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  • 1.
    Ali, Arshad
    et al.
    Hebei University.
    Mattsson, Eskil
    IVL Swedish Environmental Research Institute.
    Nissanka, Sarath Premalal
    Big-sized trees and species-functional diversity pathways mediate divergent impacts of environmental factors on individual biomass variability in Sri Lankan tropical forests2022In: Journal of Environmental Management, ISSN 0301-4797, E-ISSN 1095-8630, Vol. 315, p. 115177-115177, article id 115177Article in journal (Refereed)
    Abstract [en]

    In this study, we used the Partial Least Squares Structural Equation Models (PLS-SEMs), and other complementary analyses, on data from 189 tropical forest plots in Sri Lanka, to test the linkages amongst climate, soil, plot conditions, big-sized trees, species-functional diversity, and abiotic and biotic effects on individual biomass variability (BioVar). This study suggests that individual tree biomass variability (i.e., BioVar) should be considered for managing natural tropical forests in the context of the plant-plant interactions for species coexistence.

  • 2.
    Allison, Timothy M.
    et al.
    Biomolecular Interaction Centre, School of Physical and Chemical Sciences, University of Canterbury, Christchurch, New Zealand.
    Degiacomi, Matteo T.
    Department of Physics, Durham University, Durham, UK.
    Marklund, Erik
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
    Jovine, Luca
    Department of Biosciences and Nutrition, Karolinska Institutet, Huddinge, Sweden.
    Elofsson, Arne
    Science for Life Laboratory and Department of Biochemistry and Biophysics, Stockholm University, Solna, Sweden.
    Benesch, Justin L. P.
    Department of Chemistry, University of Oxford, Oxford, UK.
    Landreh, Michael
    Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet – Biomedicum, Stockholm, Sweden.
    Complementing machine learning‐based structure predictions with native mass spectrometry2022In: Protein Science, ISSN 0961-8368, E-ISSN 1469-896X, Vol. 31, no 6, article id e4333Article in journal (Refereed)
    Abstract [en]

    The advent of machine learning-based structure prediction algorithms such as AlphaFold2 (AF2) and RoseTTa Fold have moved the generation of accurate structural models for the entire cellular protein machinery into the reach of the scientific community. However, structure predictions of protein complexes are based on user-provided input and may require experimental validation. Mass spectrometry (MS) is a versatile, time-effective tool that provides information on post-translational modifications, ligand interactions, conformational changes, and higher-order oligomerization. Using three protein systems, we show that native MS experiments can uncover structural features of ligand interactions, homology models, and point mutations that are undetectable by AF2 alone. We conclude that machine learning can be complemented with MS to yield more accurate structural models on a small and large scale.

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  • 3.
    Amrein, Beat Anton
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Extending the Reach of Computational Approaches to Model Enzyme Catalysis2017Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Recent years have seen tremendous developments in methods for computational modeling of (bio-) molecular systems. Ever larger reactive systems are being studied with high accuracy approaches, and high-level QM/MM calculations are being routinely performed. However, applying high-accuracy methods to large biological systems is computationally expensive and becomes problematic when conformational sampling is needed. To address this challenge, classical force field based approaches such as free energy perturbation (FEP) and empirical valence bond calculations (EVB) have been employed in this work. Specifically:

    1. Force-field independent metal parameters have been developed for a range of alkaline earth and transition metal ions, which successfully reproduce experimental solvation free energies, metal-oxygen distances, and coordination numbers. These are valuable for the computational study of biological systems.

    2. Experimental studies have shown that the epoxide hydrolase from Solanum tuberosum (StEH1) is not only an enantioselective enzyme, but for smaller substrates, displays enantioconvergent behavior. For StEH1, two detailed studies, involving combined experimental and computational efforts have been performed: We first used trans-stilbene oxide to establish the basic reaction mechanism of this enzyme. Importantly, a highly conserved and earlier ignored histidine was identified to be important for catalysis. Following from this, EVB and experiment have been used to investigate the enantioconvergence of the StEH1-catalyzed hydrolysis of styrene oxide. This combined approach involved wildtype StEH1 and an engineered enzyme variant, and established a molecular understanding of enantioconvergent behavior of StEH1.

    3. A novel framework was developed for the Computer-Aided Directed Evolution of Enzymes (CADEE), in order to be able to quickly prepare, simulate, and analyze hundreds of enzyme variants. CADEE’s easy applicability is demonstrated in the form of an educational example.

    In conclusion, classical approaches are a computationally economical means to achieve extensive conformational sampling. Using the EVB approach has enabled me to obtain a molecular understanding of complex enzymatic systems. I have also increased the reach of the EVB approach, through the implementation of CADEE, which enables efficient and highly parallel in silico testing of hundreds-to-thousands of individual enzyme variants.

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  • 4.
    Amrein, Beat Anton
    et al.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Steffen-Munsberg, Fabian
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Szeler, Ireneusz
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Purg, Miha
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Kulkarni, Yashraj
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Kamerlin, Shina Caroline Lynn
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    CADEE: Computer-Aided Directed Evolution of Enzymes2017In: IUCrJ, E-ISSN 2052-2525, Vol. 4, no 1, p. 50-64Article in journal (Refereed)
    Abstract [en]

    The tremendous interest in enzymes as biocatalysts has led to extensive work in enzyme engineering, as well as associated methodology development. Here, a new framework for computer-aided directed evolution of enzymes (CADEE) is presented which allows a drastic reduction in the time necessary to prepare and analyze in silico semi-automated directed evolution of enzymes. A pedagogical example of the application of CADEE to a real biological system is also presented in order to illustrate the CADEE workflow.

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  • 5.
    Anandapadamanaban, Madhanagopal
    et al.
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, Faculty of Science & Engineering.
    Kyriakidis, Nikolaos C.
    Karolinska Univ Hosp, Sweden; UDLA, Ecuador.
    Csizmok, Veronika
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Wallenhammar, Amélie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Espinosa, Alexander C.
    Karolinska Univ Hosp, Sweden.
    Ahlner, Alexandra
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Round, Adam R.
    Grenoble Outstn, France; European XFEL GmbH, Germany.
    Trewhella, Jill
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering. Univ Sydney, Australia.
    Moche, Martin
    Karolinska Inst, Sweden.
    Wahren-Herlenius, Marie
    Karolinska Univ Hosp, Sweden.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    E3 ubiquitin-protein ligase TRIM21-mediated lysine capture by UBE2E1 reveals substrate-targeting mode of a ubiquitin-conjugating E22019In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, JOURNAL OF BIOLOGICAL CHEMISTRY, Vol. 294, no 30, p. 11404-11419Article in journal (Refereed)
    Abstract [en]

    The E3 ubiquitin-protein ligase TRIM21, of the RING-containing tripartite motif (TRIM) protein family, is a major autoantigen in autoimmune diseases and a modulator of innate immune signaling. Together with ubiquitin-conjugating enzyme E2 E1 (UBE2E1), TRIM21 acts both as an E3 ligase and as a substrate in autoubiquitination. We here report a 2.82-angstrom crystal structure of the human TRIM21 RING domain in complex with the human E2-conjugating UBE2E1 enzyme, in which a ubiquitin-targeted TRIM21 substrate lysine was captured in the UBE2E1 active site. The structure revealed that the direction of lysine entry is similar to that described for human proliferating cell nuclear antigen (PCNA), a small ubiquitin-like modifier (SUMO)-targeted substrate, and thus differs from the canonical SUMO-targeted substrate entry. In agreement, we found that critical UBE2E1 residues involved in the capture of the TRIM21 substrate lysine are conserved in ubiquitin-conjugating E2s, whereas residues critical for SUMOylation are not conserved. We noted that coordination of the acceptor lysine leads to remodeling of amino acid side-chain interactions between the UBE2E1 active site and the E2-E3 direct interface, including the so-called linchpin residue conserved in RING E3s and required for ubiquitination. The findings of our work support the notion that substrate lysine activation of an E2-E3-connecting allosteric path may trigger catalytic activity and contribute to the understanding of specific lysine targeting by ubiquitin-conjugating E2s.

  • 6.
    Anandapadmanaban, Madhanagopal
    et al.
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, Faculty of Science & Engineering.
    Pilstål, Robert
    Linköping University, Department of Physics, Chemistry and Biology, Bioinformatics. Linköping University, Faculty of Science & Engineering.
    Andrésen, Cecilia
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Trewhella, Jill
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering. University of Sydney, Australia.
    Moche, Martin
    Karolinska Institute, Sweden.
    Wallner, Björn
    Linköping University, Department of Physics, Chemistry and Biology, Bioinformatics. Linköping University, Faculty of Science & Engineering.
    Sunnerhagen, Maria
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Mutation-Induced Population Shift in the MexR Conformational Ensemble Disengages DNA Binding: A Novel Mechanism for MarR Family Derepression2016In: Structure, ISSN 0969-2126, E-ISSN 1878-4186, Vol. 24, no 8, p. 1311-1321Article in journal (Refereed)
    Abstract [en]

    MexR is a repressor of the MexAB-OprM multidrug efflux pump operon of Pseudomonas aeruginosa, where DNA-binding impairing mutations lead to multidrug resistance (MDR). Surprisingly, the crystal structure of an MDR-conferring MexR mutant R21W (2.19 angstrom) presented here is closely similar to wildtype MexR. However, our extended analysis, by molecular dynamics and small-angle X-ray scattering, reveals that the mutation stabilizes a ground state that is deficient of DNA binding and is shared by both mutant and wild-type MexR, whereas the DNA-binding state is only transiently reached by the more flexible wild-type MexR. This population shift in the conformational ensemble is effected by mutation-induced allosteric coupling of contact networks that are independent in the wild-type protein. We propose that the MexR-R21W mutant mimics derepression by small-molecule binding to MarR proteins, and that the described allosteric model based on population shifts may also apply to other MarR family members.

  • 7.
    Ancker Persson, Björn
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Structural and biophysical studies on infection phenotypes in totivirus-like viruses2023Independent thesis Advanced level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Viruses pose significant threats to human health, making it crucial to understand their structure and behavior to develop effective treatments. This study focuses on two strains of Giardia Lamblia virus (GLV) (a cat strain and human strain) a virus that infects the parasite Giardia Lamblia which in turn can infect humans or animals. Understanding viruses helps immensely with treating people who are infected as well as our ability to utilize the virus for good. The Omono River virus (OmRV) and saccharomyces cerevisae virus (ScV-L-A) is used for comparison. OmRV belongs to a group called totivirus-like viruses and ScV-L-A and GLV belong to the Totiviridae group. They have some common characteristics such as isometric virions, double-stranded RNA genomes, and 120 chemically identical subunits. What sets them apart is their hosts, viruses in the Totiviridae group only infect protozoan hosts intracellularly while OmRV only infects metazoan hosts extracellularly. GLV is interesting because it has displayed both an intra- and extracellular mode of infection. Finding out more about GLV could give us insight into viral evolution. Cryo-electron microscopy (cryoEM) is used to investigate the structure of GLV. From the cryoEM data models were created using Coot and Chimera with resolutions of 4.0 Å for the human strain and 4.2 Å for the cat strain. Found in these models were an open pore compared to the closed one of OmRV as well as lack of a C-terminal, which is used to increase stability. Signs of conserved 𝛼-helices are also found indicating their evolutionary relationship. 

