Ändra sökning
Avgränsa sökresultatet
1819202122 1001 - 1050 av 1087
RefereraExporteraLänk till träfflistan
Permanent länk
Referera
Referensformat
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Annat format
Fler format
Språk
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Annat språk
Fler språk
Utmatningsformat
  • html
  • text
  • asciidoc
  • rtf
Träffar per sida
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sortering
  • Standard (Relevans)
  • Författare A-Ö
  • Författare Ö-A
  • Titel A-Ö
  • Titel Ö-A
  • Publikationstyp A-Ö
  • Publikationstyp Ö-A
  • Äldst först
  • Nyast först
  • Skapad (Äldst först)
  • Skapad (Nyast först)
  • Senast uppdaterad (Äldst först)
  • Senast uppdaterad (Nyast först)
  • Disputationsdatum (tidigaste först)
  • Disputationsdatum (senaste först)
  • Standard (Relevans)
  • Författare A-Ö
  • Författare Ö-A
  • Titel A-Ö
  • Titel Ö-A
  • Publikationstyp A-Ö
  • Publikationstyp Ö-A
  • Äldst först
  • Nyast först
  • Skapad (Äldst först)
  • Skapad (Nyast först)
  • Senast uppdaterad (Äldst först)
  • Senast uppdaterad (Nyast först)
  • Disputationsdatum (tidigaste först)
  • Disputationsdatum (senaste först)
Markera
Maxantalet träffar du kan exportera från sökgränssnittet är 250. Vid större uttag använd dig av utsökningar.
  • 1001.
    Virkki, Minttu T.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Agrawal, Nitin
    Edsbäcker, Elin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Cristobal, Susana
    Elofsson, Arne
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Kauko, Anni
    Folding of Aquaporin 1: Multiple evidence that helix 3 can shift out of the membrane core2014Ingår i: Protein Science, ISSN 0961-8368, E-ISSN 1469-896X, Vol. 23, nr 7, s. 981-992Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The folding of most integral membrane proteins follows a two-step process: initially, individual transmembrane helices are inserted into the membrane by the Sec translocon. Thereafter, these helices fold to shape the final conformation of the protein. However, for some proteins, including Aquaporin 1 (AQP1), the folding appears to follow a more complicated path. AQP1 has been reported to first insert as a four-helical intermediate, where helix 2 and 4 are not inserted into the membrane. In a second step, this intermediate is folded into a six-helical topology. During this process, the orientation of the third helix is inverted. Here, we propose a mechanism for how this reorientation could be initiated: first, helix 3 slides out from the membrane core resulting in that the preceding loop enters the membrane. The final conformation could then be formed as helix 2, 3, and 4 are inserted into the membrane and the reentrant regions come together. We find support for the first step in this process by showing that the loop preceding helix 3 can insert into the membrane. Further, hydrophobicity curves, experimentally measured insertion efficiencies and MD-simulations suggest that the barrier between these two hydrophobic regions is relatively low, supporting the idea that helix 3 can slide out of the membrane core, initiating the rearrangement process.

  • 1002.
    Virkki, Minttu T.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Peters, Christoph
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab). Swedish e-Science Research Center (SeRC), Sweden.
    Nilsson, Daniel
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Sörensen, Therese
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Cristobal, Susana
    Wallner, Björn
    Elofsson, Arne
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab). Swedish e-Science Research Center (SeRC), Sweden.
    The Positive Inside Rule Is Stronger When Followed by a Transmembrane Helix2014Ingår i: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 426, nr 16, s. 2982-2991Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The translocon recognizes transmembrane helices with sufficient level of hydrophobicity and inserts them into the membrane. However, sometimes less hydrophobic helices are also recognized. Positive inside rule, orientational preferences of and specific interactions with neighboring helices have been shown to aid in the recognition of these helices, at least in artificial systems. To better understand how the translocon inserts marginally hydrophobic helices, we studied three naturally occurring marginally hydrophobic helices, which were previously shown to require the subsequent helix for efficient translocon recognition. We find no evidence for specific interactions when we scan all residues in the subsequent helices. Instead, we identify arginines located at the N-terminal part of the subsequent helices that are crucial for the recognition of the marginally hydrophobic transmembrane helices, indicating that the positive inside rule is important. However, in two of the constructs, these arginines do not aid in the recognition without the rest of the subsequent helix; that is, the positive inside rule alone is not sufficient. Instead, the improved recognition of marginally hydrophobic helices can here be explained as follows: the positive inside rule provides an orientational preference of the subsequent helix, which in turn allows the marginally hydrophobic helix to be inserted; that is, the effect of the positive inside rule is stronger if positively charged residues are followed by a transmembrane helix. Such a mechanism obviously cannot aid C-terminal helices, and consequently, we find that the terminal helices in multi-spanning membrane proteins are more hydrophobic than internal helices.

  • 1003.
    Virkki, Tuuli Minttu
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Peters, Christoph
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Cristobal, Susana
    Arne, Elofsson
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Insertion of marginally hydrophobic helix in EmrD2014Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    The positive inside rule governs the orientation of membrane proteins in such a way that more positively chargedamino acids are found on the inside of a cell. The exact mechanisms of how this is achieved are not well known, butit is clear that positively charged residues can facilitate the insertion of transmembrane helices that are not sufficientlyhydrophobic to be inserted otherwise. Here, we study one such helix, helix 2 in EmrD. We show that the insertionof this helix is facilitated when followed by positively charged residues and a hydrophobic helix. Surprisingly thepositively charged residues are not sufficient alone. This further strengthens our earlier observations that the last helixneeds to be more hydrophobic than previous helices.

  • 1004.
    Visa, Neus
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Percipalle, Piergiorgio
    Nuclear Functions of Actin2010Ingår i: COLD SPRING HARBOR PERSPECT B, ISSN 1943-0264, Vol. 2, nr 4, s. a000620-Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Actin participates in several essential processes in the cell nucleus. Even though the presence of actin in the nucleus was proposed more than 30 years ago, nuclear processes that require actin have been only recently identified. Actin is part of chromatin remodeling complexes; it is associated with the transcription machineries; it becomes incorporated into newly synthesized ribonucleoproteins; and it influences long-range chromatin organization. As in the cytoplasm, nuclear actin works in conjunction with different types of actin-binding proteins that regulate actin function and bridge interactions between actin and other nuclear components.

  • 1005.
    von Ballmoos, Christoph
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Adelroth, Pia
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Gennis, Robert B.
    Brzezinski, Peter
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Proton transfer in ba(3) cytochrome c oxidase from Thermus thermophilus2012Ingår i: Biochimica et Biophysica Acta - Bioenergetics, ISSN 0005-2728, E-ISSN 1879-2650, Vol. 1817, nr 4, s. 650-657Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    The respiratory heme-copper oxidases catalyze reduction of O-2 to H2O, linking this process to transmembrane proton pumping. These oxidases have been classified according to the architecture, location and number of proton pathways. Most structural and functional studies to date have been performed on the A-class oxidases, which includes those that are found in the inner mitochondrial membrane and bacteria such as Rhodobacter sphaeroides and Paracoccus denitrificans (aa(3)-type oxidases in these bacteria). These oxidases pump protons with a stoichiometry of one proton per electron transferred to the,catalytic site. The bacterial A-class oxidases use two proton pathways (denoted by letters D and K, respectively), for the transfer of protons to the catalytic site, and protons that are pumped across the membrane. The B-type oxidases such as, for example, the ba(3) oxidase from Thermus thermophilus, pump protons with a lower stoichiometry of 0.5 H+/electron and use only one proton pathway for the transfer of all protons. This pathway overlaps in space with the K pathway in the A class oxidases without showing any sequence homology though. Here, we review the functional properties of the A- and the B-class ba3 oxidases with a focus on mechanisms of proton transfer and pumping. This article is part of a Special Issue entitled: Respiratory Oxidases.

  • 1006.
    von Ballmoos, Christoph
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Lachmann, Peter
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Gennis, Robert B.
    Adelroth, Pia
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Brzezinski, Peter
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Timing of Electron and Proton Transfer in the ba(3) Cytochrome c Oxidase from Thermus thermophilus2012Ingår i: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 51, nr 22, s. 4507-4517Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Heme-copper oxidases are membrane-bound proteins that catalyze the reduction of O-2 to H2O, a highly exergonic reaction. Part of the free energy of this reaction is used for pumping of protons across the membrane. The ba(3) oxidase from Thermus thermophilus presumably uses a single proton pathway for the transfer of substrate protons used during O-2 reduction as well as for the transfer of the protons that are pumped across the membrane. The pumping stoichiometry (0.5 H+/electron) is lower than that of most other (mitochondrial-like) oxidases characterized to date (1 H+/electron). We studied the pH dependence and deuterium isotope effect of the kinetics of electron and proton transfer reactions in the ba3 oxidase. The results from these studies suggest that the movement of protons to the catalytic site and movement to a site located some distance from the catalytic site [proposed to be a proton-loading site (PLS) for pumped protons] are separated in time, which allows individual investigation of these reactions. A scenario in which the uptake and release of a pumped proton occurs upon every second transfer of an electron to the catalytic site would explain the decreased proton pumping stoichiometry compared to that of mitochondrial-like oxidases.

