Philipp
    University of Toronto.
    Muhandiram, D Ranjith
    University of Toronto.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Flemming Hansen, D
    University of Toronto.
    Konrat, Robert
    University of Vienna.
    Kay, Lewis E
    University of Toronto.
    Measuring the Signs of H-1(alpha) Chemical Shift Differences Between Ground and Excited Protein States by Off-Resonance Spin-Lock R-1 rho NMR Spectroscopy2009In: JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, ISSN 0002-7863, Vol. 131, no 31, p. 10832-10833Article in journal (Refereed)
    Abstract [en]

    Analysis of Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion NMR profiles provides the kinetics and thermodynamics of millisecond-time-scale exchange processes involving the interconversion of populated ground and invisible excited states. In addition, the absolute values of chemical, shift differences between NMR probes in the exchanging states, vertical bar Delta(pi)vertical bar, are also extracted. Herein, we present a simple experiment for obtaining the sign of H-1(alpha) Delta(pi) values by measuring off-resonance H-1(alpha) decay rates, R-1 rho, using weak proton spin-lock fields. A pair of R-1 rho values is measured with a spin-lock field applied vertical bar Delta omega vertical bar downfield and upfield of the major-state peak. In many cases, these two relaxation rates differ substantially, with the larger one corresponding to the case where the spin-lock field coincides with the resonance frequency of the probe in the minor state. The utility of the methodology is demonstrated first on a system involving protein ligand exchange and subsequently on an SH3 domain exchanging between a folded state and its on-pathway folding intermediate. With this experiment, it thus becomes possible to determine H-1(alpha) chemical shifts of the invisible excited state, which can be used as powerful restraints in defining the structural properties of these elusive conformers.

  • 14.
    Babu Moparthi, Satish
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering. Institut Fresnel, CNRS UMR 7249, Aix-Marseille Université, Marseille, France.
    Sjölander, Daniel
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Villebeck, Laila
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Jonsson, Bengt-Harald
    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.
    Carlsson, Uno
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, The Institute of Technology.
    Transient conformational remodeling of folding proteins by GroES - Individually and in concert with GroEL2014In: Journal of chemical biology, ISSN 1864-6158, E-ISSN 1864-6166, Vol. 7, no 1, p. 1-15Article, review/survey (Refereed)
    Abstract [en]

    The commonly accepted dogma of the bacterial GroE chaperonin system entails protein folding mediated by cycles of several ATP-dependent sequential steps where GroEL interacts with the folding client protein. In contrast, we herein report GroES-mediated dynamic remodeling (expansion and compression) of two different protein substrates during folding: the endogenous substrate MreB and carbonic anhydrase (HCAII), a well-characterized protein folding model. GroES was also found to influence GroEL binding induced unfolding and compression of the client protein underlining the synergistic activity of both chaperonins, even in the absence of ATP. This previously unidentified activity by GroES should have important implications for understanding the chaperonin mechanism and cellular stress response. Our findings necessitate a revision of the GroEL/ES mechanism.

  • 15.
    Bach, Anders
    et al.
    University of Copenhagen.
    Clausen, Bettina H
    University of South Denmark.
    Moller, Magda
    University of Copenhagen.
    Vestergaard, Bente
    University of Copenhagen.
    Chi, Celestine N
    Uppsala University.
    Round, Adam
    European Molecular Biol Lab.
    Sorensen, Pernille L
    University of Copenhagen.
    Nissen, Klaus B
    University of Copenhagen.
    Kastrup, Jette S
    University of Copenhagen.
    Gajhede, Michael
    University of Copenhagen.
    Jemth, Per
    Uppsala University.
    Kristensen, Anders S
    University of Copenhagen.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Lambertsen, Kate L
    University of South Denmark.
    Stromgaard, Kristian
    University of Copenhagen.
    A high-affinity, dimeric inhibitor of PSD-95 bivalently interacts with PDZ1-2 and protects against ischemic brain damage2012In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 109, no 9, p. 3317-3322Article in journal (Refereed)
    Abstract [en]

    Inhibition of the ternary protein complex of the synaptic scaffolding protein postsynaptic density protein-95 (PSD-95), neuronal nitric oxide synthase (nNOS), and the N-methyl-D-aspartate (NMDA) receptor is a potential strategy for treating ischemic brain damage, but high-affinity inhibitors are lacking. Here we report the design and synthesis of a novel dimeric inhibitor, Tat-NPEG4(IETDV)(2) (Tat-N-dimer), which binds the tandem PDZ1-2 domain of PSD-95 with an unprecedented high affinity of 4.6 nM, and displays extensive protease-resistance as evaluated in vitro by stability-measurements in human blood plasma. X-ray crystallography, NMR, and small-angle X-ray scattering (SAXS) deduced a true bivalent interaction between dimeric inhibitor and PDZ1-2, and also provided a dynamic model of the conformational changes of PDZ1-2 induced by the dimeric inhibitor. A single intravenous injection of Tat-N-dimer (3 nmol/g) to mice subjected to focal cerebral ischemia reduces infarct volume with 40% and restores motor functions. Thus, Tat-N-dimer is a highly efficacious neuroprotective agent with therapeutic potential in stroke.

  • 16.
    Bakszt, Rebecca
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Wernimont, Amy
    University of Toronto.
    Allali-Hassani, Abdellah
    University of Toronto.
    Wai Mok, Man
    University of Toronto.
    Hills, Tanya
    University of Toronto.
    Hui, Raymond
    University of Toronto.
    Pizarro, Juan C
    University of Toronto.
    The Crystal Structure of Toxoplasma gondii Pyruvate Kinase 12010In: PLOS ONE, ISSN 1932-6203, Vol. 5, no 9, p. 0012736-Article in journal (Refereed)
    Abstract [en]

    Background: Pyruvate kinase (PK), which catalyzes the final step in glycolysis converting phosphoenolpyruvate to pyruvate, is a central metabolic regulator in most organisms. Consequently PK represents an attractive therapeutic target in cancer and human pathogens, like Apicomplexans. The phylum Aplicomplexa, a group of exclusively parasitic organisms, includes the genera Plasmodium, Cryptosporidium and Toxoplasma, the etiological agents of malaria, cryptosporidiosis and toxoplasmosis respectively. Toxoplasma gondii infection causes a mild illness and is a very common infection affecting nearly one third of the worlds population. Methodology/Principal Findings: We have determined the crystal structure of the PK1 enzyme from T. gondii, with the B domain in the open and closed conformations. We have also characterized its enzymatic activity and confirmed glucose-6-phosphate as its allosteric activator. This is the first description of a PK enzyme in a closed inactive conformation without any bound substrate. Comparison of the two tetrameric TgPK1 structures indicates a reorientation of the monomers with a concomitant change in the buried surface among adjacent monomers. The change in the buried surface was associated with significant B domain movements in one of the interacting monomers. Conclusions: We hypothesize that a loop in the interface between the A and B domains plays an important role linking the position of the B domain to the buried surface among monomers through two a-helices. The proposed model links the catalytic cycle of the enzyme with its domain movements and highlights the contribution of the interface between adjacent subunits. In addition, an unusual ordered conformation was observed in one of the allosteric binding domains and it is related to a specific apicomplexan insertion. The sequence and structural particularity would explain the atypical activation by a mono-phosphorylated sugar. The sum of peculiarities raises this enzyme as an emerging target for drug discovery.

  • 17.
    Basaiawmoit, R V
    et al.
    Aarhus University.
    Oliveira, C L P
    Aarhus University.
    Runager, K
    Aarhus University.
    Sorensen, C S
    Aarhus University.
    Behrens, M A
    Aarhus University.
    Jonsson, Bengt-Harald
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Kristensen, T
    Aarhus University.
    Klintworth, G K
    Duke University.
    Enghild, J J
    Aarhus University.
    Skov Pedersen, J
    Aarhus University.
    Otzen, D E
    Aarhus University.
    SAXS Models of TGFBIp Reveal a Trimeric Structure and Show That the Overall Shape Is Not Affected by the Arg124His Mutation2011In: JOURNAL OF MOLECULAR BIOLOGY, ISSN 0022-2836, Vol. 408, no 3, p. 503-513Article in journal (Refereed)
    Abstract [en]

    Human transforming growth factor beta induced protein (TGFBIp) is composed of 683 residues, including an N-terminal cysteine-rich (EMI) domain, four homologous fasciclin domains, and an Arg-Gly-Asp (RGD) motif near the C-terminus. The protein is of interest because mutations in the TGFBI gene encoding TGFBIp lead to corneal dystrophy (CD), a condition where protein aggregates within the cornea compromise transparency. The complete three-dimensional structure of TGFBIp is not yet available, with the exception of a partial X-ray structure of the archetype FAS1 domain derived from Drosophila fasciclin-1. In this study, small-angle X-ray scattering (SAXS) models of intact wild-type (WT) human TGFBIp and a mutant (R124H) are presented. The mutation R124H leads to a variant of granular CD. The deduced structure of the TGFBIp monomer consists of four FAS1 domains in a simple "beads-on-a-string" arrangement, constructed by the superimposition of four consecutive Drosophila fasciclin domains. The SAXS-based model of the TGFBIp R124H mutant displayed no structural differences from WT. Both WT TGFBIp and the R124H mutant formed trimers at higher protein concentrations. The similar association properties and three-dimensional shape of the two proteins suggest that the mutation does not induce any major structural rearrangements, but points towards the role of other corneal-specific factors in the formation of corneal R124H deposits.