    The full text will be freely available from 2026-07-04 18:21
  • 8. Andersen, Birgit
    et al.
    Lundgren, Stina
    Dobritzsch, Doreen
    Karolinska Institutet.
    Piskur, Jure
    A recruited protease is involved in catabolism of pyrimidines2008In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 379, no 2, p. 243-250Article in journal (Refereed)
    Abstract [en]

    In nature, the same biochemical reaction can be catalyzed by enzymes having fundamentally different folds, reaction mechanisms and origins. For example, the third step of the reductive catabolism of pyrimidines, the conversion of N-carbamyl-beta-alanine to beta-alanine, is catalyzed by two beta-alanine synthase (beta ASase, EC 3.5.1.6) subfamilies. We show that the "prototype" eukaryote beta ASases, such as those from Drosophila melanogaster and Arabidopsis thaliana, are relatively efficient in the conversion of N-carbamyl-beta A compared with a representative of fungal beta ASases, the yeast Saccharomyces kluyveri beta ASase, which has a high K(m) value (71 mM). S. kluyveri beta ASase is specifically inhibited by dipeptides and tripeptides, and the apparent K(i) value of glycyl-glycine is in the same range as the substrate K(m). We show that this inhibitor binds to the enzyme active center in a similar way as the substrate. The observed structural similarities and inhibition behavior, as well as the phylogenetic relationship, suggest that the ancestor of the fungal beta ASase was a protease that had modified its profession and become involved in the metabolism of nucleic acid precursors.

  • 9.
    Andersson, Charlotta S.
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Berthold, Catrine
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Högbom, Martin
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    A Dynamic C-terminal Segment in the Mycobacterium tuberculosis Mn/Fe R2lox Protein can Assume a Helical Structure with Possible Functional ConsequencesManuscript (preprint) (Other academic)
  • 10.
    Andersson, Charlotta S.
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Öhrström, Maria
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Popović-Bijelić, Ana
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Gräslund, Astrid
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Stenmark, Pål
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Högbom, Martin
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    The manganese ion of the heterodinuclear Mn/Fe cofactor in Chlamydia trachomatis ribonucleotide reductase R2c is located at metal position 1.2012In: Journal of the American Chemical Society, ISSN 0002-7863, E-ISSN 1520-5126, Vol. 134, no 1, p. 123-125Article in journal (Refereed)
    Abstract [en]

    The essential catalytic radical of Class-I ribonucleotide reductase is generated and delivered by protein R2, carrying a dinuclear metal cofactor. A new R2 subclass, R2c, prototyped by the Chlamydia trachomatis protein was recently discovered. This protein carries an oxygen-activating heterodinuclear Mn(II)/Fe(II) metal cofactor and generates a radical-equivalent Mn(IV)/Fe(III) oxidation state of the metal site, as opposed to the tyrosyl radical generated by other R2 subclasses. The metal arrangement of the heterodinuclear cofactor remains unknown. Is the metal positioning specific, and if so, where is which ion located? Here we use X-ray crystallography with anomalous scattering to show that the metal arrangement of this cofactor is specific with the manganese ion occupying metal position 1. This is the position proximal to the tyrosyl radical site in other R2 proteins and consistent with the assumption that the high-valent Mn(IV) species functions as a direct substitute for the tyrosyl radical.

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  • 11.
    Andersson, Charlotta Selina
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Structural studies of R2 and R2–like proteins with a heterodinuclear Mn/Fe cofactor and enzymes involved in Mycobacterium tuberculosis lipid metabolism2012Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Tuberculosis is a notorious disease responsible for the deaths of 1.4 million people worldwide. A third of the world's population is infected with Mycobacterium tuberculosis, the bacterium causing the disease. The increase of multi drug-resistant strains worsens the situation, and the World Health Organization has declared tuberculosis to be a global emergency. The bacterium envelopes itself with a unique set of very long-chain lipids that play an important role in virulence and drug resistance. Therefore enzymes involved in lipid metabolism are putative drug targets. 

    To allow entry into different metabolic pathways and transmembrane transport, fatty acids have to be activated. This is done primarily by fatty acyl-CoA synthetases (ACSs). We identified an ACS possibly involved in the bacterium’s virulence and solved its structure. Structural interpretation combined with previously reported data gives us insights into the details of its function. This enzyme is known to harbor lipid substrates longer than the enzyme itself, and we now propose how this peripheral membrane protein accommodates its substrates. 

    Some of the most chemically challenging oxidations are performed by dinuclear metalloproteins belonging to the ferritin-like superfamily. We show that the ferritin-like protein, R2lox, from M. tuberculosis contains a new type of heterodinuclear Mn/Fe cofactor. This protein cofactor is capable of performing potent 2-electron oxidations as demonstrated by a novel tyrosine-valine crosslink observed in the protein. 

    Recently a new subclass of ribonucleotide reductase (RNR) R2 proteins, was identified in the intracellular pathogen Chlamydia trachomatis containing the same type of Mn/Fe cofactor mentioned above. The RNR R2 proteins use their metal site to generate a stable radical, essential for the reduction of ribonucleotides to their deoxy forms, the building blocks of DNA. With this work, we were able to characterize the architecture of this metal cofactor.

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  • 12.
    Andersson, Dick
    Linköping University, Department of Physics, Measurement Technology, Biology and Chemistry. Linköping University, The Institute of Technology.
    Conformational Studies of Protein Structure and Dynamics1999Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The protein folding problem is one of the most important issues to be solved in the field of molecular biology. The subject of this thesis mainly deals with various aspects of the folding process.

    In Paper I, near-UV CD kinetic measurements on mutants, in which one tryptophan (Trp) residue had been replaced, were performed to probe the development of asymmetric environments around specific Trp residues during the refolding of human carbonic anhydrase II (HCA II). The development of the individual (Trp) CD spectra during refolding was obtained by subtracting the CD spectrum of the mutant lacking one Trp from that of HCA II at different time points. The same method was used for the particular Trp residues to obtain the kinetic CD traces monitored at a specific wavelength (270 nm). Three Trp residues were analyzed, each probing different structural regions of the native structure. The investigated Trp residues develop their native CD bands at different rates, showing that formation of native-like tertiary structure is occurring with varying rates indifferent regions of the protein.

    The same approach was applied to the extracellular domain of human tissue factor (sTF), which contains four Trp residues (Paper II). The individual Trp CD spectra showed that all four residues contributed to the CD spectrum in almost the entire wavelength region investigated, including the far-UV region. This leads to uncertain predictions of the amounts of various types of secondary structure. Accordingly, the best prediction of secondary sTF structure content was achieved using a hypothetical Trp-free CD spectrum. The kinetic refolding results suggest that the compact asymmetric environments of the individual Trp residues in sTF are formed simultaneously, leading to the conclusion that the native tertiary structure of the whole protein is formed in a cooperative manner.

    In Paper III, the role of the metal ion cofactor for the refolding of bovine carbonic anhydrase II (BCA II) was studied from the molten globule to the native state. Refolding was possible to achieve by mere addition of the metal ion to the apomolten-globule, because the apoenzyme was less stable than the holoenzyme. The cofactor-effected refolding can be summarized as follows: 1) initially, the metal ion binds to the molten globule; 2) compaction of the metal-binding site region is then induced by the metal ion binding; 3) a functioning active center is formed; 4) finally, the native tertiary structure is generated in the outer parts of the protein.

    In paper IV the aim was to determine the nature of the tetramer contact of human extracellular superoxide dismutase (hEC-SOD). We chose a strategy in which we mapped the subunit interaction interface by studying effects of twelve different mutations in the N-terminal domain fused to HCA II. The results show that the hydrophobic side of a predicted amphiphatic a-helix (formed by residues 14-32) in the N-terminal domain is essential for the subunit interaction.

  • 13.
    Andrésen, Cecilia
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Niklasson, Markus
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Cassman Eklöf, Sofie
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Wallner, Björn
    Linköping University, Department of Physics, Chemistry and Biology, Bioinformatics. Linköping University, Faculty of Science & Engineering.
    Lundström, Patrik
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Biophysical characterization of the calmodulin-like domain of Plasmodium falciparum calcium dependent protein kinase 32017In: PLOS ONE, E-ISSN 1932-6203, Vol. 12, no 7, article id e0181721Article in journal (Refereed)
    Abstract [en]

    Calcium dependent protein kinases are unique to plants and certain parasites and comprise an N-terminal segment and a kinase domain that is regulated by a C-terminal calcium binding domain. Since the proteins are not found in man they are potential drug targets. We have characterized the calcium binding lobes of the regulatory domain of calcium dependent protein kinase 3 from the malaria parasite Plasmodium falciparum. Despite being structurally similar, the two lobes differ in several other regards. While the monomeric N-terminal lobe changes its structure in response to calcium binding and shows global dynamics on the sub-millisecond time-scale both in its apo and calcium bound states, the C-terminal lobe could not be prepared calcium-free and forms dimers in solution. If our results can be generalized to the full-length protein, they suggest that the C-terminal lobe is calcium bound even at basal levels and that activation is caused by the structural reorganization associated with binding of a single calcium ion to the N-terminal lobe.

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  • 14.
    Andér, Martin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Computational Analysis of Molecular Recognition Involving the Ribosome and a Voltage Gated K+ Channel2009Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Over the last few decades, computer simulation techniques have been established as an essential tool for understanding biochemical processes. This thesis deals mainly with the application of free energy calculations to ribosomal complexes and a cardiac ion channel.

    The linear interaction energy (LIE) method is used to explore the energetic properties of the essential process of codon–anticodon recognition on the ribosome. The calculations show the structural and energetic consequences and effects of first, second, and third position mismatches in the ribosomal decoding center.

    Recognition of stop codons by ribosomal termination complexes is fundamentally different from sense codon recognition. Free energy perturbation simulations are used to study the detailed energetics of stop codon recognition by the bacterial ribosomal release factors RF1 and RF2. The calculations explain the vastly different responses to third codon position A to G substitutions by RF1 and RF2. Also, previously unknown highly specific water interactions are identified.

    The GGQ loop of ribosomal RFs is essential for its hydrolytic activity and contains a universally methylated glutamine residue. The structural effect of this methylation is investigated. The results strongly suggest that the methylation has no effect on the intrinsic conformation of the GGQ loop, and, thus, that its sole purpose is to enhance interactions in the ribosomal termination complex.

    A first microscopic, atomic level, analysis of blocker binding to the pharmaceutically interesting potassium ion channel Kv1.5 is presented. A previously unknown uniform binding mode is identified, and experimental binding data is accurately reproduced. Furthermore, problems associated with pharmacophore models based on minimized gas phase ligand conformations are highlighted.

