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  • 201.
    Refaat, Doaa
    et al.
    Agr Res Ctr, Egypt;Beni Suef Univ, Egypt.
    Aggour, Mohamed G.
    Agr Res Ctr, Egypt.
    Farghali, Ahmed A.
    Beni Suef Univ, Egypt.
    Mahajan, Rashmi
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Wiklander, Jesper G.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Nicholls, Ian A.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Piletsky, Sergey A.
    Univ Leicester, UK.
    Strategies for Molecular Imprinting and the Evolution of MIP Nanoparticles as Plastic Antibodies-Synthesis and Applications2019In: International Journal of Molecular Sciences, ISSN 1422-0067, E-ISSN 1422-0067, Vol. 20, no 24, p. 1-21, article id 6304Article, review/survey (Refereed)
    Abstract [en]

    Materials that can mimic the molecular recognition-based functions found in biology are a significant goal for science and technology. Molecular imprinting is a technology that addresses this challenge by providing polymeric materials with antibody-like recognition characteristics. Recently, significant progress has been achieved in solving many of the practical problems traditionally associated with molecularly imprinted polymers (MIPs), such as difficulties with imprinting of proteins, poor compatibility with aqueous environments, template leakage, and the presence of heterogeneous populations of binding sites in the polymers that contribute to high levels of non-specific binding. This success is closely related to the technology-driven shift in MIP research from traditional bulk polymer formats into the nanomaterial domain. The aim of this article is to throw light on recent developments in this field and to present a critical discussion of the current state of molecular imprinting and its potential in real world applications.

  • 202.
    Ren, Yansong
    et al.
    KTH Royal Instute of Technology, Sweden.
    Hu, Lei
    KTH Royal Instute of Technology, Sweden;Jiangsu Univ, Peoples Republi of China.
    Ramström, Olof
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. KTH Royal Instute of Technology, Sweden;Univ Massachusetts, USA.
    Multienzymatic cascade synthesis of an enantiopure (2R,5R)-1,3-oxathiolane anti-HIV agent precursor2019In: Molecular Catalysis, ISSN 2468-8274, Vol. 468, p. 52-56Article in journal (Refereed)
    Abstract [en]

    An enantiopure (2R,5R)-1,3-oxathiolane was obtained using a multienzymatic cascade protocol. By employing a combination of surfactant-treated subtilisin Carlsberg and Candida antarctica lipase B, the absolute configuration of the resulting 1,3-oxathiolane ring was efficiently controlled, resulting in an excellent enantiomeric excess (> 99%). This enantiopure 1,3-oxathiolane derivative is a key precursor to anti-HIV agents, such as lamivudine, through subsequent N-glycosylation.

  • 203.
    Ren, Yansong
    et al.
    Royal Institute of Technology.
    Xie, Sheng
    Royal Institute of Technology;Hunan Univ, Peoples Republic of China.
    Grape, Erik Svensson
    Stockholm University.
    Inge, A. Ken
    Stockholm University.
    Ramström, Olof
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Royal Institute of Technology;Univ Massachusetts Lowell, USA.
    Multistimuli-Responsive Enaminitrile Molecular Switches Displaying H+-Induced Aggregate Emission, Metal Ion-Induced Turn-On Fluorescence, and Organogelation Properties2018In: Journal of the American Chemical Society, ISSN 0002-7863, E-ISSN 1520-5126, Vol. 140, no 42, p. 13640-13643Article in journal (Refereed)
    Abstract [en]

    Multistimuli-responsive enaminitrile-based configurational switches displaying aggregation-induced emission (AIE), fluorescence turn-on effects, and super gelation properties are presented. The E-isomers dominated (>97%) in neutral/basic solution, and the structures underwent precisely controlled switching around the enamine C=C bond upon addition of acid/base. Specific fluorescence output was observed in response to different external input in the solution and solid states. In response to H+, configurational switching resulted in complete formation of the nonemissive Z-H+-isomers in solution, however displaying deep-blue to blue fluorescence (Phi(F) up to 0.41) in the solid state. In response to Cu-II in the solution state, the E-isomers exhibited intense, turn-on, blue-green fluorescence, which could be turned off by addition of competitive coordination. The acid/base-activated switching, together with the induced AIE-effects, further enabled the accomplishment of a responsive superorganogelator. In nonpolar solvents, a blue-fluorescent supramolecular gel was formed upon addition of acid to the E-isomer suspension. The gelation could be reversed by addition of base, and the overall, reversible process could be repeated at least five cycles.