  • 1007.
    von Ballmoos, Christoph
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Wiedenmann, Alexander
    Dimroth, Peter
    Essentials for ATP Synthesis by F1F0 ATP Synthases2009Ingår i: Annual Review of Biochemistry, ISSN 0066-4154, E-ISSN 1545-4509, Vol. 78, s. 649-672Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    The majority of cellular energy in the form of adenosine triphosphate (ATP) is synthesized by the ubiquitous F1F0 ATP synthase. Power for ATP synthesis derives from an electrochemical proton (or Na+) gradient, which drives rotation of membranous F-0 motor components. Efficient rotation not only requires a significant driving force (Delta mu H+), consisting of membrane potential (Delta psi) and proton concentration gradient (Delta pH), but also a high proton concentration at the source P side. In vivo this is maintained by dynamic proton movements across and along the surface of the membrane. The torque-generating unit consists of the interface of the rotating c ring and the stator a subunit. Ion translocation through this unit involves a sophisticated interplay between the c-ring binding sites, the stator arginine, and the coupling ions on both sides of the membrane. c-ring rotation is transmitted to the eccentric shaft gamma-subunit to elicit conformational changes in the catalytic sites of F-1, leading to ATP synthesis.

  • 1008. Vukojevic, Vladana
    et al.
    Gräslund, Astrid
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Bakalkin, Georgy
    Fluorescence Imaging with Single-Molecule Sensitivity and Fluorescence Correlation Spectroscopy of Cell-Penetrating Neuropeptides2011Ingår i: Neuropeptides: methods and protocols / [ed] Merighi, A., Humana Press, 2011, s. 147-170Kapitel i bok, del av antologi (Refereegranskat)
    Abstract [en]

    Neuropeptide plasma membrane interactions in the absence of a corresponding specific receptor may result in neuropeptide translocation into the cell. Trans location across the plasma membrane may represent a previously unknown mechanism by which neuropeptides can signal information to the cell interior. We introduce here two complementary optical methods with single-molecule sensitivity, fluorescence imaging with avalanche photodiode detectors (APD imaging) and fluorescence correlation spectroscopy (FCS), and demonstrate how they may be applied for the analysis of neuropeptide ability to penetrate into live cells in real time. APD imaging enables us to visualize fluorescently labeled neuropeptide molecules at very low, physiologically relevant concentrations, whereas FCS enables us to characterize quantitatively their concentration and diffusion properties in different cellular compartments. Application of these methodologies for the analysis of the endogenous opioid peptide dynorphin A (Dyn A), a ligand for the kappa-opioid receptor (KOP), demonstrated that this neuropeptide may translocate across the plasma membrane of living cells and enter the cellular interior without binding to its cognate receptor.

  • 1009.
    Wagner, Samuel
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    From Biogenesis to Overexpression of Membrane Proteins in Escherichia coli2008Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    In both pro- and eukaryotes 20-30% of all genes encode alpha-helical transmembrane domain proteins, which act in various and often essential capacities. Notably, membrane proteins play key roles in disease and they constitute more than half of all known drug targets.

    The natural abundance of membrane proteins is in general too low to conveniently isolate sufficient material for functional and structural studies. Therefore, most membrane proteins have to be obtained through overexpression. Escherichia coli is one of the most successful hosts for overexpression of recombinant proteins. While the production of soluble proteins is comparably straightforward, overexpression of membrane proteins remains a challenging task. The yield of membrane localized recombinant membrane protein is usually low and inclusion body formation is a serious problem. Furthermore, membrane protein overexpression is often toxic to the host cell. Although several reasons can be postulated, the basis of these difficulties is not completely understood, preventing the design of rational strategies to improve membrane protein overexpression yields.

    The objective of my Ph.D. studies has been to improve membrane protein overexpression in E. coli by a) understanding membrane protein overexpression from the perspective of membrane protein biogenesis, b) systematically investigating the physiological response to overexpression of membrane proteins and c) engineering strains that are optimized for membrane protein overexpression based on insights resulting from these studies.

    By working toward these objectives, I was able to identify and alleviate one of the major bottlenecks of membrane protein overexpression in E. coli: saturation of the Sec-translocon could be overcome by harmonizing translation and membrane insertion of the recombinant membrane protein. This minimized the toxic effects of overexpression and thus resulted in increased membrane protein-producing biomass.

  • 1010.
    Wagner, Samuel
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Pop, Ovidiu
    Haan, Gert-Jan
    Baars, Louise
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Klepsch, Mirjam
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Genevaux, Pierre
    Luirink, Joen
    de Gier, Jan-Willem
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Biogenesis of MalF and the MalFGK2 maltose transport complex in Escherichia coli requires YidC2008Ingår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, nr 283, s. 17881-17890Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The polytopic inner membrane protein MalF is a constituent of the MalFGK2 maltose transport complex in Escherichia coli. We have studied the biogenesis of MalF using a combination of in vivo and in vitro approaches. MalF is targeted via the SRP pathway to the Sec/YidC insertion site. Despite close proximity of nascent MalF to YidC during insertion, YidC is not required for the insertion of MalF into the membrane. However, YidC is required for the stability of MalF and the formation of the MalFGK2 maltose transport complex. Our data indicate that YidC supports the folding of MalF into a stable conformation before it is incorporated into the maltose transport complex.

  • 1011.
    Wahlstedt, Helene
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    The subcellular localization of ADAR determines A-to-I editing levels in developing neuronsManuskript (preprint) (Övrigt vetenskapligt)
  • 1012.
    Wahlstedt, Helene
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Öhman, Marie
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Site-selective versus promiscuous A-to-I editing2011Ingår i: Wiley Interdiscip Reviews - RNA, ISSN 1757-7012, Vol. 2, nr 6, s. 761-771Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    RNA editing by adenosine deamination is acting on polymerase II derived transcripts in all metazoans. Adenosine-to-inosine (A-to-I) editing is mediated by the adenosine deaminase that acts on RNA (ADAR) enzymes. Two types of adenosine to inosine (A-to-I) RNA editing have been defined: site selective and hyper-editing. Typically, in site selectively edited substrates, one or a few A-to-I sites are edited in double-stranded RNA structures, frequently interrupted by single-stranded bulges and loops. Hyper-editing occurs in long stretches of duplex RNA where multiple adenosines are subjected to deamination. In this review, recent findings on editing within noncoding RNA as well as examples of site selective editing within coding regions are presented. We discuss how these two editing events have evolved and the structural differences between a site selective and hyper-edited substrate.

  • 1013.
    Wahlström, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    NMR studies on interactions between the amyloid β peptide and selected molecules2011Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Alzheimer’s disease is an incurable neurodegenerative disorder linked to the amyloid β (Aβ) peptide, a 38-43 residue peptide. The detailed molecular disease mechanism(s) is (are) unknown, but oligomeric Aβ structures are proposed to be involved.

    In common for the papers in this thesis is interactions; interactions between Aβ(1-40) and selected molecules and metal ions. The purpose has been to find out more about the structural states that Aβ can adopt, in particular the β-sheet state, which probably is linked to the oligomeric structures. The methods used have been nuclear magnetic resonance (NMR), circular dichroism (CD) and fluorescence spectroscopy using Thioflavin T (ThT).

    Upon addition of SDS/LiDS detergent or Congo red (CR) to Aβ(1-40), the initial random coil/PII-helix state was transformed into β-sheet and, in the case of detergent, a final α-helical state. In contrast to SDS/LiDS and CR, the dimeric Affibody molecule locks monomeric Aβ(1-40) in a β-hairpin state. It was found that by truncating the flexible N-terminal end of the Affibody molecule its affinity to Aβ was improved. The aggregation of Aβ(1-40) was further studied in the presence of a β-cyclodextrin dimer by a kinetic assay using ThT. Although having a weak dissociation constant in the millimolar range, the β-cyclodextrin dimer modified the aggregation pathways of Aβ.

    Finally Aβ(1-40) was studied in presence of Cu2+ and Zn2+ at physiological and low pH. Cu2+ was observed to maintain its specific binding to Aβ when decreasing the pH to 5.5 while Zn2+ behaved differently. This could be of importance in the Alzheimer’s disease brain in which the environment can become acidic due to inflammation.       

    In conclusion the results show that Aβ(1-40) is very sensitive to its environment, responding by adopting different conformations and aggregating in aqueous solutions. The β-sheet state is induced by varying molecules with different properties, properties that govern the final Aβ state.