    Generalized Born and Poisson–Boltzmann continuum models are incorporated into the LIE method to enable implicit treatment of solvent, in an effort to improve speed and convergence. The methods are evaluated and validated using a set of plasmepsin II inhibitors.

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  • 15. Antuch, W
    et al.
    Berndt, Kurt D
    Eidgenössische Technische Hochschule–Hönggerberg, Zürich, Switzerland.
    Chávez, M A
    Delfín, J
    Wüthrich, K
    The NMR solution structure of a Kunitz-type proteinase inhibitor from the sea anemone Stichodactyla helianthus.1993In: European Journal of Biochemistry, ISSN 0014-2956, E-ISSN 1432-1033, Vol. 212, p. 675-684Article in journal (Refereed)
    Abstract [en]

    The solution structure of a 55-amino-acid Kunitz-type proteinase inhibitor, ShPI, purified from the Caribbean sea anemone Stichodactyla helianthus, was determined by NMR spectroscopy. Nearly complete sequence-specific 1H-NMR assignments were obtained at pH 4.6 and 36 degrees C, and stereo-specific assignments were determined for 23 pairs of diastereotopic substituents. A data set of 666 upper distance limit constraints and 122 dihedral angle constraints collected on this basis was used as input for a structure calculation with the program DIANA. Following energy minimization with the program OPAL, the average root-mean-square diviation (RMSD) of the 20 DIANA conformers used to represent the solution structure relative to the mean structure is 61 pm for all backbone atoms N, C alpha and C', and 106 pm for all heavy atoms of residues 2-53. This high-quality solution structure of ShPI has a nearly identical molecular architecture as the bovine pancreatic trypsin inhibitor (BPTI), despite a mere 35% of sequence similarity between the two proteins. Exchange rates measured for 48 out of the 51 backbone amide protons showed that the positions of 20 slowly exchanging amide protons correlate well with hydrogen bonds involving these protons in the energy-minimized solution structure. The solution structure of ShPI is compared to the four homologous proteins for which the three-dimensional structure is also available.

  • 16. Aquila, A.
    et al.
    Barty, A.
    Bostedt, C.
    Boutet, S.
    Carini, G.
    dePonte, D.
    Drell, P.
    Doniach, S.
    Downing, K. H.
    Earnest, T.
    Elmlund, H.
    Elser, V.
    Gühr, M.
    Hajdu, Janos
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hastings, J.
    Hau-Riege, S. P.
    Huang, Z.
    Lattman, E. E.
    Maia, F. R. N. C.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Marchesini, S.
    Ourmazd, A.
    Pellegrini, C.
    Santra, R.
    Schlichting, I.
    Schroer, C.
    Spence, J. C. H.
    Vartanyants, I. A.
    Wakatsuki, S.
    Weis, W. I.
    Williams, G. J.
    The linac coherent light source single particle imaging road map2015In: Structural Dynamics, E-ISSN 2329-7778, Vol. 2, no 4, article id 041701Article in journal (Refereed)
    Abstract [en]

    Intense femtosecond x-ray pulses from free-electron laser sources allow the imag-ing of individual particles in a single shot. Early experiments at the Linac CoherentLight Source (LCLS) have led to rapid progress in the field and, so far, coherentdiffractive images have been recorded from biological specimens, aerosols, andquantum systems with a few-tens-of-nanometers resolution. In March 2014, LCLSheld a workshop to discuss the scientific and technical challenges for reaching theultimate goal of atomic resolution with single-shot coherent diffractive imaging. This paper summarizes the workshop findings and presents the roadmap towardreaching atomic resolution, 3D imaging at free-electron laser sources.

  • 17. Aquila, Andrew
    et al.
    Hunter, Mark S.
    Doak, R. Bruce
    Kirian, Richard A.
    Fromme, Petra
    White, Thomas A.
    Andreasson, Jakob
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Arnlund, David
    Bajt, Saša
    Barends, Thomas R. M.
    Barthelmess, Miriam
    Bogan, Michael J.
    Bostedt, Christoph
    Bottin, Hervé
    Bozek, John D.
    Caleman, Carl
    Coppola, Nicola
    Davidsson, Jan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Physical Chemistry.
    DePonte, Daniel P.
    Elser, Veit
    Epp, Sascha W.
    Erk, Benjamin
    Fleckenstein, Holger
    Foucar, Lutz
    Frank, Matthias
    Fromme, Raimund
    Graafsma, Heinz
    Grotjohann, Ingo
    Gumprecht, Lars
    Hajdu, Janos
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hampton, Christina Y.
    Hartmann, Andreas
    Hartmann, Robert
    Hau-Riege, Stefan
    Hauser, Günter
    Hirsemann, Helmut
    Holl, Peter
    Holton, James M.
    Hömke, André
    Johansson, Linda
    Kimmel, Nils
    Kassemeyer, Stephan
    Krasniqi, Faton
    Kühnel, Kai-Uwe
    Liang, Mengning
    Lomb, Lukas
    Malmerberg, Erik
    Marchesini, Stefano
    Martin, Andrew V.
    Maia, Filipe R.N.C.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Messerschmidt, Marc
    Nass, Karol
    Reich, Christian
    Neutze, Richard
    Rolles, Daniel
    Rudek, Benedikt
    Rudenko, Artem
    Schlichting, Ilme
    Schmidt, Carlo
    Schmidt, Kevin E.
    Schulz, Joachim
    Seibert, M. Marvin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Soltau, Heike
    Shoeman, Robert L.
    Sierra, Raymond
    Starodub, Dmitri
    Stellato, Francesco
    Stern, Stephan
    Strüder, Lothar
    Timneanu, Nicusor
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Ullrich, Joachim
    Wang, Xiaoyu
    Williams, Garth J.
    Weidenspointner, Georg
    Weierstall, Uwe
    Wunderer, Cornelia
    Barty, Anton
    Spence, John C. H.
    Chapman, Henry N.
    Time-resolved protein nanocrystallography using an X-ray free-electron laser2012In: Optics Express, E-ISSN 1094-4087, Vol. 20, no 3, p. 2706-2716Article in journal (Refereed)
    Abstract [en]

    We demonstrate the use of an X-ray free electron laser synchronized with an optical pump laser to obtain X-ray diffraction snapshots from the photoactivated states of large membrane protein complexes in the form of nanocrystals flowing in a liquid jet. Light-induced changes of Photosystem I-Ferredoxin co-crystals were observed at time delays of 5 to 10 µs after excitation. The result correlates with the microsecond kinetics of electron transfer from Photosystem I to ferredoxin. The undocking process that follows the electron transfer leads to large rearrangements in the crystals that will terminally lead to the disintegration of the crystals. We describe the experimental setup and obtain the first time-resolved femtosecond serial X-ray crystallography results from an irreversible photo-chemical reaction at the Linac Coherent Light Source. This technique opens the door to time-resolved structural studies of reaction dynamics in biological systems.

  • 18.
    Arndt, Anton
    Swedish School of Sport and Health Sciences, GIH, Department of Sport and Health Sciences, Laboratory for Biomechanics and Motor Control.
    Achilles tendon biomechanics and functional anatomy2012Conference paper (Other academic)
  • 19.
    Arndt, Anton
    et al.
    Swedish School of Sport and Health Sciences, GIH, Department of Sport and Health Sciences, Laboratory for Biomechanics and Motor Control.
    Bengtsson, Ann-Sophie
    Peolsson, Michael
    Thorstensson, Alf
    Swedish School of Sport and Health Sciences, GIH, Department of Sport and Health Sciences, Laboratory for Biomechanics and Motor Control.
    Movin, Tomas
    Non-uniform displacement within the Achilles tendon durig passive ankle joint motion.2012In: Knee Surgery, Sports Traumatology, Arthroscopy, ISSN 0942-2056, E-ISSN 1433-7347, Vol. 20, no 9, p. 1868-74Article in journal (Refereed)
    Abstract [en]

    PURPOSE:

    An initial step in the understanding of Achilles tendon dynamics is to investigate the effects of passive motion, thereby minimising muscle activation and reducing internal joint forces. Internal tendon dynamics during passive ankle joint motion have direct implications for clinical rehabilitation protocols after Achilles tendon surgery. The aim of this study was to test the hypothesis that tendon tissue displacement is different in different layers of the Achilles tendon during controlled passive ankle joint movements.

    METHODS:

    Ultrasound imaging was conducted on the right Achilles tendon of nine healthy recreationally active males. Standardised isokinetic passive dorsi-plantar-flexion movements were performed with a total range of motion of 35°. The tendon was divided into superficial, central and deep layers in the resulting B-mode ultrasound images viewed in the sagittal plane. A block-matching speckle tracking algorithm was applied post-process, with kernels for the measurement of displacement placed in each of the layers.

    RESULTS:

    The mean (SD) displacement of the Achilles tendon during passive dorsiflexion was 8.4 (1.9) mm in the superficial layer, 9.4 (1.9) mm in the central portion and 10.4 (2.1) mm in the deep layer, respectively. In all cases, the movement of the deep layer of the tendon was greater than that of the superficial one (P < 0.01).

    CONCLUSIONS:

    These results, achieved in vivo with ultrasonographic speckle tracking, indicated complex dynamic differences in different layers of the Achilles tendon, which could have implications for the understanding of healing processes of tendon pathologies and also of normal tendon function.

  • 20. Arnlund, David
    et al.
    Johansson, Linda C
    Wickstrand, Cecilia
    Barty, Anton
    Williams, Garth J
    Malmerberg, Erik
    Davidsson, Jan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Physical Chemistry.
    Milathianaki, Despina
    DePonte, Daniel P
    Shoeman, Robert L
    Wang, Dingjie
    James, Daniel
    Katona, Gergely
    Westenhoff, Sebastian
    White, Thomas A
    Aquila, Andrew
    Bari, Sadia
    Berntsen, Peter
    Bogan, Mike
    van Driel, Tim Brandt
    Doak, R Bruce
    Kjær, Kasper Skov
    Frank, Matthias
    Fromme, Raimund
    Grotjohann, Ingo
    Henning, Robert
    Hunter, Mark S
    Kirian, Richard A
    Kosheleva, Irina
    Kupitz, Christopher
    Liang, Mengning
    Martin, Andrew V
    Nielsen, Martin Meedom
    Messerschmidt, Marc
    Seibert, M Marvin
    Linac Coherent Light Source, SLAC National Accelerator Laboratory, Menlo Park, California, USA..
    Sjöhamn, Jennie
    Stellato, Francesco
    Weierstall, Uwe
    Zatsepin, Nadia A
    Spence, John C H
    Fromme, Petra
    Schlichting, Ilme
    Boutet, Sébastien
    Groenhof, Gerrit
    Chapman, Henry N
    Neutze, Richard
    Visualizing a protein quake with time-resolved X-ray scattering at a free-electron laser2014In: Nature Methods, ISSN 1548-7091, E-ISSN 1548-7105, Vol. 11, no 9, p. 923-926Article in journal (Refereed)
    Abstract [en]

    We describe a method to measure ultrafast protein structural changes using time-resolved wide-angle X-ray scattering at an X-ray free-electron laser. We demonstrated this approach using multiphoton excitation of the Blastochloris viridis photosynthetic reaction center, observing an ultrafast global conformational change that arises within picoseconds and precedes the propagation of heat through the protein. This provides direct structural evidence for a 'protein quake': the hypothesis that proteins rapidly dissipate energy through quake-like structural motions.