  • 18.
    Berglund, Anders
    et al.
    Department of Chemistry, Umeå University.
    Brorsson, Ann-Christin
    Biochemistry, Umeå University.
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Sethson, Ingmar
    Department of Chemistry, Umeå University.
    The equilibrium unfolding of MerP characterized by multivariate analysis of 2D NMR data2005In: Journal of magnetic resonance, ISSN 1090-7807, E-ISSN 1096-0856, Vol. 172, no 1, p. 24-30Article in journal (Refereed)
    Abstract [en]

    A general problem when analysing NMR spectra that reflect variations in the environment of target molecules is that different resonances are affected to various extents. Often a few resonances that display the largest frequency changes are selected as probes to reflect the examined variation, especially in the case, where the NMR spectra contain numerous resonances. Such a selection is dependent on more or less intuitive judgements and relying on the observed spectral variation being primarily caused by changes in the NMR sample. Second, recording changes observed for a few (albeit significant) resonances is inevitably accompanied by not using all available information in the analysis. Likewise, the commonly used chemical shift mapping (CSM) [Biochemistry 39 (2000) 26, Biochemistry 39 (2000) 12595] constitutes a loss of information since the total variation in the data is not retained in the projection into this single variable. Here, we describe a method for subjecting 2D NMR time-domain data to multivariate analysis and illustrate it with an analysis of multiple NMR experiments recorded at various folding conditions for the protein MerP. The calculated principal components provide an unbiased model of variations in the NMR spectra and they can consequently be processed as NMR data, and all the changes as reflected in the principal components are thereby made available for visual inspection in one single NMR spectrum. This approach is much less laborious than consideration of large numbers of individual spectra, and it greatly increases the interpretative power of the analysis. © 2004 Elsevier Inc. All rights reserved.

  • 19.
    Bivall Persson, Petter
    et al.
    Linköping University, Department of Science and Technology, Visual Information Technology and Applications (VITA). Linköping University, The Institute of Technology.
    Cooper, Matthew
    Linköping University, Department of Science and Technology, Visual Information Technology and Applications (VITA). Linköping University, The Institute of Technology.
    Tibell, Lena
    Linköping University, Department of Biomedicine and Surgery, Division of cell biology. Linköping University, Faculty of Health Sciences.
    Ainsworth, Shaaron
    Learning Sciences Research Institute, University of Nottingham, Nottingham, UK.
    Ynnerman, Anders
    Linköping University, Department of Science and Technology, Visual Information Technology and Applications (VITA). Linköping University, The Institute of Technology.
    Jonsson, Bengt-Harald
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Designing and Evaluating a Haptic System for Biomolecular Education2007In: IEEE Virtual Reality Conference, 2007. VR '07. / [ed] Sherman, W; Lin, M; Steed, A, Piscataway, NJ, USA: IEEE , 2007, p. 171-178Conference paper (Refereed)
    Abstract [en]

    In this paper we present an in situ evaluation of a haptic system, with a representative test population, we aim to determine what, if any, benefit haptics can have in a biomolecular education context. We have developed a haptic application for conveying concepts of molecular interactions, specifically in protein-ligand docking. Utilizing a semi-immersive environment with stereo graphics, users are able to manipulate the ligand and feel its interactions in the docking process. The evaluation used cognitive knowledge tests and interviews focused on learning gains. Compared with using time efficiency as the single quality measure this gives a better indication of a system's applicability in an educational environment. Surveys were used to gather opinions and suggestions for improvements. Students do gain from using the application in the learning process but the learning appears to be independent of the addition of haptic feedback. However the addition of force feedback did decrease time requirements and improved the students understanding of the docking process in terms of the forces involved, as is apparent from the students' descriptions of the experience. The students also indicated a number of features which could be improved in future development.

  • 20.
    Bivall Persson, Petter
    et al.
    Linköping University, Department of Science and Technology, Visual Information Technology and Applications (VITA). Linköping University, The Institute of Technology.
    Cooper, Matthew
    Linköping University, Department of Science and Technology, Media and Information Technology. Linköping University, Faculty of Science & Engineering.
    Ynnerman, Anders
    Linköping University, Department of Science and Technology, Visual Information Technology and Applications (VITA). Linköping University, The Institute of Technology.
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology.
    Tibell, Lena
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Biomedicine and Surgery, Division of cell biology.
    Use of Chemical Force Feedback for Multisensory Insights into Ligand Docking2007In: VII European Symposium of The Protein Society: From Proteins to Proteome, 2007, p. 151-151Conference paper (Refereed)
  • 21.
    Bivall Persson, Petter
    et al.
    Linköping University, Department of Science and Technology, Media and Information Technology. Linköping University, Faculty of Science & Engineering.
    Tibell, Lena
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Biomedicine and Surgery, Division of cell biology.
    Cooper, Matthew
    Linköping University, Department of Science and Technology, Visual Information Technology and Applications (VITA). Linköping University, The Institute of Technology.
    Ynnerman, Anders
    Linköping University, Department of Science and Technology, Visual Information Technology and Applications (VITA). Linköping University, The Institute of Technology.
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology.
    Evaluating the Effectiveness of Haptic Visualization in Biomolecular Education - Feeling Molecular Specificity in a Docking Task2006In: 12th IOSTE Symposium, Universiti Science Malaysia , 2006, p. 745-752Conference paper (Refereed)
    Abstract [en]

    Within the molecular life sciences extensive use is made of visual representations, ranging from sketches to advanced computer graphics, often used to convey abstract knowledge that is difficult for the student to grasp. This work evaluates a new visual and haptic (tactile/kinetic) tool for protein docking in an in situ learning situation by combining qualitative and quantitative methods, performing tests and interviews with students; all aiming at a proper inclusion of visualization tools into biomolecular education. Preliminary results indicate time gains, strong positive affective responses and learning gains from the tasks, however the influence of haptics needs further investigation.

  • 22.
    Bolognesi, B
    et al.
    University of Cambridge, Cambridge, UK.
    Jahn, T R
    University of Cambridge, Cambridge, UK.
    Brorsson, Ann-Christin
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology.
    Luheshi, L M
    University of Cambridge, Cambridge, UK.
    Yerbury, J J
    University of Wollongong, Wollongong, NSW, Australia .
    Crowther, D C
    University of Cambridge, Cambridge, UK.
    Dobson, C M
    University of Cambridge, Cambridge, UK.
    The N-terminus of amyloid-beta plays a crucial role in its aggregation and toxicity2010In: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 277, no Suppl. 1, p. 79-80Article in journal (Other academic)
    Abstract [en]

    The aggregation of Amyloid Beta (Aß) peptide into insolubleamyloid fibrils that deposit in the brain is one of the primarypathogenic events in Alzheimer’s disease. We have previouslyshown, using a Drosophila model of Aß toxicity, that the N terminus of the Aß peptide, despite being unstructured in themature Aß fibril, nonetheless affects Aß induced neurodegeneration in vivo. In order to understand the contribution of the N terminusof Aß to its aggregation behaviour, we have investigated anumber of rationally designed N-terminal mutants in vitro. We find that single amino acid mutations in this region affect significantlythe kinetics of Aß aggregation in vitro as measured by arange of spectroscopic techniques. Furthermore, we observe striking differences in the morphology of the aggregated speciesformed by these different Aß mutants when imaged with TEM or  AFM  and  also  in the ß-sheet  content  of their  mature  fibrils. Interestingly, mutants with an increased net charge or lower hydrophobicity tend  to show slower aggregation  kinetics, and  to form more ordered  aggregates  whereas mutations that  reduce net charge   or   increase   hydrophobicity   favour   faster   aggregation kinetics   and   poorly   structured  aggregates.   In   addition,    the exposed  hydrophobicity of aggregates  formed  in the early stages of aggregation  is correlated  to their toxicity.  These findings demonstrate  not  only that  the N-terminus of the Aß peptide  plays a crucial  role  in its aggregation  and  toxicity  but  also  suggest that this  region  of Aß  may  modulate  in vivo toxicity  by altering  the conformations of aggregates that  it forms.

  • 23.
    Brorsson, Ann-Christin
    et al.
    Department of Biochemistry, Umeå University.
    Kjellson, Annika
    Department of Biochemistry, Umeå University.
    Aronsson, Göran
    Biopool AB.
    Sethson, Ingmar
    Department of Organic Chemistry, Umeå University.
    Hambraeus, Charlotta
    University of Southern Stockholm.
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    The "two-state folder" MerP forms partially unfolded structures that show temperature dependent hydrogen exchange2004In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 340, no 2, p. 333-344Article in journal (Refereed)
    Abstract [en]

    We have analysed the folding energy landscape of the 72 amino acid protein MerP by monitoring native state hydrogen exchange as a function of temperature in the range of 7-55°C. The temperature dependence of the hydrogen exchange has allowed us to determine ΔG, ΔH and ΔCp values for the conformational processes that permit hydrogen exchange. When studied with the traditional probes, fluorescence and CD, MerP appears to behave as a typical two-state protein, but the results from the hydrogen exchange analysis reveal a much more complex energy landscape. Analysis at the individual amino acid level show that exchange is allowed from an ensemble of partially unfolded structures (i.e. intermediates) in which the stabilities at the amino acid level form a broad distribution throughout the protein. The formation of partially unfolded structures might contribute to the unusually slow folding of MerP. © 2004 Elsevier Ltd. All rights reserved.

  • 24.
    Brorsson, Ann-Christin
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Kumita, Janet R.
    University of Cambridge.
    MacLeod, Ian
    University of Cambridge.
    Bolognesi, Benedetta
    University of Cambridge.
    Speretta, Elena
    University of Cambridge.
    Luheshi, Leila M.
    University of Cambridge.
    Knowles, Tuomas P.
    University of Cambridge.
    Dobson, Christopher M.
    University of Cambridge.
    Crowther, Damian C.
    University of Cambridge.
    Methods and models in neurodegenerative and systemic protein aggregation diseases2010In: Frontiers in bioscience : a journal and virtual library, ISSN 1093-4715, Vol. 15, p. 373-396Article, review/survey (Refereed)
    Abstract [en]

    Protein misfolding and aggregation are implicated in a wide range of increasingly prevalent human diseases ranging from dementia to diabetes. In this review we discuss the current experimental strategies that are being employed in the investigation of the pathogenesis of three important protein misfolding disorders. The first, Alzheimers disease (AD), is the most prevalent neurodegenerative disease and is thought to be initiated by the aggregation of a natively unstructured peptide called amyloid beta (Abeta). We discuss methods for the characterization of the aggregation properties of Abeta in vitro and how the results of such experiments can be correlated with data from animal models of disease. We then consider another form of amyloidosis, where a systemic distribution of amyloid deposit is caused by aggregation and deposition of mutational variants of lysozyme. We describe how experiments in vitro, and more recently in vivo, have provided insights into the origins of this disease. Finally we outline the varied paradigms that have been employed in the study of the serpinopathies, and in particular, a dementia caused by neuroserpin polymerization.