  • 1014.
    Wahlström, Anna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Cukalevski, Risto
    Danielsson, Jens
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Jarvet, Jueri
    Onagi, Hideki
    Rebek, Julius, Jr.
    Linse, Sara
    Gräslund, Astrid
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Specific binding of a beta cyclodextrin dimer to the amyloid beta peptide modulates the peptide aggregation process2012Ingår i: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 51, nr 21, s. 4280-4289Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Alzheimer's disease involves progressive neuronal loss. Linked to the disease is the amyloid beta (A beta) peptide, a 38-43-amino acid peptide found in extracellular amyloid plaques in the brain. Cyclodextrins are nontoxic, cone-shaped oligosaccharides with a hydrophilic exterior and a hydrophobic cavity making them suitable hosts for aromatic guest molecules in water. beta-Cyclodextrin consists of seven alpha-D-glucopyranoside units and has been shown to reduce the level of fibrillation and neurotoxicity of A beta. We have studied the interaction between A beta and a beta-cyclodextrin dimer, consisting of two beta-cyclodextrin monomers connected by a flexible linker. The beta-cyclodextrin monomer has been found to interact with A beta(1-40) at sites Y10, F19, and/or F20 with a dissociation constant (K-D) of 3.9 +/- 2.0 mM. Here H-1-N-15 and H-1-C-13 heteronuclear single-quantum correlation nuclear magnetic resonance (NMR) spectra show that in addition, the beta-cyclodextrin monomer and dimer bind to the histidines. NMR translational diffusion experiments reveal the increased affinity of the beta-cyclodextrin dimer (apparent K-D of 1.1 +/- .5 mM) for A beta(1-40) compared to that of the beta-cyclodextrin monomer. Kinetic aggregation experiments based on thioflavin T fluorescence indicate that the dimer at 0.05-5 mM decreases the lag time of A beta aggregation, while a concentration of 10 mM increases the lag time. The beta-cyclodextrin monomer at a high concentration decreases the lag time of the aggregation. We conclude that cyclodextrin monomers and dimers have specific, modulating effects on the A beta(1-40) aggregation process. Transmission electron microscopy shows that the regular fibrillar aggregates formed by A beta(1-40) alone are replaced by a major fraction of amorphous aggregates in the presence of the beta-cyclodextrin dimer.

  • 1015.
    Wahlström, Anna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Cukalevski, Risto
    Danielsson, Jens
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Jarvet, Jüri
    Onagi, Hideki
    Rebek Jr., Julius
    Linse, Sara
    Gräslund, Astrid
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Specific binding of an engineered β-cyclodextrin dimer to the amyloid β peptide modulates the peptide aggregation processManuskript (preprint) (Övrigt vetenskapligt)
  • 1016.
    Wahlström, Anna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Hugonin, Loïc
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Perálvarez-Marín, Alex
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Jarvet, Jüri
    Gräslund, Astrid
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Secondary structure conversions of Alzheimer’s Aβ(1–40) peptide induced by membrane-mimicking detergents2008Ingår i: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 275, nr 20, s. 5117-5128Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The amyloid β peptide (Aβ) with 39–42 residues is the major component of amyloid plaques found in brains of Alzheimer’s disease patients, and soluble oligomeric peptide aggregates mediate toxic effects on neurons. The Aβ aggregation involves a conformational change of the peptide structure to β-sheet. In the present study, we report on the effect of detergents on the structure transitions of Aβ, to mimic the effects that biomembranes may have. In vitro, monomeric Aβ(1–40) in a dilute aqueous solution is weakly structured. By gradually adding small amounts of sodium dodecyl sulfate (SDS) or lithium dodecyl sulfate to a dilute aqueous solution, Aβ(1–40) is converted to β-sheet, as observed by CD at 3 °C and 20 °C. The transition is mainly a two-state process, as revealed by approximately isodichroic points in the titrations. Aβ(1–40) loses almost all NMR signals at dodecyl sulfate concentrations giving rise to the optimal β-sheet content (approximate detergent/peptide ratio = 20). Under these conditions, thioflavin T fluorescence measurements indicate a maximum of aggregated amyloid-like structures. The loss of NMR signals suggests that these are also involved in intermediate chemical exchange. Transverse relaxation optimized spectroscopy NMR spectra indicate that the C-terminal residues are more dynamic than the others. By further addition of SDS or lithium dodecyl sulfate reaching concentrations close to the critical micellar concentration, CD, NMR and FTIR spectra show that the peptide rearranges to form a micelle-bound structure with α-helical segments, similar to the secondary structures formed when a high concentration of detergent is added directly to the peptide solution.

  • 1017.
    Waldholm, Johan
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Wang, Zhi
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Brodin, David
    Tyagi, Anu
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik. University of Würzburg, Germany.
    Yu, Simei
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Theopold, Uli
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Östlund Farrants, Ann Kristin
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut.
    Visa, Neus
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    SWI/SNF regulates the alternative processing of a specific subset of pre-mRNAs in Drosophila melanogaster2011Ingår i: BMC Molecular Biology, ISSN 1471-2199, E-ISSN 1471-2199, Vol. 12, artikel-id 46Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Background: The SWI/SNF chromatin remodeling factors have the ability to remodel nucleosomes and play essential roles in key developmental processes. SWI/SNF complexes contain one subunit with ATPase activity, which in Drosophila melanogaster is called Brahma (Brm). The regulatory activities of SWI/SNF have been attributed to its influence on chromatin structure and transcription regulation, but recent observations have revealed that the levels of Brm affect the relative abundances of transcripts that are formed by alternative splicing and/or polyadenylation of the same pre-mRNA.

    Results: We have investigated whether the function of Brm in pre-mRNA processing in Drosophila melanogaster is mediated by Brm alone or by the SWI/SNF complex. We have analyzed the effects of depleting individual SWI/SNF subunits on pre-mRNA processing throughout the genome, and we have identified a subset of transcripts that are affected by depletion of the SWI/SNF core subunits Brm, Snr1 or Mor. The fact that depletion of different subunits targets a subset of common transcripts suggests that the SWI/SNF complex is responsible for the effects observed on pre-mRNA processing when knocking down Brm. We have also depleted Brm in larvae and we have shown that the levels of SWI/SNF affect the pre-mRNA processing outcome in vivo.

    Conclusions: We have shown that SWI/SNF can modulate alternative pre-mRNA processing, not only in cultured cells but also in vivo. The effect is restricted to and specific for a subset of transcripts. Our results provide novel insights into the mechanisms by which SWI/SNF regulates transcript diversity and proteomic diversity in higher eukaryotes.

  • 1018.
    Walles, Björn
    Stockholms universitet.
    Chloroplast morphogenesis in biochemical mutants1967Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
  • 1019.
    Waluk, Dominik P.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Schultz, Niklas
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Hunt, Mary C.
    Identification of glycine N-acyltransferase-like 2 (GLYATL2) as a transferase that produces N-acyl glycines in humans.2010Ingår i: The FASEB Journal, ISSN 0892-6638, E-ISSN 1530-6860, Vol. 24, nr 8, s. 2795-2803Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The discovery of glycine conjugates of long-chain fatty acids (N-acyl glycines) in the brain and other non-neuronal tissues has led to the identification of an emerging class of bioactive lipids. The biological activities of N-acyl glycines include antinociceptive, anti-inflammatory and antiproliferative effects, and activation of G-protein-coupled receptors. However, despite the fact that N-acyl glycines are emerging as a distinct lipid signaling family, pathways for their production are not fully elucidated. Here we report on the characterization of human glycine N-acyltransferase-like 2 (hGLYATL2), a member of a gene family of 4 putative glycine conjugating enzymes, and show that it synthesizes various N-acyl glycines. Recombinantly expressed hGLYATL2 efficiently conjugated oleoyl-CoA, arachidonoyl-CoA, and other medium- and long-chain acyl-CoAs to glycine. The enzyme was specific for glycine as an acceptor molecule, and preferentially produced N-oleoyl glycine. The hGLYATL2 enzyme is localized to the endoplasmic reticulum, and the mRNA shows highest expression in salivary gland and trachea, but is also detected in spinal cord and skin fibroblasts. The expression pattern and the identification of high levels of N-acyl glycines in skin and lung may indicate a role for N-acyl glycines in barrier function/immune response and the potential role of hGLYATL2 in this regard is discussed.

  • 1020.
    Waluk, Dominik P.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Sucharski, Filip
    Sipos, Laszlo
    Silberring, Jerzy
    Hunt, Mary C.
    Reversible lysine acetylation regulates the activity of human glycine n-acyltransferase-like 2 (hGLYATL2): Implications for production of glycine-conjugated signalling molecules2012Ingår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 287, nr 20, s. 16158-16167Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Lysine acetylation is a major post-translational modification of proteins, and regulates many physiological processes such as metabolism, cell migration, ageing and inflammation. Proteomic studies have identified numerous lysine-acetylated proteins in human and mouse models (Kim et al, (2006) Mol. Cell. 23, 607-618). One family of proteins identified in this study was the murine glycine N-acyltransferase (GLYAT) enzymes, which are acetylated on lysine 19 (K19). Lysine 19 is a conserved residue in human glycine N-acyltransferase-like 2 (hGLYATL2) and in several other species, showing that this residue may be important for enzyme function. Mutation of lysine 19 (K19) in recombinant hGLYATL2 to glutamine (K19Q) and arginine (K19R) resulted in a 50-80% lower production of N-oleoyl glycine and N-arachidonoylglycine, indicating that lysine 19 is important for enzyme function. LC/MS/MS confirmed that K19 is not acetylated in wild-type hGLYATL2, indicating that K19 requires to be deacetylated for full activity. The hGLYATL2 enzyme conjugates medium- and long-chain saturated and unsaturated acyl-CoA esters to glycine, resulting in the production of N-oleoyl glycine and also N-arachidonoyl glycine. N-oleoyl glycine and N-arachidonoyl glycine are structurally and functionally related to endocannabinoids and have been identified as signalling molecules that regulate functions like the perception of pain, body temperature, and also have anti-inflammatory properties. In conclusion, acetylation of lysine(s) in hGLYATL2 regulate the enzyme activity, thus linking post-translational modification of proteins with the production of biological signalling molecules, the N-acyl glycines.