  • 21.
    Artursson, Elisabet
    et al.
    Swedish Defence Research Agency, CBRN, Defence and Security, Umeå.
    Andersson, Per Ola
    Swedish Defence Research Agency, CBRN, Defence and Security, Umeå.
    Akfur, Christine
    Swedish Defence Research Agency, CBRN, Defence and Security, Umeå.
    Linusson, Anna
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Börjegren, Susanne
    Swedish Defence Research Agency, CBRN, Defence and Security, Umeå.
    Ekström, Fredrik
    Swedish Defence Research Agency, CBRN, Defence and Security, Umeå.
    Catalytic-site conformational equilibrium in nerve-agent adducts of acetylcholinesterase: Possible implications for the HI-6 antidote substrate specificity2013In: Biochemical Pharmacology, ISSN 0006-2952, E-ISSN 1356-1839, Vol. 85, no 9, p. 1389-1397Article in journal (Refereed)
    Abstract [en]

    Nerve agents such as tabun, cyclosarin and Russian VX inhibit the essential enzyme acetylcholinesterase (AChE) by organophosphorylating the catalytic serine residue. Nucleophiles, such as oximes, are used as antidotes as they can reactivate and restore the function of the inhibited enzyme. The oxime HI-6 shows a notably low activity on tabun adducts but can effectively reactivate adducts of cyclosarin and Russian VX. To examine the structural basis for the pronounced substrate specificity of HI-6, we determined the binary crystal structures of Mus musculus AChE (mAChE) conjugated by cyclosarin and Russian VX and found a conformational mobility of the side chains of Phe338 and His447. The interaction between HI-6 and tabun-adducts of AChE were subsequently investigated using a combination of time resolved fluorescence spectroscopy and X-ray crystallography. Our findings show that HI-6 binds to tabun inhibited Homo sapiens AChE (hAChE) with an IC50 value of 300 μM and suggest that the reactive nucleophilic moiety of HI-6 is excluded from the phosphorus atom of tabun. We propose that a conformational mobility of the side-chains of Phe338 and His447 is a common feature in nerve-agent adducts of AChE. We also suggest that the conformational mobility allow HI-6 to reactivate conjugates of cyclosarin and Russian VX while a reduced mobility in tabun conjugated AChE results in steric hindrance that prevents efficient reactivation.

  • 22.
    Aurelius, Oskar
    et al.
    Lund University.
    Johansson, Renzo
    Lund University.
    Bågenholm, Viktoria
    Lund University.
    Lundin, Daniel
    Stockholm University.
    Tholander, Fredrik
    Karolinska Institutet.
    Balhuizen, Alexander
    Lund University.
    Beck, Tobias
    University of Göttingen, Germany.
    Sahlin, Margareta
    Stockholm University.
    Sjöberg, Britt-Marie
    Stockholm University.
    Mulliez, Etienne
    Institut de Recherches en Technologies et Sciences pour le Vivant (iRTSV), France.
    Logan, Derek T.
    Lund University.
    The Crystal Structure of Thermotoga maritima Class III Ribonucleotide Reductase Lacks a Radical Cysteine Pre-Positioned in the Active Site2015In: PLOS ONE, E-ISSN 1932-6203, Vol. 10, no 7, article id e0128199Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductases (RNRs) catalyze the reduction of ribonucleotides to deoxyribonucleotides, the building blocks for DNA synthesis, and are found in all but a few organisms. RNRs use radical chemistry to catalyze the reduction reaction. Despite RNR having evolved several mechanisms for generation of different kinds of essential radicals across a large evolutionary time frame, this initial radical is normally always channelled to a strictly conserved cysteine residue directly adjacent to the substrate for initiation of substrate reduction, and this cysteine has been found in the structures of all RNRs solved to date. We present the crystal structure of an anaerobic RNR from the extreme thermophile Thermotoga maritima (tmNrdD), alone and in several complexes, including with the allosteric effector dATP and its cognate substrate CTP. In the crystal structure of the enzyme as purified, tmNrdD lacks a cysteine for radical transfer to the substrate pre-positioned in the active site. Nevertheless activity assays using anaerobic cell extracts from T. maritima demonstrate that the class III RNR is enzymatically active. Other genetic and microbiological evidence is summarized indicating that the enzyme is important for T. maritima. Mutation of either of two cysteine residues in a disordered loop far from the active site results in inactive enzyme. We discuss the possible mechanisms for radical initiation of substrate reduction given the collected evidence from the crystal structure, our activity assays and other published work. Taken together, the results suggest either that initiation of substrate reduction may involve unprecedented conformational changes in the enzyme to bring one of these cysteine residues to the expected position, or that alternative routes for initiation of the RNR reduction reaction may exist. Finally, we present a phylogenetic analysis showing that the structure of tmNrdD is representative of a new RNR subclass IIIh, present in all Thermotoga species plus a wider group of bacteria from the distantly related phyla Firmicutes, Bacteroidetes and Proteobacteria.

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  • 23.
    Ayranci, Diyar
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Design, expression and purification of virus-like particles derived from metagenomic studies: Virus-like Particles (VLP) of novel Partitiviridae species, Hubei.PLV 11, and novel Soutern pygmy squid flavilike virus were designed, expressed using the bac-to-bac expression system and then pruified using various methods2021Independent thesis Advanced level (degree of Master (Two Years)), 30 credits / 45 HE creditsStudent thesis
    Abstract [en]

    Viruses are entities which are made of a few genes and are reliant on obligate parasitism to propagate. Due to the obligate connection to their hosts, virus evolution is constrained to the type of host. Viruses however do transmit to evolutionary distinct hosts; in these cases, the phylogenetic relationship of the hosts usually are close. In some instances, RNA-viruses have made host jumps between evolutionary distant hosts, such as the host jump from invertebrates to vertebrates, and fungi to arthropod. Partitiviruses are double stranded RNA viruses which mainly infect fungi and plants. The defining characteristic of these double stranded RNA viruses are the double layered capsids which are formed by a single open reading frame (ORF). The capsid proteins form icosahedral virus particles which are in the magnitude of 30-40 nm. Metagenomic studies have discovered partitiviruses originating from an insect in the Odanata family, a finding which contradicts the fungal host specificity of partitiviruses. The finding of the Hubei.PLV 11 thus implies the existence of a partitiviruses containing structural elements in their capsids which could be involved in the infection of arthropods. Thus, this virus could be used as a model for a structural comparison with its fungi infecting relatives with hopes to identify common viral structural factors necessary for the infection of arthropods. For this purpose, the Hubei.PLV ORF was cloned and then transfected into insect Spodoptera frugiperda (Sf-9) cells using a baculovirus expression system, “bac-to-bac” expression system. The FLAG-tagged capsid proteins were expressed by the Sf-9 cells to be approximately 60 kDa. After ultra-centrifugation in a sucrose gradient, some spontaneous assembly into the expected ~40 nm icosahedral virus-like particles were observed using low resolution scanning electron microscopy. The observed particles were also confirmed by a dynamic light scattering experiment (DLS) and a higher resolution cryo-EM microscope. Thus, the bac-to-bac expression system can be used to produce VLPs from this genus of viruses, and this metagenomically derived virus genome. However, for future success in defining a high-resolution model of this virus, it is recommended that the Sf-9 culture volume is sufficiently high for enough particle production which is necessary for a high-resolution map. The other virus, the Southern pygmy squid Flavilike virus (SpSFV) has been suggested to be the oldest relative of the land based flaviviruses. The SpSFV was found to be the most divergent of the flaviviruses, and to infect invertebrates. Solving for the structure of the SpSFV and comparing it to vertebrate infecting flaviviruses could therefore lead to the identification of factors necessary for the adaptation to vertebrates and thus the humoral immunity by flaviviruses. The soluble E-protein was expressed using the bac-to-bac expression system. The protein was indicated to be multiglycosylated and approximately 50 kDa which is in line with other strains in the genus. Affinity chromatography did not elute this protein, likely due to the His-tag not being spatially available. Cation exchange could elute some protein, but not much from the small ~30 mL culture. To conclude, VLP assembly was confirmed by the Hubei.PLV, thus, solving for the structure is a distinct possibility when a larger Sf-9 culture is used to produce the VLPs. For the SpSFV soluble E-protein, the protein is secreted into the supernatant of the Sf-9 cultures, making purification a possibility. For this, a large Sf-9 culture can be used to produce this protein and then purify it with a cat-ion exchange chromatography.

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  • 24.
    Bacic, Luka
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology. Uppsala University.
    Molecular mechanisms underlying the activation of ALC1 nucleosome remodeling2022Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Packaging DNA into chromatin represses essential DNA-based processes, such as transcription, DNA replication, and repair. To change the accessibility of DNA, cells have evolved a set of enzymes referred to as chromatin remodelers that act on the basic repeat unit of chromatin,  the nucleosome. Chromatin remodelers are critical for normal cell physiology and development. Dysfunction or aberrant regulation of chromatin remodelers can lead to multisystem developmental disorders and cancers. DNA damage represents a major threat to eukaryotic cells. When DNA damage persists, the cell can enter programmed cell death. To avoid such a dramatic outcome, cells must rapidly recognize the DNA damage and trigger DNA repair pathways. An early event following DNA damage is the relaxation of chromatin. Chromatin relaxation depends on ATP consumption and ADP-ribosylation, where the site of DNA damage is marked with ADP-ribose units. ADP-ribose, in turn, can be recognized by the macro domain of the remodeler ALC1 (Amplified in Liver Cancer 1). ALC1 has therefore been implicated in the DNA damage response. In the absence of DNA damage, the macro domain of ALC1 is placed against its ATPase motor to inhibit its activity. However, it is unclear how ALC1, in its active state, engages the nucleosome. Moreover, the mechanism by which ALC1 is fully activated upon recruitment is poorly understood, and the impact of ALC1-catalyzed nucleosome sliding in the vicinity of a DNA damage site is unknown. This thesis investigates how ALC1 engages its substrate, the nucleosome, and how histone modifications can regulate ALC1 activity. Structural and biophysical approaches revealed an ALC1 regulatory segment that binds to the acidic patch, a prominent feature on the nucleosome surface. Further analysis showed that the interaction between ALC1 and the acidic patch is required to fully activate ALC1. Moreover, in vitro ADP-ribosylation of nucleosomes enabled us to form a stable complex of nucleosome-bound ALC1 amenable to structural determination by cryogenic electron microscopy. Our structural models visualize nucleosomal epitopes that play an important role in stimulating productive remodeling by ALC1, as confirmed by various biochemical approaches. Taken together, our data suggested a possible mechanism by which ALC1 could render DNA breaks more accessible to downstream repair factors. Since recent studies defined ALC1 as an attractive anti-cancer target, this thesis provides insights into the molecular mechanisms that regulate ALC1 activity as a potential starting point for structure-based drug development.