  • 25.
    Brorsson, Ann-Christin
    et al.
    Department of Biochemistry, Umeå University.
    Lundqvist, Martin
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Sethson, Ingmar
    Department of Organic Chemistry Umeå University.
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    GuHCl and NaCl-dependent hydrogen exchange in MerP reveals a well-defined core with an unusual exchange pattern2006In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 357, no 5, p. 1634-1646Article in journal (Refereed)
    Abstract [en]

    We have analysed hydrogen exchange at amide groups to characterise the energy landscape of the 72 amino acid residue protein MerP. From the guanidine hydrochloride (GuHCl) dependence of exchange in the pre-transitional region we have determined free energy values of exchange (ΔGHX) and corresponding m-values for individual amide protons. Detailed analysis of the exchange patterns indicates that for one set of amide protons there is a weak dependence on denaturant, indicating that the exchange is dominated by local fluctuations. For another set of amide protons a linear, but much stronger, denaturant dependence is observed. Notably, the plots of free energy of exchange versus [GuHCl] for 16 amide protons show pronounced upward curvature, and a close inspection of the structure shows that these residues form a well-defined core in the protein. The hydrogen exchange that was measured at various concentrations of NaCl shows an apparent selective stabilisation of this core. Detailed analysis of this exchange pattern indicates that it may originate from selective destabilisation of the unfolded state by guanidinium ions and/or selective stabilisation of the core in the native state by chloride ions. © 2006 Elsevier Ltd. All rights reserved.

  • 26.
    Carlsson, Uno
    et al.
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Hammarström, Per
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Lindgren, M
    Persson, M
    Freskgård, Per-Ola
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology.
    Mårtensson, Lars-Göran
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Andersson, D
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Svensson, Magdalena
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Aggregation is site-specific in carbonic anhydrase and is prevented by GroEL: The interaction leads to a more flexible structure of both the protein substrate and the chaperonin.2000In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 78, no 1, p. 202Pos-Conference paper (Other academic)
  • 27.
    Chi, Celestine N
    et al.
    University of Copenhagen.
    Engstrom, Ake
    University of Copenhagen.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Ferguson, Neil
    University College Dublin.
    Jemth, Per
    Uppsala University.
    Biophysical Characterization of the Complex between Human Papillomavirus E6 Protein and Synapse-associated Protein 972011In: JOURNAL OF BIOLOGICAL CHEMISTRY, ISSN 0021-9258, Vol. 286, no 5, p. 3597-3606Article in journal (Refereed)
    Abstract [en]

    The E6 protein of human papillomavirus (HPV) exhibits complex interaction patterns with several host proteins, and their roles in HPV-mediated oncogenesis have proved challenging to study. Here we use several biophysical techniques to explore the binding of E6 to the three PDZ domains of the tumor suppressor protein synapse-associated protein 97 (SAP97). All of the potential binding sites in SAP97 bind E6 with micromolar affinity. The dissociation rate constants govern the different affinities of HPV16 and HPV18 E6 for SAP97. Unexpectedly, binding is not mutually exclusive, and all three PDZ domains can simultaneously bind E6. Intriguingly, this quaternary complex has the same apparent hydrodynamic volume as the unliganded PDZ region, suggesting that a conformational change occurs in the PDZ region upon binding, a conclusion supported by kinetic experiments. Using NMR, we discovered a new mode of interaction between E6 and PDZ: a subset of residues distal to the canonical binding pocket in the PDZ(2) domain exhibited noncanonical interactions with the E6 protein. This is consistent with a larger proportion of the protein surface defining binding specificity, as compared with that reported previously.

  • 28.
    Chi, Celestine N.
    et al.
    Department of Medical Biochemistry and Microbiology, Uppsala University.
    Haq, S. Raza
    Department of Medical Biochemistry and Microbiology, Uppsala University.
    Rinaldo, Serena
    University of Rome, Italy.
    Dogan, Jakob
    Department of Medical Biochemistry and Microbiology, Uppsala University.
    Cutruzzolà, Francesca
    University of Rome, Italy.
    Engström, Åke
    Department of Medical Biochemistry and Microbiology, Uppsala University.
    Gianni, Stefano
    University of Rome, Italy.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Jemth, Per
    Department of Medical Biochemistry and Microbiology, Uppsala University.
    Interactions outside the Boundaries of the Canonical Binding Groove of a PDZ Domain Influence Ligand Binding2012In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 51, no 44, p. 8971-8979Article in journal (Refereed)
    Abstract [en]

    The postsynaptic density protein-95/discs large/zonula occludens-1 (PDZ) domain is a protein-protein interaction module with a shallow binding groove where protein ligands bind. However, interactions that are not part of this canonical binding groove are likely to modulate peptide binding. We have investigated such interactions beyond the binding groove for PDZ3 from PSD-95 and a peptide derived from the C-terminus of the natural ligand CRIPT. We found via nuclear magnetic resonance experiments that up to eight residues of the peptide ligand interact with the PDZ domain, showing that the interaction surface extends far outside of the binding groove as defined by the crystal structure. PDZ3 contains an extra structural element, a C-terminal helix (α3), which is known to affect affinity. Deletion of this helix resulted in the loss of several intermolecular nuclear Overhauser enhancements from peptide residues outside of the binding pocket, suggesting that α3 forms part of the extra binding surface in wild-type PDZ3. Site-directed mutagenesis, isothermal titration calorimetry, and fluorescence intensity experiments confirmed the importance of both α3 and the N-terminal part of the peptide for the affinity. Our data suggest a general mechanism in which different binding surfaces outside of the PDZ binding groove could provide sites for specific interactions.

  • 29.
    Chilkova, O.
    et al.
    Dept. of Med. Biochem./Biophysics, Umeå University, SE-901 87 Umeå, Sweden.
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Johansson, E.
    Dept. of Med. Biochem./Biophysics, Umeå University, SE-901 87 Umeå, Sweden.
    The quaternary structure of DNA polymerase e from Saccharomyces cerevisiae2003In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 278, no 16, p. 14082-14086Article in journal (Refereed)
    Abstract [en]

    DNA polymerase e (Pol e) trom Saccharomyces cerevisiae consists of four subunits (Pol2, Dpb2, Dpb3, and Dpb4) and is essential for chromosomal DNA replication. Biochemical characterizations of Pol e have been cumbersome due to protease sensitivity and the limited amounts of Pol e in cells. We have developed a protocol for overexpression and purification of Pol e from S. cerevisiae. The native four-subunit complex was purified to homogeneity by conventional chromatography. Pol e was characterized biochemically by sedimentation velocity experiments and gel filtration experiments. The stoichiometry of the four subunits was estimated to be 1:1:1:1 from colloidal Coomassie-stained gels. Based on the sedimentation coefficient (11.9 S) and the Stokes radius (74.5 Å), a molecular mass for Pol e of 371 kDa was calculated, in good agreement with the calculated molecular mass of 379 kDa for a heterotetramer. Furthermore, analytical equilibrium ultracentrifugation experiments support the proposed heterotetrameric structure of Pol e. Thus, both DNA polymerase d and Pol e are purified as monomeric complexes, in agreement with accumulating evidence that Pol d and Pol e are located on opposite strands of the eukaryotic replication fork.

  • 30.
    Dzikaite, Vijole
    et al.
    Rheumatology Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden.
    Strandberg, Linn S
    Rheumatology Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden.
    Ambrosi, Aurelie
    Rheumatology Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden.
    Jagodic, Maja
    Neuroimmunology Unit, Department of Clinical Neuroscience, Karolinska Institute, Stockholm, Sweden.
    Janson, Peter
    Clinical Allergy Research Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden.
    Khademi, Mohsen
    Neuroimmunology Unit, Department of Clinical Neuroscience, Karolinska Institute, Stockholm, Sweden.
    Salomonsson, Stina
    Rheumatology Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden.
    Ottosson, Lars
    Rheumatology Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden.
    Klauninger, Robert
    Rheumatology Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden.
    Åden, Ulrika
    Department of Woman and Child Health, Karolinska Institute, Stockholm, Sweden.
    Sonesson, Sven-Erik
    Department of Woman and Child Health, Karolinska Institute, Stockholm, Sweden.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    de Graaf, Katrien L
    Hertie Institute for clinical Brain Research, University of Tübingen, Germany.
    Kuchroo, Vijay K
    Center for Neurological Diseases, Brigham and Women’s Hospital, Harvard Medical School, Boston, USA,.
    Achour, Adnane
    Center for Infectious Medicine, Department of Medicine, Karolinska Institutet, Stockholm, Sweden.
    Winqvist, Ola
    Clinical Allergy Research Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden.
    Olsson, Tomas
    Neuroimmunology Unit, Department of Clinical Neuroscience, Karolinska Institute, Stockholm, Sweden.
    Wahren-Herlenius, Marie
    Rheumatology Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden.
    MHC Genes Determine Fetal Susceptibility in a Rat Model of Congenital Heart Block2010In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 72, no 3, p. 269-269Article in journal (Other academic)
    Abstract [en]

    n/a

  • 31.
    Ek, Anders
    et al.
    Swedish Biogas International Korea Co.
    Hallin, Sara
    Svensk Biogas FoU.
    Vallin, Lina
    Svensk Biogas FoU.
    Schnurer, Anna
    Swedish University of Agricultural Sciences, Uppsala.
    Karlsson, Martin
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Slaughterhouse waste co-digestion - Experiences from 15 years of full-scale operation2011In: World Renewable Energy Congress - Sweden 8-13 May, 2011: Volume 1 (Bioenergy Technology) / [ed] Bahram Moshfegh, Linköping University: Linköping University Electronic Press, 2011, Vol. 009, p. 64-71Conference paper (Refereed)
    Abstract [en]

    At Tekniska Verken in Linköping AB (TVAB) there is a long time experience of handling and producing biogas from large volumes of slaughterhouse waste. Experiences from research and development and plant operations have lead to the implementation of several process improving technological/biological solutions. We can in this paper describe how the improvements have had several positive effects on the process, including energy savings, better odor control, higher gas quality, increased organic loading rates and higher biogas production with maintained process stability. In addition, it is described how much of the process stability in anaerobic digestion of slaughter house waste relates to the plant operation, which allow the microbiological consortia to adapt to the substrate. Since digestion of proteinaceous substrates like slaughterhouse waste lead to high ammonia loads, special requirements in ammonia tolerance are placed on the microbiota of the anaerobic digestion. Biochemical assays revealed that the main route for methane production proceed through syntrophic acetate oxidation, which require longer retention times than methane production by acetoclastic methanogens. Thus, the long retention time of the plant, accomplished by a low dilution of the substrate, is a vital component of the process stability when treating high protein substrates like slaughterhouse waste.