  • 1021.
    Waluk, Dominik P.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Tillander, Veronika
    Schultz, Niklas
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Hunt, Mary C.
    Molecular characterization of two members of the glycine N-acyltransferase gene family in human: glycine N-acyl transferase-like 1 (GLYATL1) and glycine N-acyltransferase-like 3 (GLYATL3).Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    N-acyl amino acids are a group of endogenous lipid mediators that regulate a variety of cellular physiological functions. The discovery of N-acyl amino acids in many biological systems has allowed research to focus on their functions as well as pathways for production of these signalling lipids.

    We have previously identified that human glycine N-acyltransferase-like 2 (hGLYATL2) is involved in the enzymatic formation of N-acyl glycines. hGLYATL2 is localized in a gene cluster with other glycine N-acyltransferase genes. Here, we have characterized human glycine N-acyltransferase-like 1 (hGLYATL1) and human glycine N-acyltransferase-like 3 (hGLYATL3), which are members of this gene family. Our results show that hGLYATL1 is localized to the endoplasmic reticulum (ER) but the intracellular localization of hGLYATL3 remains to be determined. The hGLYATL1 mRNA shows highest expression in liver and kidney, whereas mRNA of hGLYATL3 is expressed in pancreas and liver. Using bioinformatics we determined the overall three-dimensional (3D) structures of hGLYATL1 and hGLYATL3 enzymes, with predicted binding site residues.

    In summary, we have characterized novel members of glycine N-acyltransferases that may be involved in the production of lipid signalling molecules, in particular N-acyl glycines.

  • 1022.
    Waluk, Dominik P.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Vielfort, Katarina
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Derakhshan, Sepide
    Aro, Helena
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Hunt, Mary C
    N-acyl taurines trigger insulin secretion by increasing calcium flux in pancreatic b-cellsManuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Pancreatic b-cells secrete insulin in response to various stimuli to control blood glucose levels. This insulin release is the result of a complex interplay between signalling, membrane potential and intracellular calcium levels. Various nutritional and hormonal factors are involved in regulating this process. N-acyl taurines are a group of fatty acids which are amidated (or conjugated) to taurine and little is known about their physiological functions. In this study, treatment of pancreatic b-cell lines (HIT-T15) and rat islet cell lines (INS-1) with N-acyl taurines (N-arachidonoyl taurine and N-oleoyl taurine), induced a high frequency of calcium oscillations in these cells. Treatment with N-arachidonoyl taurine and N-oleoyl taurine also resulted in a significant increase in insulin secretion from pancreatic b-cell lines as determined by insulin release assay and immunofluorescence (p<0.05). Our data also show that the transient receptor potential vanilloid 1 (TRPV1) channel is involved in insulin secretion in response to N-arachidonoyl taurine and N-oleoyl taurine treatment. However our data also suggest that receptors other than TRPV1 are involved in the insulin secretion response to treatment with N-oleoyl taurine.

  • 1023.
    Waluk, Dominik Paweł
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Biosynthesis and physiological functions of N-acyl amino acids2012Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    N-acyl amino acids are lipid signalling molecules that have recently been identified in biological systems. These lipids are structurally related to the endocannabinoids, although they do not activate cannabinoid receptors. In 2001, N-arachidonoyl glycine was the first signalling lipid in this group to be identified in bovine and rat brain and since then, about 50 novel N-acyl amino acids have been identified in mammalian systems. These N-acyl amino acids are involved in regulating pain processes, are anti-inflammatory and regulate body temperature, but the metabolic pathways for production and metabolism remain poorly understood.

    This thesis focussed on the identification of pathways for production and regulation of N-acyl amino acids, in particular N-acyl glycines, and in identifying physiological functions for N-acyl amino acids (particularly N-acyl taurines). Our results identified an enzymatic pathway for production of N-acyl glycines in human and we identified that the human glycine N-acyltransferase-like 2 (hGLYATL2) conjugates (amidates) medium- and long-chain, saturated and unsaturated acyl-CoAs with glycine, to produce N-acyl glycines, with the preferential production of N-oleoyl glycine. Furthermore, we have characterized two other members of the gene family of glycine N-acyltransferases (GLYATs) in human, the hGLYATL1 and hGLYATL3 that may be involved in the production of N-acyl amino acids.

    As N-acyl glycines are bioactive signalling molecules, it is likely their production requires a rapid on/off switch. The post-translational modification of proteins can result in enzyme regulation, without the need for transcriptional regulation. We have identified that hGLYATL2 is regulated by acetylation/deacetylation on lysine 19, and using mutation analysis, we show that deacetylation of lysine 19 is important for full enzyme activity.

    The physiological functions of N-acyl amino acids are not well studied to date. In this thesis, we have identified that N-arachidonoyl taurine and N-oleoyl taurine trigger insulin secretion by increasing the calcium flux in pancreatic b-cells via the activation of transient receptor potential vanilloid subfamily 1 (TRPV1).

    This work on N-acyl amino acids has led us to identify new pathways and physiological functions for these lipid signalling molecules, which advances our knowledge of the importance of these lipids in mammalian systems.

  • 1024.
    Waluk, Dominik
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Sucharski, Filip
    Sipos, Laszlo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Derakhshan, Sepide
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Sillbering, Jerzy
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Hunt, Mary
    Reversible lysine acetylation regulates the activity of human glycine N-acyltransferase 2 (hGLYATL2)-implications for the production of glycine conjugated signalling lipids2011Ingår i: Chemistry and Physics of Lipids, ISSN 0009-3084, E-ISSN 1873-2941, Vol. 164, s. s35-S35Artikel i tidskrift (Övrigt vetenskapligt)
  • 1025. Wang, Kaituo
    et al.
    Preisler, Sarah Spruce
    Zhang, Liying
    Cui, Yanxiang
    Missel, Julie Winkel
    Gronberg, Christina
    Gotfryd, Kamil
    Lindahl, Erik
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Andersson, Magnus
    Calloe, Kirstine
    Egea, Pascal F.
    Klaerke, Dan Arne
    Pusch, Michael
    Pedersen, Per Amstrup
    Zhou, Z. Hong
    Gourdon, Pontus
    Structure of the human ClC-1 chloride channel2019Ingår i: PLoS biology, ISSN 1544-9173, E-ISSN 1545-7885, Vol. 17, nr 4, artikel-id e3000218Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    ClC-1 protein channels facilitate rapid passage of chloride ions across cellular membranes, thereby orchestrating skeletal muscle excitability. Malfunction of ClC-1 is associated with myotonia congenita, a disease impairing muscle relaxation. Here, we present the cryo-electron microscopy (cryo-EM) structure of human ClC-1, uncovering an architecture reminiscent of that of bovine ClC-K and CLC transporters. The chloride conducting pathway exhibits distinct features, including a central glutamate residue (fast gate) known to confer voltage-dependence (a mechanistic feature not present in ClC-K), linked to a somewhat rearranged central tyrosine and a narrower aperture of the pore toward the extracellular vestibule. These characteristics agree with the lower chloride flux of ClC-1 compared with ClC-K and enable us to propose a model for chloride passage in voltage-dependent CLC channels. Comparison of structures derived from protein studied in different experimental conditions supports the notion that pH and adenine nucleotides regulate ClC-1 through interactions between the so-called cystathionine-beta-synthase (CBS) domains and the intracellular vestibule (slow gating). The structure also provides a framework for analysis of mutations causing myotonia congenita and reveals a striking correlation between mutated residues and the phenotypic effect on voltage gating, opening avenues for rational design of therapies against ClC-1-related diseases.

  • 1026. Wangsell, Fredrik
    et al.
    Nordeman, Patrik
    Savmarker, Jonas
    Emanuelsson, Rikard
    Jansson, Katarina
    Lindberg, Jimmy
    Rosenquist, Asa
    Samuelsson, Bertil
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för organisk kemi.
    Larhed, Mats
    Investigation of alpha-phenylnorstatine and alpha-benzylnorstatine as transition state isostere motifs in the search for new BACE-1 inhibitors2011Ingår i: Bioorganic & Medicinal Chemistry, ISSN 0968-0896, E-ISSN 1464-3391, Vol. 19, nr 1, s. 145-155Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Inhibition of the BACE-1 protease enzyme has over the recent decade developed into a promising drug strategy for Alzheimer therapy. In this report, more than 20 new BACE-1 protease inhibitors based on alpha-phenylnorstatine, alpha-benzylnorstatine, iso-serine, and beta-alanine moieties have been prepared. The inhibitors were synthesized by applying Fmoc solid phase methodology and evaluated for their inhibitory properties. The most potent inhibitor, tert-alcohol containing (R)-12 (IC(50) = 0.19 mu M) was co-crystallized in the active site of the BACE-1 protease, furnishing a novel binding mode in which the N-terminal amine makes a hydrogen bond to one of the catalytic aspartic acids.