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  • 25.
    Bacic, Luka
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Gaullier, Guillaume
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Mohapatra, Jugal
    Mao, Guanzhong
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Brackmann, Klaus
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Panfilov, Mikhail
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Liszczak, Glen
    Sabantsev, Anton
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Deindl, Sebastian
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Asymmetric nucleosome PARylation at DNA breaks mediates directional nucleosome sliding by ALC12024In: Nature Communications, E-ISSN 2041-1723, Vol. 15, no 1, article id 1000Article in journal (Refereed)
    Abstract [en]

    The chromatin remodeler ALC1 is activated by DNA damage-induced poly(ADP-ribose) deposited by PARP1/PARP2 and their co-factor HPF1. ALC1 has emerged as a cancer drug target, but how it is recruited to ADP-ribosylated nucleosomes to affect their positioning near DNA breaks is unknown. Here we find that PARP1/HPF1 preferentially initiates ADP-ribosylation on the histone H2B tail closest to the DNA break. To dissect the consequences of such asymmetry, we generate nucleosomes with a defined ADP-ribosylated H2B tail on one side only. The cryo-electron microscopy structure of ALC1 bound to such an asymmetric nucleosome indicates preferential engagement on one side. Using single-molecule FRET, we demonstrate that this asymmetric recruitment gives rise to directed sliding away from the DNA linker closest to the ADP-ribosylation site. Our data suggest a mechanism by which ALC1 slides nucleosomes away from a DNA break to render it more accessible to repair factors.

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  • 26. Banijamali, Elnaz
    et al.
    Baronti, Lorenzo
    Becker, Walter
    Sajkowska-Kozielewicz, Joanna J.
    Huang, Ting
    Palka, Christina
    Kosek, David
    Sweetapple, Lara
    Müller, Juliane
    Stone, Michael D.
    Andersson, Emma R.
    Petzold, Katja
    Department of Medical Biochemistry and Biophysics, Karolinska Institute, 17177 Stockholm, Sweden; Stellenbosch Institute for Advanced Study (STIAS), Wallenberg Research Centre at Stellenbosch University, Stellenbosch 7600, South Africa.
    RNA:RNA interaction in ternary complexes resolved by chemical probing2022In: RNA: A publication of the RNA Society, ISSN 1355-8382, E-ISSN 1469-9001, Vol. 29, no 3, p. 317-329Article in journal (Refereed)
    Abstract [en]

    RNA regulation can be performed by a second targeting RNA molecule, such as in the microRNA regulation mechanism. Selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE) probes the structure of RNA molecules and can resolve RNA:protein interactions, but RNA:RNA interactions have not yet been addressed with this technique. Here, we apply SHAPE to investigate RNA-mediated binding processes in RNA:RNA and RNA:RNA-RBP complexes. We use RNA:RNA binding by SHAPE (RABS) to investigate microRNA-34a (miR-34a) binding its mRNA target, the silent information regulator 1 (mSIRT1), both with and without the Argonaute protein, constituting the RNA-induced silencing complex (RISC). We show that the seed of the mRNA target must be bound to the microRNA loaded into RISC to enable further binding of the compensatory region by RISC, while the naked miR-34a is able to bind the compensatory region without seed interaction. The method presented here provides complementary structural evidence for the commonly performed luciferase-assay-based evaluation of microRNA binding-site efficiency and specificity on the mRNA target site and could therefore be used in conjunction with it. The method can be applied to any nucleic acid-mediated RNA- or RBP-binding process, such as splicing, antisense RNA binding, or regulation by RISC, providing important insight into the targeted RNA structure.

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  • 27. Barbercheck Epler, Chelsea R.
    et al.
    Bullitt, Esther
    Andersson, Magnus
    Umeå University, Faculty of Science and Technology, Department of Physics.
    Bacterial adhesion pili2018In: Membrane protein complexes: structure and function / [ed] J. Robin Harris, Egbert Boekema, Springer Publishing Company, 2018, , p. 18p. 1-18Chapter in book (Refereed)
    Abstract [en]

    Escherichia coli bacterial cells produce multiple types of adhesion pili that mediate cell-cell and cell-host attachments. These pili (also called 'fimbriae') are large biopolymers that are comprised of subunits assembled via a sophisticated micro-machinery into helix-like structures that are anchored in the bacterial outer membrane. They are commonly essential for initiation of disease and thus provide a potential target for antibacterial prevention and treatment. To develop new therapeutics for disease prevention and treatment we need to understand the molecular mechanisms and the direct role of adhesion pili during pathogenesis. These helix-like pilus structures possess fascinating and unique biomechanical properties that have been thoroughly investigated using high-resolution imaging techniques, force spectroscopy and fluid flow chambers. In this chapter, we first discuss the structure of pili and the micro-machinery responsible for the assembly process. Thereafter, we present methods for measurement of the biomechanics of adhesion pili, including optical tweezers. Data demonstrate unique biomechanical properties of pili that allow bacteria to sustain binding during in vivo fluid shear forces. We thereafter summarize the current biomechanical findings related to adhesion pili and show that pili biomechanical properties are niche-specific. That is, the data suggest that there is an organ-specific adaptation of pili that facilitates infection of the bacteria's target tissue. Thus, pilus biophysical properties are an important part of Escherichia coli pathogenesis, allowing bacteria to overcome hydrodynamic challenges in diverse environments.

  • 28. Barbieri, Shayla Fernanda
    et al.
    Ruthes, Andrea C.
    KTH, School of Biotechnology (BIO), Glycoscience.
    de Oliveira Petkowicz, Carmen Lucia
    Bueno de Godoy, Rossana Catie
    Sassaki, Guilherme Lanzi
    Santana Filho, Arquimedes Paixao
    Meira Silveira, Joana Lea
    Extraction, purification and structural characterization of a galactoglucomannan from the gabiroba fruit (Campomanesia xanthocarpa Berg), Myrtaceae family2017In: Carbohydrate Polymers, ISSN 0144-8617, E-ISSN 1879-1344, Vol. 174, p. 887-895Article in journal (Refereed)
    Abstract [en]

    In this study, we isolated and structurally characterized, for the first time, a galactoglucomannan (GGM) from the pulp of gabiroba, a Myrtaceae family species. The HPSEC-MALLS-RI analysis showed a homogeneous polysaccharide with molar mass of 25,340 g mol(-1). The monosaccharide composition showed that the GGM consisted of Man:Glc:Gal in a molar ratio of 1:1:0.6. Methylation and 1D and 2D NMR analyses suggested that the main chain of the GGM consisted of beta-D-Glcp and beta-D-Manp units (1 -> 4)-linked. The alpha-D-Galp substitutions occur mainly at O-6 position of beta-D-Manp units. The glycosidic linkages of the GGM were evident by the presence of the characteristic signals of 4-O-substituted residues at delta 78.6/3.69 for both beta-D-Glcp and beta-D-Manp. Furthermore, the 0-6 substitutions for both beta-D-Glcp and beta-D-Manp units were confirmed by signals at delta 67.1/4.00 and 3.93. The interglycosidic correlations, obtained through the analysis of the HMBC spectrum, further confirm the structure. (C) 2017 Elsevier Ltd. All rights reserved.

  • 29. Barends, Thomas R. M.
    et al.
    Hartmann, Elisabeth
    Griese, Julia J.
    Beitlich, Thorsten
    Kirienko, Natalia V.
    Ryjenkov, Dmitri A.
    Reinstein, Jochen
    Shoeman, Robert L.
    Gomelsky, Mark
    Schlichting, Ilme
    Structure and mechanism of a bacterial light-regulated cyclic nucleotide phosphodiesterase2009In: Nature, ISSN 0028-0836, E-ISSN 1476-4687, Vol. 459, p. 1015-1018Article in journal (Refereed)
    Abstract [en]

    The ability to respond to light is crucial for most organisms. BLUF is a recently identified photoreceptor protein domain that senses blue light using a FAD chromophore. BLUF domains are present in various proteins from the Bacteria, Euglenozoa and Fungi. Although structures of single-domain BLUF proteins have been determined, none are available for a BLUF protein containing a functional output domain; the mechanism of light activation in this new class of photoreceptors has thus remained poorly understood. Here we report the biochemical, structural and mechanistic characterization of a full-length, active photoreceptor, BlrP1 (also known as KPN_01598), from Klebsiella pneumoniae. BlrP1 consists of a BLUF sensor domain and a phosphodiesterase EAL output domain which hydrolyses cyclic dimeric GMP (c-di-GMP). This ubiquitous second messenger controls motility, biofilm formation, virulence and antibiotic resistance in the Bacteria. Crystal structures of BlrP1 complexed with its substrate and metal ions involved in catalysis or in enzyme inhibition provide a detailed understanding of the mechanism of the EAL-domain c-di-GMP phosphodiesterases. These structures also sketch out a path of light activation of the phosphodiesterase output activity. Photon absorption by the BLUF domain of one subunit of the antiparallel BlrP1 homodimer activates the EAL domain of the second subunit through allosteric communication transmitted through conserved domain-domain interfaces.

  • 30.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Automated liquid-handling systems for submicroliter crystallization2007In: Protein Crystallization Strategies for Structural Genomics / [ed] Naomi E. Chayen, La Jolla, California: International University Line , 2007, p. 57-73Chapter in book (Other academic)
  • 31.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Succeeding with seeding: some practical advice2007In: Evolving Methods for Macromolecular Crystallography / [ed] Read, Randy J.; Sussman, Joel L., 2007, p. 1-10Conference paper (Refereed)
    Abstract [en]

    Seeding is a powerful and versatile method for optimizing crystal growth conditions. This article discusses, from a practical point of view, what seeding is, the selection and transfer of seeds, and into what conditions they should be transferred. The most common causes of failures in seeding experiments are also analyzed.

  • 32.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology.
    The RAPID crystallization strategy for structure-based inhibitor design2009In: From Molecules to Medicines: Structure of Biological Macromolecules and Its Relevance in Combating New Diseases and Bioterrorism / [ed] J. Sussman and P. Spadon, Dordrecht, The Netherlands: Springer , 2009, p. 11-19Chapter in book (Other academic)
  • 33. Bergh, Cathrine
    et al.
    Rovšnik, Urška
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Howard, Rebecca J.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). KTH Royal Institute of Technology, Sweden.
    Lindahl, Erik
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). KTH Royal Institute of Technology, Sweden.
    Discovery of lipid binding sites in a ligand-gated ion channel by integrating simulations and cryo-EM2023In: eLIFE, E-ISSN 2050-084X, Vol. 12, article id RP86016Article in journal (Refereed)
    Abstract [en]

    Ligand-gated ion channels transduce electrochemical signals in neurons and other excitable cells. Aside from canonical ligands, phospholipids are thought to bind specifically to the transmembrane domain of several ion channels. However, structural details of such lipid contacts remain elusive, partly due to limited resolution of these regions in experimental structures. Here, we discovered multiple lipid interactions in the channel GLIC by integrating cryo-electron microscopy and large-scale molecular simulations. We identified 25 bound lipids in the GLIC closed state, a conformation where none, to our knowledge, were previously known. Three lipids were associated with each subunit in the inner leaflet, including a buried interaction disrupted in mutant simulations. In the outer leaflet, two intrasubunit sites were evident in both closed and open states, while a putative intersubunit site was preferred in open-state simulations. This work offers molecular details of GLIC-lipid contacts particularly in the ill-characterized closed state, testable hypotheses for state-dependent binding, and a multidisciplinary strategy for modeling protein-lipid interactions.