  • 204.
    Richter, Florian
    et al.
    University of Washington, USA.
    Leaver-Fay, Andrew
    University of North Carolina, USA.
    Khare, Sagar D
    University of Washington, USA.
    Bjelic, Sinisa
    University of Washington, USA.
    Baker, David
    University of Washington, USA.
    De novo enzyme design using Rosetta32011In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 6, no 5Article in journal (Refereed)
    Abstract [en]

    The Rosetta de novo enzyme design protocol has been used to design enzyme catalysts for a variety of chemical reactions, and in principle can be applied to any arbitrary chemical reaction of interest. The process has four stages: 1) choice of a catalytic mechanism and corresponding minimal model active site, 2) identification of sites in a set of scaffold proteins where this minimal active site can be realized, 3) optimization of the identities of the surrounding residues for stabilizing interactions with the transition state and primary catalytic residues, and 4) evaluation and ranking the resulting designed sequences. Stages two through four of this process can be carried out with the Rosetta package, while stage one needs to be done externally. Here, we demonstrate how to carry out the Rosetta enzyme design protocol from start to end in detail using for illustration the triosephosphate isomerase reaction.

  • 205.
    Roman, Pawel
    et al.
    Wageningen Univ, Netherlands;Wetsus, Netherlands.
    Klok, Johannes B. M.
    Wetsus, Netherlands;Paqell, Netherlands.
    Sousa, João A. B.
    Wageningen Univ, Netherlands;Wetsus, Netherlands.
    Broman, Elias
    Linnaeus University, Faculty of Health and Life Sciences, Department of Biology and Environmental Science.
    Dopson, Mark
    Linnaeus University, Faculty of Health and Life Sciences, Department of Biology and Environmental Science.
    Zessen, Erik Van
    Paques BV, Netherlands.
    Bijmans, Martijn F. M.
    Wetsus, Netherlands.
    Sorokin, Dimitry Y.
    Russian Acad Sci, Russia;Delft Univ Technol, Netherlands.
    Janssen, Albert J. H.
    Wageningen Univ, Netherlands;Shell Technol Ctr Bangalore, India.
    Selection and Application of Sulfide Oxidizing Microorganisms Able to Withstand Thiols in Gas Biodesulfurization Systems2016In: Environmental Science and Technology, ISSN 0013-936X, E-ISSN 1520-5851, Vol. 50, no 23, p. 12808-12815Article in journal (Refereed)
    Abstract [en]

    After the first commercial applications of a new biological process for the removal of hydrogen sulfide (H2S) from low pressure biogas, the need arose to broaden the operating window to also enable the removal of organosulfur compounds from high pressure sour gases. In this study we have selected microorganisms from a full-scale biodesulfurization system that are capable of withstanding the presence of thiols. This full-scale unit has been in stable operation for more than 10 years. We investigated the microbial community by using high-throughput sequencing of 16S rRNA gene amplicons which showed that methanethiol gave a competitive advantage to bacteria belonging to the genera Thioalkalibacter (Halothiobacillaceae family) and Alkalilimnicola (Ectothiorhosdospiraceae family). The sulfide-oxidizing potential of the acclimatized population was investigated under elevated thiol loading rates (4.5–9.1 mM d–1), consisting of a mix of methanethiol, ethanethiol, and propanethiol. With this biomass, it was possible to achieve a stable bioreactor operation at which 80% of the supplied H2S (61 mM d–1) was biologically oxidized to elemental sulfur. The remainder was chemically produced thiosulfate. Moreover, we found that a conventionally applied method for controlling the oxygen supply to the bioreactor, that is, by maintaining a redox potential set-point value, appeared to be ineffective in the presence of thiols.

  • 206.
    Rosengren, Johan
    et al.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    McManus, Ailsa
    Craik, David
    The structural and functional diversity of naturally occurring antimicrobial peptides2002In: Current Medicinal Chemistry - Anti Infective Agents, Vol. 1, no 4, p. 319-341Article, review/survey (Other academic)
    Abstract [en]