  • 1027. Wanngren, Johanna
    et al.
    Lara Vasques, Patricia
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Öjemalm, Karin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Maioli, Silvia
    Moradi, Nasim
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Chen, Lu
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Tjernberg, Lars O.
    Lundkvist, Johan
    Nilsson, IngMarie
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Karlström, Helena
    Changed membrane integration and catalytic site conformation are two mechanisms behind the increased Aβ42/Aβ40 ratio by presenilin 1 familial Alzheimer-linked mutations.2014Ingår i: FEBS Open Bio, E-ISSN 2211-5463, Vol. 4, s. 393-406Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The enzyme complex γ-secretase generates amyloid β-peptide (Aβ), a 37-43-residue peptide associated with Alzheimer disease (AD). Mutations in presenilin 1 (PS1), the catalytical subunit of γ-secretase, result in familial AD (FAD). A unifying theme among FAD mutations is an alteration in the ratio Aβ species produced (the Aβ42/Aβ40 ratio), but the molecular mechanisms responsible remain elusive. In this report we have studied the impact of several different PS1 FAD mutations on the integration of selected PS1 transmembrane domains and on PS1 active site conformation, and whether any effects translate to a particular amyloid precursor protein (APP) processing phenotype. Most mutations studied caused an increase in the Aβ42/Aβ40 ratio, but via different mechanisms. The mutations that caused a particular large increase in the Aβ42/Aβ40 ratio did also display an impaired APP intracellular domain (AICD) formation and a lower total Aβ production. Interestingly, seven mutations close to the catalytic site caused a severely impaired integration of proximal transmembrane/hydrophobic sequences into the membrane. This structural defect did not correlate to a particular APP processing phenotype. Six selected FAD mutations, all of which exhibited different APP processing profiles and impact on PS1 transmembrane domain integration, were found to display an altered active site conformation. Combined, our data suggest that FAD mutations affect the PS1 structure and active site differently, resulting in several complex APP processing phenotypes, where the most aggressive mutations in terms of increased Aβ42/Aβ40 ratio are associated with a decrease in total γ-secretase activity.

  • 1028. Wanschers, Bas F. J.
    et al.
    Szklarczyk, Radek
    van den Brand, Mariel A. M.
    Jonckheere, An
    Suijskens, Janneke
    Smeets, Roel
    Rodenburg, Richard J.
    Stephan, Katharina
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Helland, Ingrid B.
    Elkamil, Areej
    Rootwelt, Terje
    Ott, Martin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    van den Heuvel, Lambert
    Nijtmans, Leo G.
    Huynen, Martijn A.
    A mutation in the human CBP4 ortholog UQCC3 impairs complex III assembly, activity and cytochrome b stability2014Ingår i: Human Molecular Genetics, ISSN 0964-6906, E-ISSN 1460-2083, Vol. 23, nr 23, s. 6356-6365Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Complex III (cytochrome bc(1)) is a protein complex of the mitochondrial inner membrane that transfers electrons from ubiquinol to cytochrome c. Its assembly requires the coordinated expression of mitochondrial-encoded cytochrome b and nuclear-encoded subunits and assembly factors. Complex III deficiency is a severe multisystem disorder caused by mutations in subunit genes or assembly factors. Sequence-profile-based orthology predicts C11orf83, hereafter named UQCC3, to be the ortholog of the fungal complex III assembly factor CBP4. We describe a homozygous c.59T > A missense mutation in UQCC3 from a consanguineous patient diagnosed with isolated complex III deficiency, displaying lactic acidosis, hypoglycemia, hypotonia and delayed development without dysmorphic features. Patient fibroblasts have reduced complex III activity and lower levels of the holocomplex and its subunits than controls. They have no detectable UQCC3 protein and have lower levels of cytochrome b protein. Furthermore, in patient cells, cytochrome b is absent from a high-molecular-weight complex III. UQCC3 is reduced in cells depleted for the complex III assembly factors UQCC1 and UQCC2. Conversely, absence of UQCC3 in patient cells does not affect UQCC1 and UQCC2. This suggests that UQCC3 functions in the complex III assembly pathway downstream of UQCC1 and UQCC2 and is consistent with what is known about the function of Cbp4 and of the fungal orthologs of UQCC1 and UQCC2, Cbp3 and Cbp6. We conclude that UQCC3 functions in complex III assembly and that the c.59T > A mutation has a causal role in complex III deficiency.

  • 1029.
    Warholm, Per
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Light, Sara
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Identification of a Non-Pentapeptide Region Associated with Rapid Mycobacterial Evolution2016Ingår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 11, nr 5, artikel-id e0154059Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    A large portion of the coding capacity of Mycobacterium tuberculosis is devoted to the production of proteins containing several copies of the pentapeptide-2 repeat, namely the PE/PPE_MPTR proteins. Protein domain repeats have a variety of binding properties and are involved in protein-protein interactions as well as binding to other ligands such as DNA and RNA. They are not as common in prokaryotes, compared to eukaryotes, but the enrichment of pentapeptide-2 repeats in Mycobacteria constitutes an exception to that rule. The genes encoding the PE/PPE_MPTR proteins have undergone many rearrangements and here we have identified the expansion patterns across the Mycobacteria. We have performed a reclassification of the PE/PPE_MPTR proteins using cohesive regions rather than sparse domain architectures. It is clear that these proteins have undergone large insertions of several pentapeptide-2 domains appearing adjacent to one another in a repetitive pattern. Further, we have identified a non-pentapeptide motif associated with rapid mycobacterial evolution. The sequence composition of this region suggests a different structure compared to pentapeptide-2 repeats. By studying the evolution of the PE/PPE_MPTR proteins, we have distinguished features pertaining to tuberculosis-inducing species. Further studies of the non-pentapeptide region associated with repeat expansions promises to shed light on the pathogenicity of Mycobacterium tuberculosis.

  • 1030.
    Webling, Kristin E
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Design, Synthesis and Characterization of Galanin Receptor Selective Ligands2017Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Galanin is a 29/30 amino acid long bioactive peptide discovered over 30 years ago when C-terminally amidated peptides were isolated from porcine intestines. The name galanin originates from a combination of the first and last amino acids - G from glycine and the rest from alanine. The first 15 amino acids are highly conserved throughout species, which indicates that the N-terminus is important for receptor recognition and binding. Galanin exerts its effects by binding to three different G protein-coupled receptors, which all differ according to regional distribution, the affinity for shortened galanin fragments, as well as the intracellular G-protein signaling cascade used. When first discovered, galanin was found to cause muscle contraction as well as hyperglycemia.  Over the years, galanin has been reported to be involved in a wide variety of biological functions, for example food intake and neurogenesis, and pathological functions, for example epilepsy and depression.

    Determining the specific involvement of the three different galanin receptors in biological and pathological processes is limited by the small amount of galanin receptor selective/specific ligands available as research tools. Furthermore, the fast degradation of peptides limits the administration routes in animal studies.

    This thesis aims at developing new galanin receptor-selective ligands to help delineate the involvement of the three different galanin receptors.

    Paper 1 presents the shortest galanin fragment with a galanin receptor 2 specific binding preference where only a single amino acid substitution was made, Ala5Ser in galanin (2-11). In addition, G-protein coupled receptor signaling were evaluated through both a classical second messenger assay and a real-time label-free technique in cells overexpressing the receptor as well as low receptor expression.

    Paper 2 demonstrates that the neuroprotective effects of galanin in a kainic acid-induced excitotoxic animal model were mediated through galanin receptor 1. Furthermore, a new robust protocol for evaluating G-protein signaling using a label-free real time impedance technique was presented and compared to two different classical second-messenger assays.

    Paper 3 presents a series of systemically active galanin receptor 2 selective ligands subsequently evaluated in two different depression-like animal models.

    Paper 4 investigates a mutated form of human galanin which was found in epilepsy patients and binding and signaling properties of the mutated associated ligand p.(A39E) was examined.

    In conclusion, this thesis presents the discovery of eight new galanin ligands, which can be used to evaluate the galaninergic system as well as to help investigate the possible use of peptides as pharmaceuticals in different diseases.

  • 1031.
    Webling, Kristin
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Groves-Chapman, Jessica L.
    Runesson, Johan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Saar, Indrek
    Lang, Andreas
    Sillard, Rannar
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Jakovenko, Erik
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Kofler, Barbara
    Holmes, Philip V.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    Pharmacological stimulation of GAL1R but not GAL2R attenuates kainic acid-induced neuronal cell death in the rat hippocampus2016Ingår i: Neuropeptides, ISSN 0143-4179, E-ISSN 1532-2785, Vol. 58, s. 83-92Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The neuropeptide galanin is widely distributed in the central and peripheral nervous systems and part of a bigger family of bioactive peptides. Galanin exerts its biological activity through three G-protein coupled receptor subtypes, GAL1–3R. Throughout the last 20 years, data has accumulated that galanin can have a neuroprotective effect presumably mediated through the activation of GAL1R and GAL2R. In order to test the pharmaceutical potential of galanin receptor subtype selective ligands to inhibit excitotoxic cell death, the GAL1R selective ligand M617 and the GAL2R selective ligand M1145 were compared to the novel GAL1/2R ligand M1154, in their ability to reduce the excitotoxic effects of intracerebroventricular injected kainate acid in rats.