  • 32.
    Espinosa, Alexander
    et al.
    Rheumatology Unit, Department of Medicine, CMM L8:04, Karolinska Institutet, SE-171 76 Stockholm, Sweden.
    Hennig, Janosch
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Ambrosi, Aurélie
    Rheumatology Unit, Department of Medicine, CMM L8:04, Karolinska Institutet, SE-171 76 Stockholm, Sweden.
    Anandapadamanaban, Madhanagopal
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Sandberg Abelius, Martina
    Linköping University, Department of Clinical and Experimental Medicine, Division of Microbiology and Molecular Medicine. Linköping University, Faculty of Medicine and Health Sciences.
    Sheng, Yi
    Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, Canada.
    Nyberg, Filippa
    Department of Clinical Sciences at Danderyd Hospital, Karolinska Institute, Stockholm, Sweden.
    Arrowsmith, Cheryl H.
    Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, Canada.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Wahren-Herlenius, Marie
    Rheumatology Unit, Department of Medicine, CMM L8:04, Karolinska Institutet, SE-171 76 Stockholm, Sweden.
    Anti-Ro52 Autoantibodies from Patients with Sjögren's Syndrome Inhibit the Ro52 E3 Ligase Activity by Blocking the E3/E2 Interface2011In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 286, no 42, p. 36478-36491Article in journal (Refereed)
    Abstract [en]

    Ro52 (TRIM21) is an E3 ligase of the tripartite motif family that negatively regulates proinflammatory cytokine production by ubiquitinating transcription factors of the interferon regulatory factor family. Autoantibodies to Ro52 are present in patients with lupus and Sjögren's syndrome, but it is not known if these autoantibodies affect the function of Ro52. To address this question, the requirements for Ro52 E3 ligase activity were first analyzed in detail. Scanning a panel of E2 ubiquitin-conjugating enzymes, we found that UBE2D1–4 and UBE2E1–2 supported the E3 ligase activity of Ro52 and that the E3 ligase activity of Ro52 was dependent on its RING domain. We also found that the N-terminal extensions in the class III E2 enzymes affected their interaction with Ro52. Although the N-terminal extension in UBE2E3 made this E2 enzyme unable to function together with Ro52, the N-terminal extensions in UBE2E1 and UBE2E2 allowed for a functional interaction with Ro52. Anti-Ro52-positive patient sera and affinity-purified anti-RING domain autoantibodies inhibited the E3 activity of Ro52 in ubiquitination assays. Using NMR, limited proteolysis, ELISA, and Ro52 mutants, we mapped the interactions between Ro52, UBE2E1, and anti-Ro52 autoantibodies. We found that anti-Ro52 autoantibodies inhibited the E3 ligase activity of Ro52 by sterically blocking the E2/E3 interaction between Ro52 and UBE2E1. Our data suggest that anti-Ro52 autoantibodies binding the RING domain of Ro52 may be actively involved in the pathogenesis of rheumatic autoimmune disease by inhibiting Ro52-mediated ubiquitination.

  • 33.
    Fernandes, Aristi Potamitou
    et al.
    Karolinska Institutet, Stockholm.
    Fladvad, Malin
    Karolinska Institutet, Stockholm.
    Berndt, Carsten
    Karolinska Institutet, Stockholm.
    Andrésen, Cecilia
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Lillig, Christopher Horst
    Karolinska Institutet, Stockholm.
    Neubauer, Peter
    University of Oulu, Finland.
    Sunnerhagen, Maria
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Holmgren, Arne
    Karolinska Institutet, Stockholm.
    Vlamis-Gardikas, Alexios
    Karolinska Institutet, Stockholm.
    A Novel Monothiol Glutaredoxin (Grx4) from Escherichia coli Can Serve as a Substrate for Thioredoxin Reductase2005In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 280, no 26, p. 24544-24552Article in journal (Refereed)
    Abstract [en]

    Glutaredoxins are ubiquitous proteins that catalyze the reduction of disulfides via reduced glutathione (GSH). Escherichia coli has three glutaredoxins (Grx1, Grx2, and Grx3), all containing the classic dithiol active site CPYC. We report the cloning, expression, and characterization of a novel monothiol E. coli glutaredoxin, which we name glutaredoxin 4 (Grx4). The protein consists of 115 amino acids (12.7 kDa), has a monothiol (CGFS) potential active site and shows high sequence homology to the other monothiol glutaredoxins and especially to yeast Grx5. Experiments with gene knock-out techniques showed that the reading frame encoding Grx4 was essential. Grx4 was inactive as a GSH-disulfide oxidoreductase in a standard glutaredoxin assay with GSH and hydroxyethyl disulfide in a complete system with NADPH and glutathione reductase. An engineered CGFC active site mutant did not gain activity either. Grx4 in reduced form contained three thiols, and treatment with oxidized GSH resulted in glutathionylation and formation of a disulfide. Remarkably, this disulfide of Grx4 was a direct substrate for NADPH and E. coli thioredoxin reductase, whereas the mixed disulfide was reduced by Grx1. Reduced Grx4 showed the potential to transfer electrons to oxidized E. coli Grx1 and Grx3. Grx4 is highly abundant (750–2000 ng/mg of total soluble protein), as determined by a specific enzyme-link immunosorbent assay, and most likely regulated by guanosine 3′,5′-tetraphosphate upon entry to stationary phase. Grx4 was highly elevated upon iron depletion, suggesting an iron-related function for the protein.

  • 34.
    Fladvad, Malin
    et al.
    Karolinska Institutet, Stockholm.
    Bellanda, Massimo
    University of Padova, Italy.
    Fernandes, Aristi Potamitou
    Karolinska Institutet, Stockholm.
    Mammi, Stefano
    University of Padova, Italy.
    Vlamis-Gardikas, Alexios
    Karolinska Institutet, Stockholm.
    Holmgren, Arne
    Karolinska Institutet, Stockholm.
    Sunnerhagen, Maria
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Molecular Mapping of Functionalities in the Solution Structure of Reduced Grx4, a Monothiol Glutaredoxin from Escherichia coli2005In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 280, no 26, p. 24553-24561Article in journal (Refereed)
  • 35.
    Fladvad, Malin
    et al.
    Department of Medical Biochemistry and Biophysics Karolinska Institutet, Stockholm.
    Zhou, Kaison
    Molecular Biotechnology IFM, Linköping University.
    Moshref, Ahmed
    Department of Medical Biochemistry and Biophysics Karolinska Institutet, Stockholm.
    Pursglove, Sharon
    School of Molecular and Microbial Biosciences University of Sydney.
    Säfsten, Pär
    Biacore AB, Uppsala.
    Sunnerhagen, Maria
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    N and C-terminal Sub-regions in the c-Myc Transactivation Region and their Joint Role in Creating Versatility in Folding and Binding2005In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 246, p. 175-189Article in journal (Refereed)
  • 36.
    Fucile, Geoffrey
    et al.
    University of Toronto.
    Garcia, Christel
    University of Toronto.
    Carlsson, Jonas
    Linköping University, Department of Physics, Chemistry and Biology, Bioinformatics . Linköping University, The Institute of Technology.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Christendat, Dinesh
    University of Toronto.
    Structural and biochemical investigation of two Arabidopsis shikimate kinases: The heat-inducible isoform is thermostable2011In: Protein Science, ISSN 0961-8368, E-ISSN 1469-896X, Vol. 20, no 7, p. 1125-1136Article in journal (Refereed)
    Abstract [en]

    The expression of plant shikimate kinase (SK; EC 2.7.1.71), an intermediate step in the shikimate pathway to aromatic amino acid biosynthesis, is induced under specific conditions of environmental stress and developmental requirements in an isoform-specific manner. Despite their important physiological role, experimental structures of plant SKs have not been determined and the biochemical nature of plant SK regulation is unknown. The Arabidopsis thaliana genome encodes two SKs, AtSK1 and AtSK2. We demonstrate that AtSK2 is highly unstable and becomes inactivated at 37 degrees C whereas the heat-induced isoform, AtSK1, is thermostable and fully active under identical conditions at this temperature. We determined the crystal structure of AtSK2, the first SK structure from the plant kingdom, and conducted biophysical characterizations of both AtSK1 and AtSK2 towards understanding this mechanism of thermal regulation. The crystal structure of AtSK2 is generally conserved with bacterial SKs with the addition of a putative regulatory phosphorylation motif forming part of the adenosine triphosphate binding site. The heat-induced isoform, AtSK1, forms a homodimer in solution, the formation of which facilitates its relative thermostability compared to AtSK2. In silico analyses identified AtSK1 site variants that may contribute to AtSK1 stability. Our findings suggest that AtSK1 performs a unique function under heat stress conditions where AtSK2 could become inactivated. We discuss these findings in the context of regulating metabolic flux to competing downstream pathways through SK-mediated control of steady state concentrations of shikimate.