  • 34.
    Bergkvist, Liza
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering. Van Andel Res Inst, MI USA.
    Du, Zhen
    Univ Cambridge, England; Univ Cambridge, England.
    Elovsson, Greta
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Appelqvist, Hanna
    Linköping University, Department of Biomedical and Clinical Sciences, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Itzhaki, Laura S.
    Univ Cambridge, England.
    Kumita, Janet R.
    Univ Cambridge, England.
    Kågedal, Katarina
    Linköping University, Department of Biomedical and Clinical Sciences, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Brorsson, Ann-Christin
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Mapping pathogenic processes contributing to neurodegeneration in Drosophila models of Alzheimers disease2020In: FEBS Open Bio, E-ISSN 2211-5463, Vol. 10, no 3, p. 338-350Article in journal (Refereed)
    Abstract [en]

    Alzheimers disease (AD) is the most common form of dementia, affecting millions of people and currently lacking available disease-modifying treatments. Appropriate disease models are necessary to investigate disease mechanisms and potential treatments. Drosophila melanogaster models of AD include the A beta fly model and the A beta PP-BACE1 fly model. In the A beta fly model, the A beta peptide is fused to a secretion sequence and directly overexpressed. In the A beta PP-BACE1 model, human A beta PP and human BACE1 are expressed in the fly, resulting in in vivo production of A beta peptides and other A beta PP cleavage products. Although these two models have been used for almost two decades, the underlying mechanisms resulting in neurodegeneration are not yet clearly understood. In this study, we have characterized toxic mechanisms in these two AD fly models. We detected neuronal cell death and increased protein carbonylation (indicative of oxidative stress) in both AD fly models. In the A beta fly model, this correlates with high A beta(1-42) levels and down-regulation of the levels of mRNA encoding lysosomal-associated membrane protein 1, lamp1 (a lysosomal marker), while in the A beta PP-BACE1 fly model, neuronal cell death correlates with low A beta(1-42) levels, up-regulation of lamp1 mRNA levels and increased levels of C-terminal fragments. In addition, a significant amount of A beta PP/A beta antibody (4G8)-positive species, located close to the endosomal marker rab5, was detected in the A beta PP-BACE1 model. Taken together, this study highlights the similarities and differences in the toxic mechanisms which result in neuronal death in two different AD fly models. Such information is important to consider when utilizing these models to study AD pathogenesis or screening for potential treatments.

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  • 35.
    Berndt, Kurt D
    et al.
    Eidgenössische Technische Hochschule-Hönggerberg, Zürich, Switzerland.
    Beunink, J
    Schröder, W
    Wüthrich, K
    Designed replacement of an internal hydration water molecule in BPTI: structural and functional implications of a glycine-to-serine mutation.1993In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 32, p. 4564-4570Article in journal (Refereed)
    Abstract [en]

    The three-dimensional structure of the basic pancreatic trypsin inhibitor (BPTI) contains four internal water molecules, which form a total of nine intermolecular hydrogen bonds with the BPTI polypeptide chain. To investigate the effect of such internal hydration on protein structure and stability, we displaced one of the internal water molecules in a recombinant BPTI analogue, BPTI(G36S), in which Gly 36 is replaced by serine. The replacement of a water molecule by the seryl side chain was established by the absence of the protein-water nuclear Overhauser effects (NOE) that had been attributed to the water molecule near Gly 36 in wild-type BPTI and by the presence of new, intramolecular NOEs to the hydroxyl proton of Ser 36. BPTI(G36S) has slightly reduced thermal stability compared to BPTI, corresponding to a destabilization by delta (delta G) approximately 0.7 kcal/M in 6 M guanidinium hydrochloride solution. Additionally, the stabilities of the complexes formed between BPTI(G36S) and trypsin, plasmin, or kallikrein are significantly reduced when compared to the corresponding complexes with wild-type BPTI.

  • 36.
    Berndt, Kurt D
    et al.
    Eidgenossische TH-Honggerberg, Zürich, Switzerland.
    Güntert, P
    Wüthrich, K
    Nuclear-Magnetic-Resonance Solution Structure of Dendrotoxin-K from the Venom of Dendroaspis-Polylepis-Polylepis1993In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 234, no 3, p. 735-750Article in journal (Refereed)
  • 37.
    Berndt, Kurt D
    et al.
    Eidgenösische Technische Hochschule-Hönggerberg, Zürich, Switzerland.
    Güntert, Peter
    Orbons, Leonard P.M.
    Wüthrich, Kurt
    Determination of a high-quality nuclear magnetic resonance solution structure of the bovine pancreatic trypsin inhibitor and comparison with three crystal structures1992In: Journal of Molecular Biology, Vol. 227, p. 757-775Article in journal (Refereed)
    Abstract [en]

    A high-quality three-dimensional structure of the bovine pancreatic trypsin inhibitor (BPTI) in aqueous solution was determined by 1H nuclear magnetic resonance (n.m.r.) spectroscopy and compared to the three available high-resolution X-ray crystal structures. A newly collected input of 642 distance constraints derived from nuclear Overhauser effects and 115 dihedral angle constraints was used for the structure calculations with the program DIANA, followed by restrained energy minimization with the program AMBER. The BPTI solution structure is represented by a group of 20 conformers with an average root-mean-square deviation (RMSD) relative to the mean solution structure of 0.43 A for backbone atoms and 0.92 A for all heavy atoms of residues 2 to 56. The pairwise RMSD values of the three crystal structures relative to the mean solution structure are 0.76 to 0.85 A for the backbone atoms and 1.24 to 1.33 A for all heavy atoms of residues 2 to 56. Small local differences in backbone atom positions between the solution structure and the X-ray structures near residues 9, 25 to 27, 46 to 48 and 52 to 58, and conformational differences for individual amino acid side-chains were analyzed for possible correlations with intermolecular protein-protein contacts in the crystal lattices, using the pairwise RMSD values among the three crystal structures as a reference.

  • 38.
    Berndt, Kurt D
    et al.
    Eidgenössische TH-Hönggerberg, Zürich, Switzerland.
    Güntert, Peter
    Wüthrich, Kurt
    Conformational sampling by NMR solution structures calculated with the program DIANA evaluated by comparison with long-time molecular dynamics calculations in explicit water1996In: Proteins: Structure, Function, and Bioinformatics, ISSN 0887-3585, E-ISSN 1097-0134, Vol. 24, p. 304-313Article in journal (Refereed)
    Abstract [en]

    The NMR solution structure of bovine pancreatic trypsin inhibitor (BPTI) obtained by distance geometry calculations with the program DIANA is compared with groups of conformers generated by molecular dynamics (MD) simulations in explicit water at ambient temperature and pressure. The MD simulations started from a single conformer and were free or restrained either by the experimental NOE distance restraints or by time-averaged restraints; the groups of conformers were collected either in 10 ps intervals during 200 ps periods of simulation, or in 50 ps intervals during a 1 ns period of simulation. Overall, these comparisons show that the standard protein structure determination protocol with the program DIANA provides a picture of the protein structure that is in agreement with MD simulations using "realistic" potential functions over a nanosecond timescale. For well-constrained molecular regions there is a trend in the free MD simulation of duration 1 ns that the sampling of the conformation space is slightly increased relative to the DIANA calculations. In contrast, for surface-exposed side-chains that are less extensively constrained by the NMR data, the DIANA conformers tend to sample larger regions of conformational space than conformers selected from any of the MD trajectories. Additional insights into the behavior of surface side-chains come from comparison of the MD runs of 200 ps or 1 ns duration. In this time range the sampling of conformation space by the protein surface depends strongly on the length of the simulation, which indicates that significant side-chain transitions occur on the nanosecond timescale and that much longer simulations will be needed to obtain statistically significant data on side-chain dynamics.

  • 39.
    Berntsson, Elina
    et al.
    Department of Biochemistry and Biophysics, Arrhenius Laboratories, Stockholm University, Stockholm, Sweden; Department of Chemistry and Biotechnology, Tallinn University of Technology, Tallinn, Estonia.
    Vosough, Faraz
    Department of Biochemistry and Biophysics, Arrhenius Laboratories, Stockholm University, Stockholm, Sweden.
    Svantesson, Teodor
    Department of Biochemistry and Biophysics, Arrhenius Laboratories, Stockholm University, Stockholm, Sweden.
    Pansieri, Jonathan
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Iashchishyn, Igor
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Ostojic, Lucija
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Dong, Xiaolin
    Department of Biochemistry and Biophysics, Arrhenius Laboratories, Stockholm University, Stockholm, Sweden.
    Paul, Suman
    Department of Biochemistry and Biophysics, Arrhenius Laboratories, Stockholm University, Stockholm, Sweden.
    Jarvet, Jüri
    Department of Biochemistry and Biophysics, Arrhenius Laboratories, Stockholm University, Stockholm, Sweden; National Institute of Chemical Physics and Biophysics, Tallinn, Estonia.
    Roos, Per M.
    Institute of Environmental Medicine, Karolinska Institutet, Nobels Väg 13, Stockholm, Sweden; Department of Clinical Physiology, Capio St. Göran Hospital, St. Göransplan 1, Stockholm, Sweden.
    Barth, Andreas
    Department of Biochemistry and Biophysics, Arrhenius Laboratories, Stockholm University, Stockholm, Sweden.
    Morozova-Roche, Ludmilla
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Gräslund, Astrid
    Department of Biochemistry and Biophysics, Arrhenius Laboratories, Stockholm University, Stockholm, Sweden.
    Wärmländer, Sebastian K T S
    Chemistry Section, Arrhenius Laboratories, Stockholm University, Stockholm, Sweden.
    Residue-specific binding of Ni(II) ions influences the structure and aggregation of amyloid beta (Aβ) peptides2023In: Scientific Reports, E-ISSN 2045-2322, Vol. 13, no 1, article id 3341Article in journal (Refereed)
    Abstract [en]

    Alzheimer's disease (AD) is the most common cause of dementia worldwide. AD brains display deposits of insoluble amyloid plaques consisting mainly of aggregated amyloid-β (Aβ) peptides, and Aβ oligomers are likely a toxic species in AD pathology. AD patients display altered metal homeostasis, and AD plaques show elevated concentrations of metals such as Cu, Fe, and Zn. Yet, the metal chemistry in AD pathology remains unclear. Ni(II) ions are known to interact with Aβ peptides, but the nature and effects of such interactions are unknown. Here, we use numerous biophysical methods-mainly spectroscopy and imaging techniques-to characterize Aβ/Ni(II) interactions in vitro, for different Aβ variants: Aβ(1-40), Aβ(1-40)(H6A, H13A, H14A), Aβ(4-40), and Aβ(1-42). We show for the first time that Ni(II) ions display specific binding to the N-terminal segment of full-length Aβ monomers. Equimolar amounts of Ni(II) ions retard Aβ aggregation and direct it towards non-structured aggregates. The His6, His13, and His14 residues are implicated as binding ligands, and the Ni(II)·Aβ binding affinity is in the low µM range. The redox-active Ni(II) ions induce formation of dityrosine cross-links via redox chemistry, thereby creating covalent Aβ dimers. In aqueous buffer Ni(II) ions promote formation of beta sheet structure in Aβ monomers, while in a membrane-mimicking environment (SDS micelles) coil-coil helix interactions appear to be induced. For SDS-stabilized Aβ oligomers, Ni(II) ions direct the oligomers towards larger sizes and more diverse (heterogeneous) populations. All of these structural rearrangements may be relevant for the Aβ aggregation processes that are involved in AD brain pathology.