    The peptide ligands were evaluated in vitro for their binding preference in a competitive 125I-galanin receptor subtype binding assay, and G-protein signaling was evaluated using both classical signaling and a label-free real-time technique. Even though there was no significant difference in the time course or severity of the kainic acid induced epileptic behavior in vivo, administration of either M617 or M1154 before kainic acid administration significantly attenuated the neuronal cell death in the hippocampus. Our results indicate the potential therapeutic value of agonists selective for GAL1R in the prevention of neuronal cell death. 

  • 1032.
    Webling, Kristin
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Runesson, Johan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lang, Andreas
    Saar, Indrek
    Kofler, Barbara
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    Ala(5)-galanin (2-11) is a GAL2R specific galanin analogue2016Ingår i: Neuropeptides, ISSN 0143-4179, E-ISSN 1532-2785, Vol. 60, s. 75-82Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    It is over 30years since the regulatory peptide galanin was discovered by Professor Mutt and co-workers. Galanin exerts its effects by binding to three galanin G-protein coupled receptors, namely GAL1R, GAL2R and GAL3R. Each galanin receptor has a different distribution in the central nervous system and the peripheral nervous system as well as distinctive signaling pathways, which implicates that the receptors are involved in different biological- and pathological effects. The delineation of the galaninergic system is however difficult due to a lack of stable, specific galanin receptor ligands. Herein, a new short GAL2R specific ligand, Ala(5)-galanin (2-11), is presented. The galanin (2-11) modified analogue Ala(5)-galanin (2-11) was tested in (125)I-galanin competitive binding studies for the three galanin receptors and the G-protein coupled receptor signaling properties was tested by the ability to influence second-messenger molecules like inositol phosphate and cyclic adenosine monophosphate. In addition, two different label-free real-time assays, namely EnSpire® based on an optical biosensor and xCELLigence® based on an electric biosensor, were used for evaluating the signaling properties using cell lines with different levels of receptor expression. Ala(5)-galanin (2-11) was subsequently found to be a full agonist for GAL2R with more than 375-fold preference for GAL2R compared to both GAL1R and GAL3R. The single amino acid substitution of serine to alanine at position 5 in the short ligand galanin (2-11) resulted in a ligand subsequently unable to bind neither GAL3R nor GAL1R, even at concentrations as high as 0.1mM.

  • 1033. Weibrecht, Irene
    et al.
    Lundin, Elin
    Kiflemariam, Sara
    Mignardi, Marco
    Grundberg, Ida
    Larsson, Chatarina
    Koos, Björn
    Nilsson, Mats
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab). Uppsala universitet.
    Söderberg, Ola
    In situ detection of individual mRNA molecules and protein complexes or post-translational modifications using padlock probes combined with the in situ proximity ligation assay2013Ingår i: Nature Protocols, ISSN 1754-2189, E-ISSN 1750-2799, Vol. 8, nr 2, s. 355-372Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Analysis at the single-cell level is essential for the understanding of cellular responses in heterogeneous cell populations, but it has been difficult to perform because of the strict requirements put on detection methods with regard to selectivity and sensitivity (i.e., owing to the cross-reactivity of probes and limited signal amplification). Here we describe a 1.5-d protocol for enumerating and genotyping mRNA molecules in situ while simultaneously obtaining information on protein interactions or post-translational modifications; this is achieved by combining padlock probes with in situ proximity ligation assays (in situ PLA). In addition, we provide an example of how to design padlock probes and how to optimize staining conditions for fixed cells and tissue sections. Both padlock probes and in situ PLA provide the ability to directly visualize single molecules by standard microscopy in fixed cells or tissue sections, and these methods may thus be valuable for both research and diagnostic purposes.

  • 1034.
    Weidner, Jessica M.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Barragan, Antonio
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Tightly regulated migratory subversion of immune cells promotes the dissemination of Toxoplasma gondii2014Ingår i: International Journal of Parasitology, ISSN 0020-7519, E-ISSN 1879-0135, Vol. 44, nr 2, s. 85-90Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    While the spread of Toxoplasma gondii within the infected human or animal host is associated with pathology, the pathways of dissemination have remained enigmatic. From the time point of entry into the gut, to the quiescent chronic infection in the central nervous system, Toxoplasma is detected and surveyed by immune cells that populate the tissues, for example dendritic cells. Paradoxically, this protective migratory function of leukocytes appears to be targeted by Toxoplasma to mediate its dissemination in the organism. Recent findings show that tightly regulated events take place shortly after host cell invasion that promote the migratory activation of infected dendritic cells. Here, we review the emerging knowledge on how this obligate intracellular protozoan orchestrates the subversion of leukocytes to achieve systemic dissemination and reach peripheral organs where pathology manifests.

  • 1035.
    Westberg, Emelie
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för material- och miljökemi (MMK).
    Hedebrant, Ulla
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för material- och miljökemi (MMK).
    Haglund, Johanna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för material- och miljökemi (MMK).
    Alsberg, Tomas
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för tillämpad miljövetenskap (ITM).
    Eriksson, Johan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för material- och miljökemi (MMK).
    Seidel, Albrecht
    Törnqvist, Margareta
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för material- och miljökemi (MMK).
    Conditions for sample preparation and quantitative HPLC/MS-MS analysis of bulky adducts to serum albumin with diolepoxides of polycyclic aromatic hydrocarbons as models2014Ingår i: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 406, nr 5, s. 1519-1530Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Stable adducts to serum albumin (SA) from electrophilic and genotoxic compounds/metabolites can be used as biomarkers for quantification of the corresponding in vivo dose. In the present study, conditions for specific analysis of stable adducts to SA formed from carcinogenic polycyclic aromatic hydrocarbons (PAH) were evaluated in order to achieve a sensitive and reproducible quantitative method. Bulky adducts from diolepoxides (DE) of PAH, primarily DE of benzo[a]pyrene (BPDE) and also DE of dibenzo[a,l]pyrene (DBPDE) and dibenzo[a,h]anthracene (DBADE), were used as model compounds. The alkylated peptides obtained after enzymatic hydrolysis of human SA modified with the different PAHDE were principally PAHDE-His-Pro, PAHDE-His-Pro-Tyr and PAHDE-Lys. Alkaline hydrolysis under optimised conditions gave the BPDE-His as the single analyte of alkylated His, but also indicated degradation of this adduct. It was not possible to obtain the BPDE-His as one analyte from BPDE-alkylated SA through modifications of the enzymatic hydrolysis. The BPDE-His adduct was shown to be stable during the weak acidic conditions used in the isolation of SA. Enrichment by HPLC or SPE, but not butanol extraction, gave good recovery, using Protein LoBind tubes. A simple internal standard (IS) approach using SA modified with other PAHDE as IS was shown to be applicable. A robust analytical procedure based on digestion with pronase, enrichment by HPLC or SPE, and analysis with HPLC/MS-MS electrospray ionisation was achieved. A good reproducibility (coefficient of variation (CV) 11 %) was obtained, and the achieved limit of detection for the studied PAHDE, using standard instrumentation, was approximately 1 fmol adduct/mg SA analysing extract from 5 mg SA.

  • 1036.
    Westerlund, Kristina
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Moran, Sean D.
    Privett, Heidi K.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Hay, Sam
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Jarvet, Juri
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Gibney, Brian R.
    Tommos, Cecilia
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Making a single-chain four-helix bundle for redox chemistry studies2008Ingår i: Protein Engineering Design & Selection, ISSN 1741-0126, E-ISSN 1741-0134, Vol. 21, nr 11, s. 645-652Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The construction and characteristics of the stable and well-structured alpha W-4 protein are described. The 117-residue, single-chain protein has a molecular weight of 13.1 kDa and is designed to fold into a four-helix bundle. Experimental characterization of the expressed and purified protein shows a 69.8 +/- 0.8% helical content over a 5.5-10.0 pH range. The protein is thermostable with a T-M > 355 K and has a free energy of unfolding as measured by chemical denaturation of -4.7 kcal mol(-1) at 25 degrees C and neutral pH. One-dimensional (1D) proton and 2D N-15-HSQC spectra show narrow, well-dispersed spectral lines consistent with a uniquely structured alpha-helical protein. Analytical ultracentrifugation and NMR data show that the protein is monomeric over a broad protein concentration range. The 324 nm emission maximum of the unique Trp-106 is consistent with a sequestered position of the aromatic residue. Additionally, differential pulse voltammetry characterization indicates an elevated peak potential for Trp-106 when the protein is folded (pH range 7.0-8.5) relative to partly unfolded (pH range 11.4-13.2). The oxidation of Trp-106 is coupled to proton release as shown by a 53 +/- 3 mV/pH unit dependence of the peak potential over the 7.0-8.5 pH range.

  • 1037.
    Westman, Jacob
    Stockholms universitet, Naturvetenskapliga fakulteten.
    Synthesis of oligosaccharides related to heparin and heparan sulphate and their binding to fibroblast growth factors1995Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
  • 1038. Whitelam, Stephen
    et al.
    Geissler, Phillip L.
    Pronk, Sander
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Microscopic implications of S-DNA2010Ingår i: Physical review. E, ISSN 1539-3755, Vol. 82, nr 2, s. 21907-Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Recent experiments [J. van Mameren et al., Proc. Natl. Acad. Sci. U.S.A. 106, 18231 (2009)] provide a detailed spatial picture of overstretched DNA, showing that under certain conditions the two strands of the double helix separate at about 65 pN. It was proposed that this observation rules out the existence of an elongated, hybridized form of DNA (S-DNA). Here, we argue that the S-DNA picture is consistent with the observation of unpeeling during overstretching. We demonstrate that assuming the existence of S-DNA does not imply DNA overstretching to consist of the complete or near-complete conversion of the molecule from B to S form. Instead, this assumption implies in general a more complex dynamic coexistence of hybridized and unhybridized forms of DNA. We argue that such coexistence can rationalize several recent experimental observations.