  • 37.
    Gertow, K.
    et al.
    Department of Medicine, Atherosclerosis Research Unit, King Gustaf V Research Institute, S-171 76 Stockholm, Sweden.
    Bellanda, M.
    Department of Organic Chemistry, University of Padova, 35131 Padova, Italy.
    Eriksson, P.
    Department of Medicine, Atherosclerosis Research Unit, King Gustaf V Research Institute, S-171 76 Stockholm, Sweden.
    Boquist, S.
    Department of Medicine, Atherosclerosis Research Unit, King Gustaf V Research Institute, S-171 76 Stockholm, Sweden.
    Hamsten, A.
    Department of Medicine, Atherosclerosis Research Unit, King Gustaf V Research Institute, S-171 76 Stockholm, Sweden.
    Sunnerhagen, Maria
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Fisher, R.M.
    Department of Medicine, Atherosclerosis Research Unit, King Gustaf V Research Institute, S-171 76 Stockholm, Sweden, King Gustaf V Research Institute, Karolinska Hospital, S-171 76 Stockholm, Sweden.
    Genetic and Structural Evaluation of Fatty Acid Transport Protein-4 in Relation to Markers of the Insulin Resistance Syndrome2004In: Journal of Clinical Endocrinology and Metabolism, ISSN 0021-972X, E-ISSN 1945-7197, Vol. 89, no 1, p. 392-399Article in journal (Refereed)
    Abstract [en]

    Disturbances in fatty acid metabolism are involved in the etiology of insulin resistance and the related dyslipidemia, hypertension, and procoagulant state. The fatty acid transport proteins (FATPs) are implicated in facilitated cellular uptake of nonesterified fatty acids (NEFAs), thus potentially regulating NEFA concentrations and metabolism. The aim of this study was to investigate polymorphic loci in the FATP4 gene with respect to associations with fasting and postprandial lipid and lipoprotein variables and markers of insulin resistance in 608 healthy, middle-aged Swedish men and to evaluate possible mechanisms behind any associations observed. Heterozygotes for a Gly209Ser polymorphism (Ser allele frequency 0.05) had significantly lower body mass index and, correcting for body mass index, significantly lower triglyceride concentrations, systolic blood pressure, insulin concentrations, and homeostasis model assessment index compared with common homozygotes. A three-dimensional model of the FATP4 protein based on structural and functional similarity with adenylate-forming enzymes revealed that the variable residue 209 is exposed in a region potentially involved in protein-protein interactions. Furthermore, the model indicated functional regions with respect to NEFA transport and acyl-coenzyme A synthase activity and membrane association. These findings propose FATP4 as a candidate gene for the insulin resistance syndrome and provide a structural basis for understanding FATP function in NEFA transport and metabolism.

  • 38.
    Ghasriani, Houman
    et al.
    University of Ottawa.
    Ducat, Thierry
    University of Ottawa.
    Hart, Chris T
    University of Ottawa.
    Hafizi, Fatima
    University of Ottawa.
    Chang, Nina
    University of Ottawa.
    Al-Baldawi, Ali
    University of Ottawa.
    Ayed, Saud H
    University of Ottawa.
    Lundström, Patrik
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Dillon, Jo-Anne R
    University of Saskatchewan.
    Goto, Natalie K
    University of Ottawa.
    Appropriation of the MinD protein-interaction motif by the dimeric interface of the bacterial cell division regulator MinE2010In: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, ISSN 0027-8424, Vol. 107, no 43, p. 18416-18421Article in journal (Refereed)
    Abstract [en]

    MinE is required for the dynamic oscillation of Min proteins that restricts formation of the cytokinetic septum to the midpoint of the cell in gram negative bacteria. Critical for this oscillation is MinD-binding by MinE to stimulate MinD ATP hydrolysis, a function that had been assigned to the first similar to 30 residues in MinE. Previous models based on the structure of an autonomously folded dimeric C-terminal fragment suggested that the N-terminal domain is freely accessible for interactions with MinD. We report here the solution NMR structure of the full-length MinE dimer from Neisseria gonorrhoeae, with two parts of the N-terminal domain forming an integral part of the dimerization interface. Unexpectedly, solvent accessibility is highly restricted for residues that were previously hypothesized to directly interact with MinD. To delineate the true MinD-binding region, in vitro assays for MinE-stimulated MinD activity were performed. The relative MinD-binding affinities obtained for full-length and N-terminal peptides from MinE demonstrated that residues that are buried in the dimeric interface nonetheless participate in direct interactions with MinD. According to results from NMR spin relaxation experiments, access to these buried residues may be facilitated by the presence of conformational exchange. We suggest that this concealment of MinD-binding residues by the MinE dimeric interface provides a mechanism for prevention of nonspecific interactions, particularly with the lipid membrane, to allow the free diffusion of MinE that is critical for Min protein oscillation.

  • 39.
    Gustafsson, Robert
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology.
    Biophysical characterization of the *5 protein variant of human thiopurine methyltransferase by NMR spectroscopy2012Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Human thiopurine methyltransferase (TPMT) is an enzyme involved in the metabolism of thiopurine drugs, which are widely used in leukemia and inflammatory bowel diseases such as ulcerative colitis and Crohn´s disease. Due to genetic polymorphisms, approximately 30 protein variants are present in the population, some of which have significantly lowered activity. TPMT *5 (Leu49Ser) is one of the protein variants with almost no activity. The mutation is positioned in the hydrophobic core of the protein, close to the active site.

    Hydrogen exchange rates measured with NMR spectroscopy for N-terminally truncated constructs of TPMT *5 and TPMT *1 (wild type) show that local stability and hydrogen bonding patterns are changed by the mutation Leu49Ser. Most residues exhibit faster exchange rates and a lower local stability in TPMT *5 in comparison with TPMT *1. Changes occur close to the active site but also throughout the entire protein. Calculated overall stability is similar for the two constructs, so the measured changes are due to local stability.

    Protein dynamics measured with NMR relaxation experiments show that both TPMT *5 and TPMT *1 are monomeric in solution. Millisecond dynamics exist in TPMT *1 but not in TPMT *5, even though a few residues exhibit a faster dynamic. Dynamics on nanosecond to picosecond time scale have changed but no clear trends are observable.

  • 40.
    Göransson, Anna-Lena
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    The Alzheimer Aβ Peptide: Identification of Properties Distinctive for Toxic Prefibrillar Species2012Licentiate thesis, comprehensive summary (Other academic)
    Abstract [en]

    Proteins must have specific conformations to function correctly inside cells. However, sometimes they adopt the wrong conformation, causing dysfunction and disease. A number of amyloid diseases are caused by misfolded proteins that form amyloid fibrils. One such disease is Alzheimer’s disease (AD). The protein involved in this deadly disease is the amyloid β (Aβ) peptide. The formation of soluble prefibrillar oligomeric Aβ species has been recognized as an important factor in the development of AD. The aim of work described in this thesis was to investigate which properties of these oligomeric species can be linked to toxicity. We approached this task by comparing the aggregation behavior and biophysical properties of aggregates formed by variants of the Aβ peptide that have been shown to differ in neurotoxicity when expressed in the central nervous system (CNS) of Drosophila melanogaster. A combined set involving different fluorescent probes was used in parallell with transmission electron microscopy. The toxicity of species formed during the aggregation process was examined by exposing human SH-SY5Y neuroblastoma cells to Aβ aggregates. We deduced that there is a correlation between cell toxicity and the propensity of the Aβ peptide to form small prefibrillar assemblies at an early stage of aggregation in vitro. Moreover, these prefibrillar species were characterized by their ability to be recognized by pentamer formyl thiophene acetic acid (p-FTAA) and the presence of exposed hydrophobic patches. We also found that larger aggregates did not induce cell death.

    List of papers
    1. Identification of distinct physiochemical properties of the toxic prefibrillar species formed by Aβ peptide variants
    Open this publication in new window or tab >>Identification of distinct physiochemical properties of the toxic prefibrillar species formed by Aβ peptide variants
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    2012 (English)In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 420, no 4, p. 895-900Article in journal (Refereed) Published
    Abstract [en]

    The formation of amyloid-β peptide (Aβ) aggregates at an early stage during the self-assembly process is an important factor in the development of Alzheimer’s disease. The toxic effect is believed to be exerted by prefibrillar species of Aβ. It is therefore important to identify which prefibrillar species are toxic and characterize their distinct properties. In the present study, we investigated the in vitro aggregation behavior of Aβ-derived peptides possessing different levels of neurotoxic activity, using fluorescence spectroscopy in combination with transmission electron microscopy. The toxicity of various Aβ aggregates was assessed by using cultures of human neuroblastoma cells. Through combined use of the fluorescence probe 8-anilino-1-napthalenesulfonate (ANS) and the novel luminescent probe pentamer formyl thiophene acetic acid (p-FTAA), we were able to identify those Aβ peptide-derived prefibrillar species which exhibited cellular toxicity. In particular, species, which formed early during the aggregation process and showed strong p-FTAA and ANS fluorescence, were the species that possessed toxic activities. Moreover, by manipulating the aggregation conditions, it was possible to change the capacity of the Aβ peptide to form nontoxic versus toxic species.