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  • 40.
    Bhowmick, Asmit
    et al.
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Simon, Philipp S.
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Bogacz, Isabel
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Hussein, Rana
    Department of Biology, Humboldt-Universität zu Berlin, Berlin, Germany.
    Zhang, Miao
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Makita, Hiroki
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Ibrahim, Mohamed
    Department of Biology, Humboldt-Universität zu Berlin, Berlin, Germany.
    Chatterjee, Ruchira
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Doyle, Margaret D.
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Cheah, Mun Hon
    Molecular Biomimetics, Department of Chemistry-Ångström, Uppsala University, Uppsala, Sweden.
    Chernev, Petko
    Molecular Biomimetics, Department of Chemistry-Ångström, Uppsala University, Uppsala, Sweden.
    Fuller, Franklin D.
    Linac Coherent Light Source, SLAC National Accelerator Laboratory, CA, Menlo Park, United States.
    Fransson, Thomas
    Department of Physics, AlbaNova University Center, Stockholm University, Stockholm, Sweden.
    Alonso-Mori, Roberto
    Linac Coherent Light Source, SLAC National Accelerator Laboratory, CA, Menlo Park, United States.
    Brewster, Aaron S.
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Sauter, Nicolas K.
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Bergmann, Uwe
    Department of Physics, University of Wisconsin-Madison, WI, Madison, United States.
    Dobbek, Holger
    Department of Biology, Humboldt-Universität zu Berlin, Berlin, Germany.
    Zouni, Athina
    Department of Biology, Humboldt-Universität zu Berlin, Berlin, Germany.
    Messinger, Johannes
    Umeå University, Faculty of Science and Technology, Department of Chemistry. Molecular Biomimetics, Department of Chemistry-Ångström, Uppsala University, Uppsala, Sweden.
    Kern, Jan
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Yachandra, Vittal K.
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Yano, Junko
    Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, CA, Berkeley, United States.
    Going around the Kok cycle of the water oxidation reaction with femtosecond X-ray crystallography2023In: IUCrJ, E-ISSN 2052-2525, Vol. 10, no 6, p. 642-655Article, review/survey (Refereed)
    Abstract [en]

    The water oxidation reaction in photosystem II (PS II) produces most of the molecular oxygen in the atmosphere, which sustains life on Earth, and in this process releases four electrons and four protons that drive the downstream process of CO2 fixation in the photosynthetic apparatus. The catalytic center of PS II is an oxygen-bridged Mn4Ca complex (Mn4CaO5) which is progressively oxidized upon the absorption of light by the chlorophyll of the PS II reaction center, and the accumulation of four oxidative equivalents in the catalytic center results in the oxidation of two waters to dioxygen in the last step. The recent emergence of X-ray free-electron lasers (XFELs) with intense femtosecond X-ray pulses has opened up opportunities to visualize this reaction in PS II as it proceeds through the catalytic cycle. In this review, we summarize our recent studies of the catalytic reaction in PS II by following the structural changes along the reaction pathway via room-temperature X-ray crystallography using XFELs. The evolution of the electron density changes at the Mn complex reveals notable structural changes, including the insertion of OX from a new water molecule, which disappears on completion of the reaction, implicating it in the O-O bond formation reaction. We were also able to follow the structural dynamics of the protein coordinating with the catalytic complex and of channels within the protein that are important for substrate and product transport, revealing well orchestrated conformational changes in response to the electronic changes at the Mn4Ca cluster.

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  • 41.
    Björkelid, Christofer
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Enzymes in the Mycobacterium tuberculosis MEP and CoA Pathways Targeted for Structure-Based Drug Design2012Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Tuberculosis, caused by the pathogenic bacteria Mycobacterium tuberculosis, is one of the most widespread and deadly infectious diseases today. Treatment of tuberculosis relies on antibiotics that were developed more than 50 years ago. These are now becoming ineffective due to the emergence of antibiotic resistant strains of the bacteria.

    The aim of the research in this thesis was to develop new antibiotics for tuberculosis treatment. To this end, we targeted enzymes from two essential biosynthetic pathways in M. tuberculosis for drug development. The methylerythritol phosphate (MEP) pathway synthesizes a group of compounds called isoprenoids. These compounds have essential roles in all living organisms. The fact that humans utilize a different pathway for isoprenoid synthesis makes the MEP pathway enzymes attractive targets for drug development. We have determined the structures of two essential enzymes from this pathway by X-ray crystallography: 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR) and 2-C-methyl-D-erythritol 4-phosphate cytidylyltransferase (IspD). These are the first structures of these enzymes from M. tuberculosis. Additionally, structures of the IspD enzyme from the related bacteria Mycobacterium smegmatis were determined. We have characterized these enzymes and evaluated the efficiency of a number of inhibitors of the DXR enzyme by biochemical methods. Crystal structures of DXR in complex with some of these inhibitors were also determined.

    The second pathway of interest for drug development is the universal pathway for Coenzyme A biosynthesis. Enzymes in this pathway have essential roles in all living organisms. However, the bacterial enzymes have little similarity to the human homologues. We have determined a number of structures of the M. tuberculosis pantothenate kinase (PanK), the regulatory enzyme of this pathway, in complex with two new classes of inhibitory compounds, and evaluated these by biochemical methods.

    The structures and biochemical characterization of these enzymes provide us with detailed information about their functions and broadens our knowledge of these bacteria. Biochemical and structural information about new inhibitors of these enzymes serve as a starting point for future development of antibiotics against tuberculosis.

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  • 42.
    Björkelid, Christofer
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Raichurkar, Anand Kumar V.
    AstraZeneca India Private Limited.
    Mukherjee, Kakoli
    AstraZeneca India Private Limited.
    Malolanarasimhan, Krishnan
    AstraZeneca India Private Limited.
    Bandodkar, Balachandra
    AstraZeneca India Private Limited.
    Jones, T. Alwyn
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Structural and biochemical characterization of compounds inhibiting Mycobacterium tuberculosis Pantothenate Kinase2013In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 288, no 25, p. 18260-18270Article in journal (Refereed)
    Abstract [en]

    Mycobacterium tuberculosis, the bacterial causative agent oftuberculosis, currently affects millions of people. The emergence of drug-resistant strains makes development of new antibiotics targeting the bacterium a global health priority. Pantothenate kinase, a key enzyme in the universal biosynthesis of the essential cofactor CoA, was targeted in this study to find new tuberculosis drugs. The biochemicalcharacterizations of two new classes of compounds that inhibitpantothenate kinase from M. tuberculosis are described, along with crystal structures of their enzyme-inhibitor complexes. These represent the first crystal structures of this enzyme with engineered inhibitors. Both classes of compounds bind in the active site of the enzyme, overlapping with the binding sites of the natural substrate and product, pantothenateand phosphopantothenate, respectively. One class of compounds also interferes with binding of the cofactor ATP. The complexes were crystallized in two crystal forms, one of which is in a new space group for this enzyme and diffracts to the highest resolution reported for anypantothenate kinase structure. These two crystal forms allowed, for the first time, modeling of the cofactor-binding loop in both open and closed conformations. The structures also show a binding mode of ATP different from that previously reported for the M. tuberculosis enzyme but similar to that in the pantothenate kinases of other organisms.

  • 43.
    Blissing, Annica
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Thiopurine S-methyltransferase - characterization of variants and ligand binding2017Licentiate thesis, comprehensive summary (Other academic)
    Abstract [en]

    Thiopurine S-methyltransferase (TPMT) belongs to the Class I S-adenosylmethionine-dependent methyltransferase (SAM-MT) super family of structurally related proteins. Common to the members of this large protein family is the catalysis of methylation reactions using S-adenosylmethionine (SAM) as a methyl group donor, although SAM-MTs act on a wide range of different substrates and carry out numerous biologically important functions. While the natural function of TPMT is unknown, this enzyme is involved in the metabolism of thiopurines, a class of pharmaceutical substances administered in treatment of immune-related disorders. Specifically, methylation by TPMT inactivates thiopurines and their metabolic intermediates, which reduces the efficacy of clinical treatment and increases the risk of adverse side effects. To further complicate matters, TPMT is a polymorphic enzyme with over 40 naturally occurring variants known to date, most of which exhibit lowered methylation activity towards thiopurines. Consequently, there are individual variations in TPMTmediated thiopurine inactivation, and the administered dose has to be adjusted prior to clinical treatment to avoid harmful side effects.

    Although the clinical relevance of TPMT is well established, few studies have investigated the molecular causes of the reduced methylation activity of variant proteins. In this thesis, the results of biophysical characterization of two variant proteins, TPMT*6 (Y180F) and TPMT*8 (R215H), are presented. While the properties of TPMT*8 were indistinguishable from those of the wild-type protein, TPMT*6 was found to be somewhat destabilized. Interestingly, the TPMT*6 amino acid substitution did not affect the functionality or folding pattern of the variant protein. Therefore, the decreased in vivo functionality reported for TPMT*6 is probably caused by increased proteolytic degradation in response to the reduced stability of this protein variant, rather than loss of function.

    Also presented herein are novel methodological approaches for studies of TPMT and its variants. Firstly, the advantages of using 8-anilinonaphthalene-1-sulfonic acid (ANS) to probe TPMT tertiary structure and active site integrity are presented. ANS binds exclusively to the native state of TPMT with high affinity (KD ~ 0.2 μm) and a 1:1 ratio. The stability of TPMT was dramatically increased by binding of ANS, which was shown to co-localize with the structurally similar adenine moiety of the cofactor SAM. Secondly, an enzyme activity assay based on isothermal titration calorimetry (ITC) is presented. Using this approach, the kinetics of 6-MP and 6-TG methylation by TPMT has been characterized.