  • 1039. Whittle, Andrew J.
    et al.
    Carobbio, Stefania
    Martins, Luis
    Slawik, Marc
    Hondares, Elayne
    Jesus Vazquez, Maria
    Morgan, Donald
    Csikasz, Robert I.
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut, Avdelningen för fysiologi.
    Gallego, Rosalia
    Rodriguez-Cuenca, Sergio
    Dale, Martin
    Virtue, Samuel
    Villarroya, Francesc
    Cannon, Barbara
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut, Avdelningen för fysiologi.
    Rahmouni, Kamal
    Lopez, Miguel
    Vidal-Puig, Antonio
    BMP8B Increases Brown Adipose Tissue Thermogenesis through Both Central and Peripheral Actions2012Ingår i: Cell, ISSN 0092-8674, E-ISSN 1097-4172, Vol. 149, nr 4, s. 871-885Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Thermogenesis in brown adipose tissue (BAT) is fundamental to energy balance and is also relevant for humans. Bone morphogenetic proteins (BMPs) regulate adipogenesis, and, here, we describe a role for BMP8B in the direct regulation of thermogenesis. BMP8B is induced by nutritional and thermogenic factors in mature BAT, increasing the response to noradrenaline through enhanced p38MAPK/CREB signaling and increased lipase activity. Bmp8b(-/-) mice exhibit impaired thermogenesis and reduced metabolic rate, causing weight gain despite hypophagia. BMP8B is also expressed in the hypothalamus, and Bmp8b(-/-) mice display altered neuropeptide levels and reduced phosphorylation of AMP-activated protein kinase (AMPK), indicating an anorexigenic state. Central BMP8B treatment increased sympathetic activation of BAT, dependent on the status of AMPK in key hypothalamic nuclei. Our results indicate that BMP8B is a thermogenic protein that regulates energy balance in partnership with hypothalamic AMPK. BMP8B may offer a mechanism to specifically increase energy dissipation by BAT.

  • 1040. Wickstrom, David
    et al.
    Wagner, Samuel
    Baars, Louise
    Ytterberg, A. Jimmy
    Klepsch, Mirjam
    van Wijk, Klaas J.
    Luirink, Joen
    de Gier, Jan-Willem
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Consequences of Depletion of the Signal Recognition Particle in Escherichia coli2011Ingår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 286, nr 6, s. 4598-4609Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Thus far, the role of the Escherichia coli signal recognition particle (SRP) has only been studied using targeted approaches. It has been shown for a handful of cytoplasmic membrane proteins that their insertion into the cytoplasmic membrane is at least partially SRP-dependent. Furthermore, it has been proposed that the SRP plays a role in preventing toxic accumulation of mistargeted cytoplasmic membrane proteins in the cytoplasm. To complement the targeted studies on SRP, we have studied the consequences of the depletion of the SRP component Fifty-four homologue (Ffh) in E. coli using a global approach. The steady-state proteomes and the proteome dynamics were evaluated using one- and two-dimensional gel analysis, followed by mass spectrometry-based protein identification and immunoblotting. Our analysis showed that depletion of Ffh led to the following: (i) impaired kinetics of the biogenesis of the cytoplasmic membrane proteome; (ii) lowered steady-state levels of the respiratory complexes NADH dehydrogenase, succinate dehydrogenase, and cytochrome bo(3) oxidase and lowered oxygen consumption rates; (iii) increased levels of the chaperones DnaK and GroEL at the cytoplasmic membrane; (iv) a sigma(32) stress response and protein aggregation in the cytoplasm; and (v) impaired protein synthesis. Our study shows that in E. coli SRP-mediated protein targeting is directly linked to maintaining protein homeostasis and the general fitness of the cell.

  • 1041.
    Wieslander, Lars
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Björk, Petra
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Nucleocytoplasmic mRNP export is an integral part of mRNP biogenesis2011Ingår i: Chromosoma, ISSN 0009-5915, E-ISSN 1432-0886, Vol. 120, nr 1, s. 23-38Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Nucleocytoplasmic export and biogenesis of mRNPs are closely coupled. At the gene, concomitant with synthesis of the pre-mRNA, the transcription machinery, hnRNP proteins, processing, quality control and export machineries cooperate to release processed and export competent mRNPs. After diffusion through the interchromatin space, the mRNPs are translocated through the nuclear pore complex and released into the cytoplasm. At the nuclear pore complex, defined compositional and conformational changes are triggered, but specific cotranscriptionally added components are retained in the mRNP and subsequently influence the cytoplasmic fate of the mRNP. Processes taking place at the gene locus and at the nuclear pore complex are crucial for integrating export as an essential part of gene expression. Spatial, temporal and structural aspects of these events have been highlighted in analyses of the Balbiani ring genes

  • 1042.
    Wigerius, Michael
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Roles of mammalian Scribble in polarity signaling, virus offense and cell-fate determination2010Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Mammalian Scribble is a target for proteins encoded by human papilloma virus, retro- and flaviviruses. Tick-borne encephalitis virus (TBEV) is a flavivirus that have evolved distinct strategies to escape antiviral responses. Information of how flaviviruses intrude on cell integrity comes from understanding of the roles that host-factors play when they interfere with viruses. The first part of this thesis describes a novel interaction between the TBEVNS5 protein and Scribble. The importance of the interaction was demonstrated by RNAi-mediated depletion of Scribble, which prevented suppression of JAK-STAT signaling by NS5. Together, these results define Scribble as a novel target for NS5.

    TBEV is known to cause central nervous system disease TBE in humans that can lead to cognitive dysfunction. A unifying theme in CNS related diseases are defects in neuronal extensions. We therefore addressed the effects of TBEV expression in PC12 cell differentiation, which is characterized by extensive neurite growth. Our data show that TBEVNS5 suppresses neurite outgrowth through the Rho GTPase Rac1. These findings provide evidence that Rac1 is an indirect target of NS5 in neurite inhibition. Scribble was recently implicated in spine morphogenesis. Thus, we tested the role of Scribble in neurite elongation. Depletion of Scribble in PC12 cells, reduced neurite density but increased length of those remaining. Moreover, Scribble bound components in the Ras/ERK cascade in a growth factor dependent manner. Together, these results demonstrate that Scribble controls neurite elongation by scaffolding MAPK components. Moreover, as loss of dendritic spines, actin-rich protrusions on neurons, is a feature in cognitive dysfunction we speculate that cognitive dysfunction in TBE might involve disturbed Scribble expression by NS5.

    We also investigated the binding between NS1 of Influenza A virus and Scribble. The PDZ domains of Scribble are usually selective for specific C-terminal motifs in proteins. Because NS1 has a canonical PDZ motif we tested if binding to Scribble depends on this motif. We found that Scribble binds NS1; the association is dependent on the NS1 C-terminus that is recognized by PDZ3-4 of Scribble. Together, these results suggest that Scribble is a target for the H5N1 NS1 protein 

  • 1043. Wiklund, Magda-Lena
    et al.
    Steinert, Stefanie
    Junell, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Hultmark, Dan
    Stoven, Svenja
    The N-terminal half of the Drosophila Rel/NF-kappa B factor Relish, REL-68, constitutively activates transcription of specific Relish target genes2009Ingår i: Developmental and Comparative Immunology, ISSN 0145-305X, E-ISSN 1879-0089, Vol. 33, nr 5, s. 690-696Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The Rel/NF-kappa B transcription factor Relish is a major regulator of the antimicrobial response in Drosophila. Upon immune challenge, Relish is cleaved to generate two fragments, the DNA-binding transcription factor REL-68 and the I kappa B-like REL-49. Using transgenic fly strains we show here that overexpression of REL-68 separately from REL-49 is sufficient to activate strong constitutive transcription of the Diptericin gene, but little constitutive or inducible transcription of Attacin and Cecropin, two other Relish target genes. Their transcription may therefore require additional modifications of Relish. However, phosphorylation of the conserved serine residue S431 is not involved in such modifications. This is unlike p65 and Dorsal, which are modulated by phosphorylation at their homologous site. In contrast to other I kappa B proteins, overexpression of REL-49 had no inhibitory effect on Relish-dependent transcription. Instead, we propose that the C-terminal I kappa B-like domain executes a scaffolding and recruiting function for full activation of Relish.

  • 1044.
    Wikström, Malin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Synthesis and protein curing abilities of membrane glycolipids2006Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    There are many types of membrane lipids throughout Nature. Still little is known about synthesizing pathways and how different lipids affect the embedded membrane proteins. The most common lipids are glycolipids since they dominate plant green tissue. Glycolipids also exist in mammal cells as well as in most Gram-positive bacteria. Glycosyltransferases (GTs) catalyze the final enzymatic steps for these glycolipids. In the bacteria Acholeplasma laidlawii and Streptococcus pneumonie and in the plant Arabidopsis thaliana, GTs for mono-/di-glycosyl-diacylglycerol (-DAG) are suggested to be regulated to keep a certain membrane curvature close to a bilayer/nonbilayer phase transition. The monoglycosylDAGs are nonbilayer-prone with small headgroups, hence by themselves they will not form bilayer structures.