    Place, publisher, year, edition, pages
    Wiley-Blackwell, 2012
    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-73185 (URN)10.1016/j.bbrc.2012.03.097 (DOI)000303619100034 ()
    Note

    funding agencies|Swedish National Graduate School in Science, Technology and Mathematics Education Research (Fon-tD)||Swedish Alzheimers Foundation||Soderberg foundation||

    Available from: 2011-12-21 Created: 2011-12-21 Last updated: 2017-12-08Bibliographically approved
    2. Dissecting the Aggregation Events of Alzheimer’s disease Associated Aβ peptide Variants by the Combined use of Different Fluorescent Probes
    Open this publication in new window or tab >>Dissecting the Aggregation Events of Alzheimer’s disease Associated Aβ peptide Variants by the Combined use of Different Fluorescent Probes
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    The formation of soluble prefibrillar oligomeric species of the amyloid β peptide (Aβ) has been implicated as a causative agent in the development of Alzheimer’s disease (AD). It is therefore important to characterize the properties of these aggregates, which precede the formation of amyloid fibrils. We studied the in vitro aggregation process of two Aβ40 peptide variants through the combined use of four different fluorescent probes and transmission electron microscopy. Previous studies have shown that these two studied Aβ40 variants exhibit different levels of neurodegeneration when expressed in the central nervous system of Drosophila melanogaster. In the present study, we demonstrate distinct differences in aggregate morphology and their binding properties to different fluorescent probes during in vitro fibrillation of these Aβ peptides. Our results indicate a potential link between the observed neurodegenerative properties and the biophysical properties of distinct aggregated Aβ species.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-76739 (URN)
    Available from: 2012-04-18 Created: 2012-04-18 Last updated: 2014-04-08
  • 41.
    Göransson, Anna-Lena
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Kanmert, Daniel
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Nilsson, K. Peter R.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Kågedal, Katarina
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Experimental Pathology.
    Brorsson, Ann-Christin
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Identification of distinct physiochemical properties of the toxic prefibrillar species formed by Aβ peptide variants2012In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 420, no 4, p. 895-900Article in journal (Refereed)
    Abstract [en]

    The formation of amyloid-β peptide (Aβ) aggregates at an early stage during the self-assembly process is an important factor in the development of Alzheimer’s disease. The toxic effect is believed to be exerted by prefibrillar species of Aβ. It is therefore important to identify which prefibrillar species are toxic and characterize their distinct properties. In the present study, we investigated the in vitro aggregation behavior of Aβ-derived peptides possessing different levels of neurotoxic activity, using fluorescence spectroscopy in combination with transmission electron microscopy. The toxicity of various Aβ aggregates was assessed by using cultures of human neuroblastoma cells. Through combined use of the fluorescence probe 8-anilino-1-napthalenesulfonate (ANS) and the novel luminescent probe pentamer formyl thiophene acetic acid (p-FTAA), we were able to identify those Aβ peptide-derived prefibrillar species which exhibited cellular toxicity. In particular, species, which formed early during the aggregation process and showed strong p-FTAA and ANS fluorescence, were the species that possessed toxic activities. Moreover, by manipulating the aggregation conditions, it was possible to change the capacity of the Aβ peptide to form nontoxic versus toxic species.

  • 42.
    Göransson, Anna-Lena
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Otieno, Mildred
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Nilsson, Peter
    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.
    Nilsson, K. Peter R
    Linköping University, Department of Physics, Chemistry and Biology, Biomolecular and Organic Electronics. Linköping University, The Institute of Technology.
    Brorsson, Ann-Christin
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Dissecting the Aggregation Events of Alzheimer’s disease Associated Aβ peptide Variants by the Combined use of Different Fluorescent ProbesManuscript (preprint) (Other academic)
    Abstract [en]

    The formation of soluble prefibrillar oligomeric species of the amyloid β peptide (Aβ) has been implicated as a causative agent in the development of Alzheimer’s disease (AD). It is therefore important to characterize the properties of these aggregates, which precede the formation of amyloid fibrils. We studied the in vitro aggregation process of two Aβ40 peptide variants through the combined use of four different fluorescent probes and transmission electron microscopy. Previous studies have shown that these two studied Aβ40 variants exhibit different levels of neurodegeneration when expressed in the central nervous system of Drosophila melanogaster. In the present study, we demonstrate distinct differences in aggregate morphology and their binding properties to different fluorescent probes during in vitro fibrillation of these Aβ peptides. Our results indicate a potential link between the observed neurodegenerative properties and the biophysical properties of distinct aggregated Aβ species.

  • 43.
    Hammarström, Per
    et al.
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Protein denaturation and the denatured state2005In: Encyclopedia of Life Sciences, Wiley-Blackwell , 2005Chapter in book (Other (popular science, discussion, etc.))
    Abstract [en]

    Protein denaturation experiments are routinely used to determine protein stability and to elucidate structural and dynamic effects of mutations, cofactors and ligands. Denatured states of proteins have gained wide interest in recent years owing to their fundamental importance in a wide variety of phenomena such as deciphering the protein folding problem and the molecular understanding of many diseases.

  • 44.
    Hammarström, Per
    et al.
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Kalman, B.
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Carlsson, Uno
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Pyrene Excimer Fluorescence as a Proximity Probe for Investigation of Residual Structure in the Unfolded State of Human Carbonic Anhydrase II1997In: FEBS Letters, ISSN 0014-5793, E-ISSN 1873-3468, Vol. 420, p. 63-68Article in journal (Refereed)
  • 45.
    Hammarström, Per
    et al.
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Persson, Malin
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Freskgård, Per-Ola
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Mårtensson, Lars-Göran
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Andersson, D.
    Jonsson, Bengt-Harald
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology .
    Carlsson, Uno
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Biochemistry.
    Structural mapping of an aggregation nucleation site in a molten-globule intermediate1999In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 274, p. 32897-32903Article in journal (Refereed)
  • 46.
    Hansen, Alexandar L
    et al.
    University of Toronto.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Velyvis, Algirdas
    University of Toronto.
    Kay, Lewis E
    University of Toronto.
    Quantifying Millisecond Exchange Dynamics in Proteins by CPMG Relaxation Dispersion NMR Using Side-Chain H-1 Probes2012In: Journal of the American Chemical Society, ISSN 0002-7863, E-ISSN 1520-5126, Vol. 134, no 6, p. 3178-3189Article in journal (Refereed)
    Abstract [en]

    A Carr-Purcell-Meiboom-Gill relaxation dispersion experiment is presented for quantifying millisecond time-scale chemical exchange at side-chain H-1 positions in proteins. Such experiments are not possible in a fully protonated molecule because of magnetization evolution from homonuclear scalar couplings that interferes with the extraction of accurate transverse relaxation rates. It is shown, however, that by using a labeling strategy whereby proteins are produced using {C-13,H-1}-glucose and D2O a significant number of isolated side-chain H-1 spins are generated, eliminating such effects. It thus becomes possible to record H-1 dispersion profiles at the beta positions of Ass, Cys, Ser, His, Phe, Tyr, and Trp as well as the gamma positions of Glx, in addition to the methyl side-chain moieties. This brings the total of amino acid side-chain positions that can be simultaneously probed using a single H-1 dispersion experiment to 16. The utility of the approach is demonstrated with an application to the four-helix bundle colicin E7 immunity protein, Im7, which folds via a partially structured low populated intermediate that interconverts with the folded, ground state on the millisecond time-scale. The extracted H-1 chemical shift differences at side-chain positions provide valuable restraints in structural studies of invisible, excited states, complementing backbone chemical shifts that are available from existing relaxation dispersion experiments.

  • 47.
    Haq, S Raza
    et al.
    Uppsala University.
    Chi, Celestine N
    Uppsala University.
    Bach, Anders
    University of Copenhagen.
    Dogan, Jakob
    Uppsala University.
    Engstrom, Ake
    Uppsala University.
    Hultqvist, Greta
    Uppsala University.
    Karlsson, O Andreas
    Uppsala University.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Montemiglio, Linda C
    University of Roma La Sapienza.
    Stromgaard, Kristian
    University of Copenhagen.
    Gianni, Stefano
    University of Roma La Sapienza.
    Jemth, Per
    Uppsala University.
    Side-Chain Interactions Form Late and Cooperatively in the Binding Reaction between Disordered Peptides and PDZ Domains2012In: Journal of the American Chemical Society, ISSN 0002-7863, E-ISSN 1520-5126, Vol. 134, no 1, p. 599-605Article in journal (Refereed)
    Abstract [en]

    Intrinsically disordered proteins are very common and mediate numerous protein-protein and protein-DNA interactions. While it is clear that these interactions are instrumental for the life of the mammalian cell, there is a paucity of data regarding their molecular binding mechanisms. Here we have used short peptides as a model system for intrinsically disordered proteins. Linear free energy relationships based on rate and equilibrium constants for the binding of these peptides to ordered target proteins, PDZ domains, demonstrate that native side-chain interactions form mainly after the rate-limiting barrier for binding and in a cooperative fashion. This finding suggests that these disordered peptides first form a weak encounter complex with non-native interactions. The data do not support the recent notion that the affinities of intrinsically disordered proteins toward their targets are generally governed by their association rate constants. Instead, we observed the opposite for peptide-PDZ interactions, namely, that changes in K-d correlate with changes in k(off).

  • 48. Hellman, Jan
    et al.
    Ek, Anders
    Sundberg, Carina
    Linköping University, The Tema Institute, Department of Water and Environmental Studies. Linköping University, Faculty of Arts and Sciences.
    Johansson, Mariana
    Svensson, Bo
    Linköping University, The Tema Institute, Department of Water and Environmental Studies. Linköping University, Faculty of Arts and Sciences.
    Karlsson, Martin
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, The Institute of Technology.
    Mechanisms of increased methane production through re-circulation of magnetic biomass carriers in an experimental continuously stirred tank reactor2010Conference paper (Refereed)
    Abstract [en]

    Magnetite particles were used in a semi-continuous process as magnetic biomass carriers to separate and re-introduce microorganisms in a CSTR reactor. In comparison to a control reactor the methane content during the semi-continuous process was elevated when magnetite particles were used. The difference was most apparent during the fermentative step directly after feeding and upon direct spiking with volatile fatty acids. Total DNA quantification of the separated magnetite particles revealed high association of microorganisms. Furthermore, quantitative real-time PCR analysis of the associated microbial consortia indicated that the hydrogenotrophic Methanobacteriales was overrepresented at the particle surface. Thus, the increased methane production could be coupled to both the crowding and shorter interspecies distances between the groups involved in anaerobic digestion, as well as a preferential adsorption of hydrogenotrophs. By bringing the hydrogenotrophs closer to the primary fermentative bacteria and increasing their relative number the produced hydrogen during acidogenesis is more effectively utilized and more carbon dioxide is converted to methane. Furthermore, by the same cause, the rate of acetogenesis increased as the hydrogenotrophs more effectively could consume the hydrogen produced and thereby keep the hydrogen partial pressure low.