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  • 44.
    Boström, Frida
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Biology.
    The influence of differentially expressed Nicotina tabacum Rubisco small subunit on holoenzyme structure2022Independent thesis Advanced level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Characterization of Rubisco plays a crucial role when it comes to the development and understanding of carbon sequestration in plants. This project took place at BMC in Uppsala, in the Gunn lab, and aimed to structure three Rubisco structures and analyze these with regard to the assembly pathway of the biogenesis of Rubisco but also how fast the reaction of binding of atmospheric carbon dioxide takes place with regard to different isoforms of the small subunit. The structural regulations led to the conclusion that an additional step in the assembly pathway would be added when one side of Rubisco had the chaperone BSD2 bound while the other side of Rubisco had the small subunit bound.The different subunits are believed to effect the structure of the LSu. The result also indicate that when the SSu are binding to the LSu octomer the interactions between the BSD2 and the LSu changes. This indicats that the SSu could indirectly facilitate the binding of the SSu on the other side by affecting the interactions of the LSu and the BSD2. Therefore the cooperative binding of the different subunits would be interesting to further evaluate.

    The NtL8B4(S-T1)4 , which is the first model for this structure to be determined, and therefore extended the assembly pathway for the biogenesis of higher plants, had the CABP bound, indicating that this intermediate structure could be analytically competent. This hypothesis is only based on the analyses of the structural determination, therefore further studies are needed to determine whether this is legitimate.

    Teknisk-naturvetenskapliga fakulteten, Uppsala universitet. Utgivni

  • 45.
    Boukharta, Lars
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Computational Modelling of Ligand Complexes with G-Protein Coupled Receptors, Ion Channels and Enzymes2014Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Accurate predictions of binding free energies from computer simulations are an invaluable resource for understanding biochemical processes and drug action. The primary aim of the work described in the thesis was to predict and understand ligand binding to several proteins of major pharmaceutical importance using computational methods.

    We report a computational strategy to quantitatively predict the effects of alanine scanning and ligand modifications based on molecular dynamics free energy simulations. A smooth stepwise scheme for free energy perturbation calculations is derived and applied to a series of thirteen alanine mutations of the human neuropeptide Y1 G-protein coupled receptor and a series of eight analogous antagonists. The robustness and accuracy of the method enables univocal interpretation of existing mutagenesis and binding data. We show how these calculations can be used to validate structural models and demonstrate their ability to discriminate against suboptimal ones. Site-directed mutagenesis, homology modelling and docking were further used to characterize agonist binding to the human neuropeptide Y2 receptor, which is important in feeding behavior and an obesity drug target.  In a separate project, homology modelling was also used for rationalization of mutagenesis data for an integron integrase involved in antibiotic resistance.

    Blockade of the hERG potassium channel by various drug-like compounds, potentially causing serious cardiac side effects, is a major problem in drug development. We have used a homology model of hERG to conduct molecular docking experiments with a series of channel blockers, followed by molecular dynamics simulations of the complexes and evaluation of binding free energies with the linear interaction energy method. The calculations are in good agreement with experimental binding affinities and allow for a rationalization of three-dimensional structure-activity relationships with implications for design of new compounds. Docking, scoring, molecular dynamics, and the linear interaction energy method were also used to predict binding modes and affinities for a large set of inhibitors to HIV-1 reverse transcriptase. Good agreement with experiment was found and the work provides a validation of the methodology as a powerful tool in structure-based drug design. It is also easily scalable for higher throughput of compounds.

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  • 46. Boutet, Sébastien
    et al.
    Lomb, Lukas
    Williams, Garth J
    Barends, Thomas R M
    Aquila, Andrew
    Doak, R Bruce
    Weierstall, Uwe
    DePonte, Daniel P
    Steinbrener, Jan
    Shoeman, Robert L
    Messerschmidt, Marc
    Barty, Anton
    White, Thomas A
    Kassemeyer, Stephan
    Kirian, Richard A
    Seibert, M Marvin
    Montanez, Paul A
    Kenney, Chris
    Herbst, Ryan
    Hart, Philip
    Pines, Jack
    Haller, Gunther
    Gruner, Sol M
    Philipp, Hugh T
    Tate, Mark W
    Hromalik, Marianne
    Koerner, Lucas J
    van Bakel, Niels
    Morse, John
    Ghonsalves, Wilfred
    Arnlund, David
    Bogan, Michael J
    Caleman, Carl
    Fromme, Raimund
    Hampton, Christina Y
    Hunter, Mark S
    Johansson, Linda C
    Katona, Gergely
    Kupitz, Christopher
    Liang, Mengning
    Martin, Andrew V
    Nass, Karol
    Redecke, Lars
    Stellato, Francesco
    Timneanu, Nicusor
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Wang, Dingjie
    Zatsepin, Nadia A
    Schafer, Donald
    Defever, James
    Neutze, Richard
    Fromme, Petra
    Spence, John C H
    Chapman, Henry N
    Schlichting, Ilme
    High-resolution protein structure determination by serial femtosecond crystallography2012In: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 337, no 6092, p. 362-364Article in journal (Refereed)
    Abstract [en]

    Structure determination of proteins and other macromolecules has historically required the growth of high-quality crystals sufficiently large to diffract x-rays efficiently while withstanding radiation damage. We applied serial femtosecond crystallography (SFX) using an x-ray free-electron laser (XFEL) to obtain high-resolution structural information from microcrystals (less than 1 micrometer by 1 micrometer by 3 micrometers) of the well-characterized model protein lysozyme. The agreement with synchrotron data demonstrates the immediate relevance of SFX for analyzing the structure of the large group of difficult-to-crystallize molecules.

  • 47.
    Breidenstein, Annika
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Umeå University, Faculty of Medicine, Wallenberg Centre for Molecular Medicine at Umeå University (WCMM).
    ter Beek, Josy
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Umeå University, Faculty of Medicine, Wallenberg Centre for Molecular Medicine at Umeå University (WCMM).
    Berntsson, Ronnie
    Umeå University, Faculty of Medicine, Wallenberg Centre for Molecular Medicine at Umeå University (WCMM). Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Structural and functional characterization of TraI from pKM101 reveals basis for DNA processing2023In: Life Science Alliance, E-ISSN 2575-1077, Vol. 6, no 4, article id e202201775Article in journal (Refereed)
    Abstract [en]

    Type 4 secretion systems are large and versatile protein machineries that facilitate the spread of antibiotic resistance and other virulence factors via horizontal gene transfer. Conjugative type 4 secretion systems depend on relaxases to process the DNA in preparation for transport. TraI from the well-studied conjugative plasmid pKM101 is one such relaxase. Here, we report the crystal structure of the trans-esterase domain of TraI in complex with its substrate oriT DNA, highlighting the conserved DNA-binding mechanism of conjugative relaxases. In addition, we present an apo structure of the trans-esterase domain of TraI that includes most of the flexible thumb region. This allows us for the first time to visualize the large conformational change of the thumb subdomain upon DNA binding. We also characterize the DNA binding, nicking, and religation activity of the trans-esterase domain, helicase domain, and full-length TraI. Unlike previous indications in the literature, our results reveal that the TraI trans-esterase domain from pKM101 behaves in a conserved manner with its homologs from the R388 and F plasmids.

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  • 48. Brunne, R M
    et al.
    Berndt, Kurt D
    ETH Hönggerberg, Zürich, Switzerland.
    Güntert, P
    Wüthrich, K
    van Gunsteren, W F
    Güntert, P
    Wüthrich, K
    Van Gunsteren, W F
    Structure and internal dynamics of the bovine pancreatic trypsin inhibitor in aqueous solution from long-time molecular dynamics simulations1995In: Proteins: Structure, Function, and Bioinformatics, ISSN 0887-3585, E-ISSN 1097-0134, Vol. 23, no 1, p. 49-62Article in journal (Refereed)
    Abstract [en]

    Structural and dynamic properties of bovine pancreatic trypsin inhibitor (BPTI) in aqueous solution are investigated using two molecular dynamics (MD) simulations: one of 1.4 ns length and one of 0.8 ns length in which atom-atom distance bounds derived from NMR spectroscopy are included in the potential energy function to make the trajectory satisfy these experimental data more closely. The simulated properties of BPTI are compared with crystal and solution structures of BPTI, and found to be in agreement with the available experimental data. The best agreement with experiment was obtained when atom-atom distance restraints were applied in a time-averaged manner in the simulation. The polypeptide segments found to be most flexible in the MD simulations coincide closely with those showing differences between the crystal and solution structures of BPTI.

  • 49.
    Buijs, Jos
    et al.
    Uppsala University, Sweden.
    Ramström, Margareta
    Uppsala University, Sweden.
    Danfelter, Mikael
    Uppsala University, Sweden.
    Larsericsdotter, Helén
    Mälardalen University, Department of Biology and Chemical Engineering.
    Håkansson, Per
    Mälardalen University, Department of Biology and Chemical Engineering.
    Oscarsson, Sven
    Mälardalen University, Department of Biology and Chemical Engineering.
    Localized changes in the structural stability of myoglobin upon adsorption onto silica particles, as studied with hydrogen/deuterium exchange mass spectrometry2003In: Journal of Colloid and Interface Science, ISSN 0021-9797, Vol. 263, no 2, p. 441-448Article in journal (Refereed)
    Abstract [en]

    A new method is presented for monitoring the conformational stability of various parts of a protein that is physically adsorbed onto nanometer-sized silica particles. The method employs hydrogen/deuterium (H/D) exchange of amide hydrogens, a process that is extremely sensitive to structural features of proteins. The resulting mass increase is analyzed with Fourier transform ion cyclotron resonance (FTICR) mass spectrometry. Higher structural specificity is obtained by enzymatically cleaving the adsorbed proteins prior to mass spectrometric analysis. The mass increases of four peptic fragments of myoglobin are followed as a function of the H/D exchange time. The four peptic fragments cover 90% of the myoglobin structure. Two of the peptic fragments, located in the middle of the myoglobin sequence and close to the heme group, do not show any adsorption-induced changes in their structural stability, whereas the more stable C- and N-terminal fragments are destabilized. Interestingly, for the N-terminal fragment, comprising residues 1–29, two distinct and equally large conformational populations are observed. One of these populations has a stability similar to that in solution (−23 kJ/mol), whereas the other population is highly destabilized upon adsorption (−11 kJ/mol).

  • 50. Carbera Rodrigues, Carlos
    et al.
    Ekström, Fredrik
    Wolf, Jana
    Seibt, Henrik
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Stier, Gunter
    Grundström, Christin
    Hung, Shen-Hua
    Shingler, Victoria
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Sauer, Uwe H.
    Umeå University, Faculty of Science and Technology, Department of Chemistry. Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    The crystal structure of the DmpR AAA+ domain suggests a novel mode of ATP-dependent transcriptional controlManuscript (preprint) (Other academic)
1234567 1 - 50 of 495
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