    Here we have determined the genes encoding the main glycolipids of A. laidlawii and S. pneumonie. We have also shown that these GTs belong to a large enzyme group widely spread in Nature, and that all four enzymes are differently regulated by membrane lipids. The importance of different lipid properties were traced in a lipid mutant of Escherichia coli lacking the major (75 %), nonbilayer-prone/zwitterionic, lipid phosphatidylethanolamine. Introducing the genes for the GTs of A. laidlawii and two analogous genes from A. thaliana yielded new strains containing 50 percent of glyco-DAG lipids. The monoglyco-DAG strains contain significant amounts of nonbilayer-prone lipids while the diglyco-DAG strains contain no such lipids. Comparing these new strains for viability and the state of membrane-associated functions made it possible to connect different functions to certain lipid properties. In summary, a low surface charge density of anionic lipids is important in E.coli membranes, but this fails to be supportive if the diluting species have a too large headgroup. This indicates that a certain magnitude of the curvature stress is crucial for the membrane bilayer in vivo.

  • 1045.
    Wikström, Malin
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Xie, J.
    Bogdanov, M.
    Mileyovskaya, E.
    Heacock, P.
    Wieslander, Åke
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Dowhan, W.
    Monoglucosyldiacylglycerol, a foreign lipid, can substitute for phosphatidylethanolamine in essential membrane-associated functions in Escherichia coli2004Ingår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 279, nr 11, s. 10484-10493Artikel i tidskrift (Refereegranskat)
  • 1046.
    Wilhelmsson, Christine
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Proteomics of the Drosophila hemolymph clot and the function of transglutaminase2009Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Insects rely on a fast and effective coagulation and wound response to avoid loss of body fluids and immobilize pathogens. Arthropod coagulation is in some respect equivalent to vertebrate coagulation but most factors and the regulation of coagulation systems seem not to be phylogenetically conserved. To get a more complete picture of insect clotting we studied the molecular and functional nature of Drosophila hemolymph coagulation.

    We developed new proteomic methods to collect Drosophila clotting factors. Several candidate factors were identified, including both predicted and novel clot proteins. Five putative TG (transglutaminase) substrates were found and we could also demonstrate that the clot is involved in immobilization of bacteria. Further investigating the role of TG we found TG to be important for Drosophila coagulation and that Fondue is a major substrate of the enzyme. Using fon RNAi knockdown we showed that Fondue affects the physical properties of the clot. A fon-GFP fusion construct was generated to follow its expression. The cuticle and the clot were labelled suggesting that Fondue is incorporated into both cuticle and clot. Clot properties and composition were affected by inhibiting TG chemically (MDC) and genetically (RNAi). Moreover, interaction between Fondue and Eig71Ee was demonstrated. Previous results indicated that coagulation could have an immune function. In hemolymph preparations, containing selected microorganisms, small deposits were seen on the microbial surfaces. The contents of these were investigated, revealing the presence of procoagulants. The targeting of microbes is instant and depends on TG and its substrates. Entomopathogenic nematode infections were performed to validate the functional importance of TG. TG RNAi knockdown larvae showed increased mortality, supporting an immune function for TG. Altogether, our data provide a more comprehensive picture of Drosophila immunity, and may further improve the understanding of innate immunity in general.

  • 1047.
    Wincent, Emma
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    6-Formylindolo[3,2-b]carbazole (FICZ) metabolism2007Konferensbidrag (Övrig (populärvetenskap, debatt, mm))
  • 1048.
    Wincent, Emma
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Bengtsson, Johanna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Bardbori, Afshin Mohammadi
    Alsberg, Tomas
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för tillämpad miljövetenskap (ITM).
    Luecke, Sandra
    Rannug, Ulf
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Rannug, Agneta
    Inhibition of cytochrome P4501-dependent clearance of the endogenous agonist FICZ as a mechanism for activation of the aryl hydrocarbon receptor2012Ingår i: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 109, nr 12, s. 4479-4484Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Altered systemic levels of 6-formylindolo[3,2-b]carbazole (FICZ), an enigmatic endogenous ligand for the aryl hydrocarbon receptor (AHR), may explain adverse physiological responses evoked by small natural and anthropogenic molecules as well as by oxidative stress and light. We demonstrate here that several different chemical compounds can inhibit the metabolism of FICZ, thereby disrupting the autoregulatory feedback control of cytochrome P4501 systems and other proteins whose expression is regulated by AHR. FICZ is both the most tightly bound endogenous agonist for the AHR and an ideal substrate for cytochrome CYP1A1/1A2 and 1B1, thereby also participating in an autoregulatory loop that keeps its own steady-state concentration low. At very low concentrations FICZ influences circadian rhythms, responses to UV light, homeostasis associated with pro-and anti-inflammatory processes, and genomic stability. Here, we demonstrate that, if its metabolic clearance is compromised, femtomolar background levels of this compound in cell-culture medium are sufficient to up-regulate CYP1A1 mRNA and enzyme activity. The oxidants UVB irradiation and hydrogen peroxide and the model AHR antagonist 3'-methoxy-4'-nitroflavone all inhibited induction of CYP1A1 enzyme activity by FICZ or 2,3,7,8-tetrachlorodibenzo-p-dioxin, thereby subsequently elevating intracellular levels of FICZ and activating AHR. Taken together, these findings support an indirect mechanism of AHR activation, indicating that AHR activation by molecules with low affinity actually may reflect inhibition of FICZ metabolism and raising questions about the reported promiscuity of the AHR. Accordingly, we propose that prolonged induction of AHR activity through inhibition of CYP1 disturbs feedback regulation of FICZ levels, with potential detrimental consequences.

  • 1049. Witkowska, Ewa
    et al.
    Nowakowski, Michal
    Oleszczuk, Marta
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Filip, Katarzyna
    Ciszewska, Malgorzata
    Chung, Nga N.
    Schiller, Peter W.
    Wojcik, Jacek
    Izdebski, Jan
    Ureido group containing cyclic dermorphin(1-7) analogues: synthesis, biology and conformation2007Ingår i: Journal of Peptide Science, ISSN 1075-2617, E-ISSN 1099-1387, Vol. 13, nr 8, s. 519-528Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Six cyclic peptides related to dermorphin(1-7) have been synthesized. The synthesis of linear peptides containing diamino acid residues in positions 2 and 4 was carried out on a 4-methylbenzhydrylamine resin, and cyclization was achieved by treatment with bis-(4-nitrophenyl)carbonate to form a urea unit. The peptides were tested in the guinea-pig ileum (GPI) and mouse vas deferens (MVD) assays. Diverse opioid agonist activities were observed, depending on the size of the ring. The results were compared with those obtained earlier for 1-4 dermorphin analogues. The conformations of all six dermorphin analogues were studied. The conformational space of the peptides was examined using the electrostatically driven Monte Carlo method. On the basis of NMR data, an ensemble of conformations was obtained for each peptide. The opioid activity profiles of the compounds are discussed in the light of the structural data.

  • 1050. Wojcik, Anna
    et al.
    Broclawik, Ewa
    Siegbahn, Per E. M.
    Stockholms universitet, Naturvetenskapliga fakulteten, Fysikum.
    Borowski, Tomasz
    Mechanism of Benzylic Hydroxylation by 4-Hydroxymandelate Synthase: A Computational Study2012Ingår i: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 51, nr 47, s. 9570-9580Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Hydroxymandelate synthase (HMS) and 4-hydroxyphenylpyruvate dioxygenase (HPPD) are highly related enzymes using the same substrates but catalyzing hydroxylation reactions yielding different products. The first The first steps of the HMS and I-IPPD catalytic reactions are believed to proceed in the same way and lead to an Fe(IV)=O-hydroxyphenylacetate (HPA) intermediate. Further, down the, catalytic cycles, HMS uses Fe(IV)=O to perform hydroxylation of the benzylic carbon, Whereas in HPPD, the reactive oxoferryl intermediate attacks the aromatic ring of HPA. This study focuses on this part of the HMS catalytic cycle that starts from the oxoferryl intermediate and aims to identify interactions within the active site that are responsible for enzyme specificity. To this end, a HMS-Fe(IV)=O-HPA complex was modeled with molecular dynamics simulations On the basis. of the molecular: dynamics. equilibrated structure, active site model suitable for quantum chemical Investigations was constructed and used for density functional theory. (B3LYP) Calculations of the mechanism of the native reaction of HMS, i.e., benzylic hydroxylation, and the alternative electrophilic attack on the ring, which is a step Of the HPPD catalytic cycle: The most important, result of this study is the finding that the conformation of the Ser201 side chain in the second coordination shell has a key role in directing the of Fe(IV)=O. into either the HMS or the HPPD channel

1819202122 1001 - 1050 av 1087
RefereraExporteraLänk till träfflistan
Permanent länk
Referera
Referensformat
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Annat format
Fler format
Språk
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Annat språk
Fler språk
Utmatningsformat
  • html
  • text
  • asciidoc
  • rtf