  • 49.
    Helmfors, Linda
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, Faculty of Science & Engineering.
    Boman, Andrea
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Civitelli, Livia
    Linköping University, Department of Clinical and Experimental Medicine, Division of Neuro and Inflammation Science. Linköping University, Faculty of Medicine and Health Sciences.
    Nath, Sangeeta
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Sandin, Linnea
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Janefjord, Camilla
    Linköping University, Department of Clinical and Experimental Medicine, Division of Neuro and Inflammation Science. Linköping University, Faculty of Medicine and Health Sciences.
    McCann, Heather
    Neuroscience Research Australia and University of New South Wales, Australia.
    Zetterberg, Henrik
    Clinical Neurochemistry Laboratory, Department of Neuroscience and Physiology, Sahlgrenska University Hospital, Mölndal, Sweden / UCL Institute of Neurology, Queen Square, London, United Kingdom.
    Blennow, Kaj
    Clinical Neurochemistry Laboratory, Department of Neuroscience and Physiology, Sahlgrenska University Hospital, Mölndal, Sweden.
    Halliday, Glenda
    UCL Institute of Neurology, Queen Square, London, United Kingdom.
    Brorsson, Ann-Christin
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology. Linköping University, Faculty of Science & Engineering.
    Kågedal, Katarina
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Protective properties of lysozyme on β-amyloid pathology: implications for Alzheimer disease2015In: Neurobiology of Disease, ISSN 0969-9961, E-ISSN 1095-953X, Vol. 83, p. 122-133Article in journal (Refereed)
    Abstract [en]

    The hallmarks of Alzheimer disease are amyloid-β plaques and neurofibrillary tangles accompanied by signs of neuroinflammation. Lysozyme is a major player in the innate immune system and has recently been shown to prevent the aggregation of amyloid-β1-40 in vitro. In this study we found that patients with Alzheimer disease have increased lysozyme levels in the cerebrospinal fluid and lysozyme co-localized with amyloid-β in plaques. In Drosophila neuronal co-expression of lysozyme and amyloid-β1-42 reduced the formation of soluble and insoluble amyloid-β species, prolonged survival and improved the activity of amyloid-β1-42 transgenic flies. This suggests that lysozyme levels rise in Alzheimer disease as a compensatory response to amyloid-β increases and aggregation. In support of this, in vitro aggregation assays revealed that lysozyme associates with amyloid-β1-42 and alters its aggregation pathway to counteract the formation of toxic amyloid-β species. Overall, these studies establish a protective role for lysozyme against amyloid-β associated toxicities and identify increased lysozyme in patients with Alzheimer disease. Therefore, lysozyme has potential as a new biomarker as well as a therapeutic target for Alzheimer disease.

  • 50.
    Hennig, Janosch
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Biotechnology . Linköping University, The Institute of Technology.
    Structure-function studies on TRIM21/Ro52, a protein involved in autoimmune diseases2009Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Several members of the tripartite motif (TRIM) protein family are involvedin antiviral activity and immunity and have been linked to severaldiseases. TRIM21, the main object of this thesis, is involved in Sjögrensyndrome (SS) and systemic lupus erythematosus (SLE), where patientsoften have autoantibodies against different epitopes on TRIM21. Duringthe course of this study a role of TRIM21 in regulation of proinflammatorycytokines and autoimmunity emerged. The aim of this thesis is to providea better understanding of the structure-function relationship of TRIM21.A conformational epitope in the coiled-coil domain of TRIM21 has beencharacterized, whose autoantibodies cause congenital heart block. A widerange of biophysical methods were employed to establish a model of theprotein domain arrangement of TRIM21, and functional implications werederived. By sequence comparisons, TRIM proteins were classified into threesubgroups, sharing a conserved amphipathic helix in the region, linkingthe conserved N-terminal Zn2+-binding domains RING and B-box, calledthe RING-B-box linker (RBL). A structural dependence of this region on theRING has been observed and a model of the RING-RBL was derived frombioinformatics and proteolysis data. Anti-RING-RBL antibodies inhibit theE3 ligase activity of TRIM21 in ubiquitination. Interferon regulatory factors(IRFs), the substrate for TRIM21-dependent ubiquitination could thereforeretain their high cellular levels after stress-induced inflammation, increasingthe susceptibility to SS and SLE. According to NMR data, the antibodiesbind to the Zn2+-binding loop regions of the RING, which usually bind tothe E2 conjugating enzyme. Antibodies against the C-terminus of the RBLregion do not inhibit the E3 ligase activity.

    List of papers
    1. Structurally derived mutations define congenital heart block-related epitopes within the 200-239 amino acid stretch of the Ro52 protein
    Open this publication in new window or tab >>Structurally derived mutations define congenital heart block-related epitopes within the 200-239 amino acid stretch of the Ro52 protein
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    2005 (English)In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 61, no 2, p. 109-118Article in journal (Refereed) Published
    Abstract [en]

    Congenital heart block is a passively transferred autoimmune condition, which affects the children of mothers with Ro/SSA autoantibodies. During pregnancy, the antibodies are transported across the placenta and affect the fetus. We have previously demonstrated that antibodies directed to the 200-239 amino acid (aa) stretch of the Ro52 component of the Ro/SSA antigen correlate with the development of congenital heart block. In this report, we investigated the antibody-antigen interaction of this target epitope in detail at a molecular and structural level. Peptides representing aa 200-239 (p200) with structurally derived mutations were synthesized to define the epitopes recognized by two Ro52 human monoclonal antibodies, S3A8 and M4H1, isolated from patient-derived phage display libraries. Analyses by ELISA, circular dichroism and MALDI-TOF-MS demonstrate that the antibody recognition is dependent on a partly a-helical fold within the putative leucine zipper of the 200-239 aa stretch and that the two human anti-p200 monoclonal antibodies, M4H1 and S3A8, recognize different epitopic structures within the p200 peptide. In addition, we investigated the representation of each fine specificity within the sera of mothers with children born with congenital heart block, and in such sera, antibodies of the S3A8 idiotype were more commonly detected and at higher levels than M4H1-like antibodies.

    National Category
    Engineering and Technology
    Identifiers
    urn:nbn:se:liu:diva-45521 (URN)10.1111/j.0300-9475.2005.01542.x (DOI)
    Available from: 2009-10-11 Created: 2009-10-11 Last updated: 2017-12-13
    2. Structural, functional and immunologic characterization of folded subdomains in the Ro52 protein targeted in Sjögren's syndrome
    Open this publication in new window or tab >>Structural, functional and immunologic characterization of folded subdomains in the Ro52 protein targeted in Sjögren's syndrome
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    2006 (English)In: Molecular Immunology, ISSN 0161-5890, E-ISSN 1872-9142, Vol. 43, no 6, p. 588-598Article in journal (Refereed) Published
    Abstract [en]

    Ro52, one of the major autoantigens in the rheumatic disease Sjögren's syndrome (SS), belongs to the tripartite motif (TRIM) or RING-B-box-coiled-coil (RBCC) protein family, thus comprising an N-terminal RING, followed by a B-box and a coiled-coil region. Several different proteomic functions have been suggested for Ro52, including DNA binding, protein interactions and Zn 2+-binding. To analyze the presence and/or absence of these functions and, in particular, map those to different subregions, the modular composition of the Ro52 protein was experimentally characterized. Two structured parts of Ro52 were identified, corresponding to the RING-B-box and the coiled-coil regions, respectively. Secondary structure analysis by circular dichroism (CD) spectroscopy indicated that the two subregions are independently structured. The entire RING-B-box region displayed Zn2+-dependent stabilization against proteolysis in the presence of Zn2+, indicating functional Zn2+-binding sites in both the RING and the B-box. However, no stabilization with DNA was detected, irrespective of Zn2+, thus suggesting that the RING-B-box region does not bind DNA. Oligomerization of the coiled-coil was investigated by analytical ultracentrifugation and in a mammalian two-hybrid system. Both methods show weak homodimer affinity, in parity with other coiled-coil domains involved in regulatory interactions. The C-terminal B30.2 region was rapidly degraded both during cellular expression and refolding, indicating a less stable structure. Immunologic analysis of the stable protein regions with sera from patients with Sjögren's syndrome shows that immunodominant epitopes to a large extent are localized in the structurally stable parts of Ro52. The results form a basis for further Ro52 functional studies on the proteome level. © 2005 Elsevier Ltd. All rights reserved.

    Keywords
    Immunologi, Ro52, Biofysik
    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-38851 (URN)10.1016/j.molimm.2005.04.013 (DOI)45868 (Local ID)45868 (Archive number)45868 (OAI)
    Available from: 2009-10-10 Created: 2009-10-10 Last updated: 2017-12-13
    3. Structural organization and Zn2+ -dependent subdomain interactions involving autoantigenic epitopes in the Ring-B-box-coiled-coil (RBCC) region of Ro52
    Open this publication in new window or tab >>Structural organization and Zn2+ -dependent subdomain interactions involving autoantigenic epitopes in the Ring-B-box-coiled-coil (RBCC) region of Ro52
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    2005 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 280, no 39, p. 33250-33261Article in journal (Refereed) Published
    Abstract [en]

    Ro52 is one of the major autoantigens targeted in the autoimmune disease Sjögren syndrome. By sequence similarity, Ro52 belongs to the RING-B-box-coiled-coil (RBCC) protein family. Disease-related antibodies bind Ro52 in a conformation-dependent way both in the coiled-coil region and in the Zn2+-binding Ring-B-box region. Primarily associated with Sjögren syndrome, Ro52 autoantibodies directed to a specific, partially structured epitope in the coiled-coil region may also induce a congenital heart block in the fetus of pregnant Ro52-positive mothers. To improve our understanding of the pathogenic effects of autoantibody binding to the Zn 2+-binding region, a multianalytical mapping of its structural, biophysical, and antigenic properties is presented. Structure content and ligand binding of subregions, dissected by peptide synthesis and subcloning, were analyzed by fluorescence and circular dichroism spectroscopy. A novel matrix-assisted laser desorption ionization time-of-flight mass spectrometry strategy for time-resolved proteolysis experiments of large protein domains was developed to facilitate analysis and to help resolve the tertiary arrangement of the entire RBCC subregion. The linker region between the RING and B-box motifs is crucial for full folding, and Zn2+ affinity of the RING-B-box region is further protected in the entire RBCC region and appears to interact with the coiled-coil region. Murine monoclonal antibodies raised toward the RING-B-box region were primarily directed toward the linker, further supporting a highly functional role for the linker in a cellular environment. Taken together with our previous analysis of autoantigenic epitopes in the coiled-coil region, localization of autoantigenic epitopes in Ro52 appears closely related to molecular functionalities. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.

    National Category
    Engineering and Technology
    Identifiers
    urn:nbn:se:liu:diva-50421 (URN)10.1074/jbc.M503066200 (DOI)
    Available from: 2009-10-11 Created: 2009-10-11 Last updated: 2017-12-12
    4. The fellowship of the RING: The RING-B-box linker region (RBL) interacts with the RING in TRIM21/Ro52, contributes to an autoantigenic epitope in Sjögren's syndrome, and is an integral and conserved region in TRIM proteins
    Open this publication in new window or tab >>The fellowship of the RING: The RING-B-box linker region (RBL) interacts with the RING in TRIM21/Ro52, contributes to an autoantigenic epitope in Sjögren's syndrome, and is an integral and conserved region in TRIM proteins
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    2008 (English)In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 377, no 2, p. 431-449Article in journal (Refereed) Published
    Abstract [en]

    Ro52 is a major autoantigen that is targeted in the autoimmune disease Sjögren syndrome and belongs to the tripartite motif (TRIM) protein family. Disease-related antigenic epitopes are mainly found in the coiled-coil domain of Ro52, but one such epitope is located in the Zn2+-binding region, which comprises an N-terminal RING followed by a B-box, separated by a ∼40-residue linker peptide. In the present study, we extend the structural, biophysical, and immunological knowledge of this RING-B-box linker (RBL) by employing an array of methods. Our bioinformatic investigations show that the RBL sequence motif is unique to TRIM proteins and can be classified into three distinct subtypes. The RBL regions of all three subtypes are as conserved as their known flanking domains, and all are predicted to comprise an amphipathic helix. This helix formation is confirmed by circular dichroism spectroscopy and is dependent on the presence of the RING. Immunological studies show that the RBL is part of a conformation-dependent epitope, and its antigenicity is likewise dependent on a structured RING domain. Recombinant Ro52 RING-RBL exists as a monomer in vitro, and binding of two Zn2+ increases its stability. Regions stabilized by Zn2+ binding are identified by limited proteolysis and matrix-assisted laser desorption/ionization mass spectrometry. Furthermore, the residues of the RING and linker that interact with each other are identified by analysis of protection patterns, which, together with bioinformatic and biophysical data, enabled us to propose a structural model of the RING-RBL based on modeling and docking experiments. Sequence similarities and evolutionary sequence patterns suggest that the results obtained from Ro52 are extendable to the entire TRIM protein family.

    Keywords
    Ro52; TRIM21; RING; linker; zinc binding
    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-12887 (URN)10.1016/j.jmb.2008.01.005 (DOI)
    Available from: 2008-01-28 Created: 2008-01-28 Last updated: 2017-12-14Bibliographically approved
    5. Anti-Ro52 Autoantibodies from Patients with Sjögren's Syndrome Inhibit the Ro52 E3 Ligase Activity by Blocking the E3/E2 Interface
    Open this publication in new window or tab >>Anti-Ro52 Autoantibodies from Patients with Sjögren's Syndrome Inhibit the Ro52 E3 Ligase Activity by Blocking the E3/E2 Interface
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    2011 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 286, no 42, p. 36478-36491Article in journal (Refereed) Published
    Abstract [en]

    Ro52 (TRIM21) is an E3 ligase of the tripartite motif family that negatively regulates proinflammatory cytokine production by ubiquitinating transcription factors of the interferon regulatory factor family. Autoantibodies to Ro52 are present in patients with lupus and Sjögren's syndrome, but it is not known if these autoantibodies affect the function of Ro52. To address this question, the requirements for Ro52 E3 ligase activity were first analyzed in detail. Scanning a panel of E2 ubiquitin-conjugating enzymes, we found that UBE2D1–4 and UBE2E1–2 supported the E3 ligase activity of Ro52 and that the E3 ligase activity of Ro52 was dependent on its RING domain. We also found that the N-terminal extensions in the class III E2 enzymes affected their interaction with Ro52. Although the N-terminal extension in UBE2E3 made this E2 enzyme unable to function together with Ro52, the N-terminal extensions in UBE2E1 and UBE2E2 allowed for a functional interaction with Ro52. Anti-Ro52-positive patient sera and affinity-purified anti-RING domain autoantibodies inhibited the E3 activity of Ro52 in ubiquitination assays. Using NMR, limited proteolysis, ELISA, and Ro52 mutants, we mapped the interactions between Ro52, UBE2E1, and anti-Ro52 autoantibodies. We found that anti-Ro52 autoantibodies inhibited the E3 ligase activity of Ro52 by sterically blocking the E2/E3 interaction between Ro52 and UBE2E1. Our data suggest that anti-Ro52 autoantibodies binding the RING domain of Ro52 may be actively involved in the pathogenesis of rheumatic autoimmune disease by inhibiting Ro52-mediated ubiquitination.

    Place, publisher, year, edition, pages
    American Society for Biochemistry and Molecular Biology, 2011
    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-53170 (URN)10.1074/jbc.M111.241786 (DOI)000296538300033 ()
    Note

    Funding agencies|Swedish Research Council||Swedish Foundation for Strategic Research||VINNOVA||CeNano||Swedish Cancer Society||Karolinska Institutet||Linkoping University||King Gustaf Vs 80-Year Foundation||Heart-Lung Foundation||Stockholm County Council||Gustafsson Foundation||Soderberg Foundation||National Cancer Institute of Canada||Swedish Rheumatism Association||Wallenberg Foundation||

    Available from: 2010-01-18 Created: 2010-01-18 Last updated: 2017-12-12Bibliographically approved
    6. MTMDAT: Automated analysis and visualization of mass spectrometry data for tertiary and quaternary structure probing of proteins
    Open this publication in new window or tab >>MTMDAT: Automated analysis and visualization of mass spectrometry data for tertiary and quaternary structure probing of proteins
    2008 (English)In: Bioinformatics, ISSN 1367-4803, E-ISSN 1367-4811, Vol. 24, no 10, p. 1310-1312Article in journal (Refereed) Published
    Abstract [en]

    In structural biology and -genomics, nuclear magnetic resonance (NMR) spectroscopy and crystallography are the methods of choice, but sample requirements can be hard to fulfil. Valuable structural information can also be obtained by using a combination of limited proteolysis and mass spectrometry, providing not only knowledge of how to improve sample conditions for crystallization trials or NMR spectrosopy by gaining insight into subdomain identities but also probing tertiary and quaternary structure, folding and stability, ligand binding, protein interactions and the location of post-translational modifications. For high-throughput studies and larger proteins, however, this experimentally fast and easy approach produces considerable amounts of data, which until now has made the evaluation exceedingly laborious if at all manually possible. MTMDAT, equipped with a browser-like graphical user interface, accelerates this evaluation manifold by automated peak picking, assignment, data processing and visualization. © 2008 The Author(s).

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-42790 (URN)10.1093/bioinformatics/btn116 (DOI)68779 (Local ID)68779 (Archive number)68779 (OAI)
    Available from: 2009-10-10 Created: 2009-10-10 Last updated: 2017-12-13
    7. MTMDAT-HADDOCK: high-throughput, data-driven protein complex structure modeling
    Open this publication in new window or tab >>MTMDAT-HADDOCK: high-throughput, data-driven protein complex structure modeling
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Background: MTMDAT is a recently developed program facilitating the analysis of mass spectrometry data of proteins and biomolecular complexes (Hennig et al, 2008), which were structurally probed by limited proteolysis. This can provide information about stable fragments of multidomain proteins, yields tertiary and quaternary structure data, and residue-wise origins of stability changes can be determined.

    Results: A new feature allows for the direct identification of residues that are involved in complex formation bycomparing the mass spectra of bound and unbound proteins after proteolysis. If 3D structures of the unboundcomponents are available, this data can be used to restrain data-driven docking to calculate the structure ofthe complex. Here, we provide a new implementation of MTMDAT, with a pipeline to the data-driven dockingprogram HADDOCK (Dominguez et al., 2003; De Vries et al., 2007), streamlining the entire procedure. This,together with usability improvements in MTMDAT, enables direct high-throughput modelling of protein complexesfrom mass spectrometry data.

    Conclusions: MTMDAT can be downloaded from http://cms.ifm.liu.se/chemistry/molbiotech/maria¯sunnerhagens¯group/mtmdat/ (windows- or unix-based, with or without HADDOCK pipeline) together withthe manual and example files. The program is free for academic/non-commercial purposes.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-53169 (URN)
    Available from: 2010-01-18 Created: 2010-01-18 Last updated: 2010-01-18
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