    Antimicrobial peptides occur in a diverse range of organisms from microorganisms to insects, plants and animals. Although they all have the common function of inhibiting or killing invading microorganisms they achieve this function using an extremely diverse range of structural motifs. Their sizes range from approximately 10-90 amino acids. Most carry an overall positive charge, reflecting a preferred mode of electrostatic interaction with negatively charged microbial membranes. This article describes the structural diversity of a representative set of antimicrobial peptides divided into five structural classes: those with -helical structure, those with -sheet structure, those with mixed helical / - sheet structure, those with irregular structure, and those incorporating a macrocyclic structure. There is a significant diversity in both the size and charge of molecules within each of these classes and between the classes. The common feature of their three-dimensional structures is, however, that they have a degree of amphipathic character in which there is separate localisation of hydrophobic regions and positively charged regions. An emerging trend amongst antimicrobial proteins is the discovery of more macrocyclic analogues. Cyclisation appears to impart an additional degree of stability on these molecules and minimizes proteolytic cleavage. In conclusion, there appear to be a number of promising opportunities for the development of novel clinically useful antimicrobial peptides based on knowledge of the structures of naturally occurring antimicrobial molecules.

  • 207.
    Rosengren, K. Johan
    et al.
    The University of Queensland, Institute for Molecular Bioscience, Brisbane, QLD 4072, Australia.
    Craik, David
    The University of Queensland, Institute for Molecular Bioscience, Brisbane, QLD 4072, Australia.
    How bugs make lassos2009In: Chemistry and Biology, ISSN 1074-5521, E-ISSN 1879-1301, Vol. 16, no 12, p. 1211-1212Article in journal (Refereed)
  • 208.
    Ruiz-Pavon, Lorena
    et al.
    Division of Molecular Genetics, Department of Physics, Chemistry and Biology, Linköping University and School of Natural Sciences, Linnaeus university.
    Karlsson, Patrik M.
    Division of Molecular Genetics, Department of Physics, Chemistry and Biology, Linköping University .
    Carlsson, Jonas
    Division of Bioinformatics, Department of Physics, Chemistry and Biology, Linköping University .
    Samyn, Dieter R.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Persson, Bengt L.
    Division of Bioinformatics, Department of Physics, Chemistry and Biology, Linköping University .
    Persson, Bengt L.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Spetea, Cornelia
    Division of Molecular Genetics, Department of Physics, Chemistry and Biology, Linköping University .
    Functionally important amino acids in the Arabidopsis thylakoid phosphate transporter: Homology modeling and site-directed mutagenesis2010In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 49, no 30, p. 6430-6439Article in journal (Refereed)
    Abstract [en]

    The anion transporter 1 (ANTR1) from Arabidopsis thaliana, homologous to the mammalian members of the solute carrier 17 (SLC17) family, is located in the chloroplast thylakoid membrane. When expressed heterologously in Escherichia coli, ANTR1 mediates a Na+-dependent active transport of inorganic phosphate (Pi). The aim of this study was to identify amino acid residues involved in Pi binding and translocation by ANTR1 and in the Na+ dependence of its activity. A three-dimensional structural model of ANTR1 was constructed using the crystal structure of glycerol 3-phosphate/phosphate antiporter from E. coli as a template. Based on this model and multiple sequence alignments, five highly conserved residues in plant ANTRs and mammalian SLC17 homologues have been selected for site-directed mutagenesis, namely, Arg-120, Ser-124, and Arg-201 inside the putative translocation pathway and Arg-228 and Asp-382 exposed at the cytoplasmic surface of the protein. The activities of the wild-type and mutant proteins have been analyzed using expression in E. coli and radioactive Pi transport assays and compared with bacterial cells carrying an empty plasmid. The results from Pi- and Na+-dependent kinetics indicate the following: (i) Arg-120 and Arg-201 may be important for binding and translocation of the substrate; (ii) Ser-124 may function as a transient binding site for Na+ ions in close proximity to the periplasmic side; (iii) Arg-228 and Asp-382 may participate in interactions associated with protein conformational changes required for full transport activity. Functional characterization of ANTR1 should provide useful insights into the function of other plant and mammalian SLC17 homologous transporters.

  • 209. Rydström, J
    et al.
    Eytan, G D
    Eytan, E
    Persson, Bengt L.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Mitochondrial nicotinamide nucleotide transhydrogenase: A redox-driven single polypeptide proton pump1987In: DT Diaphorase: a quinone reductase with special functions in cell metabolism and detoxication; proceedings of an international conference held at the Arrhenius Laboratory, University of Stockholm, 1-4 June, 1986 / [ed] Lars Ernster, Cambridge: Cambridge University Press , 1987, p. 83-88Chapter in book (Other academic)
  • 210. Rydström, J
    et al.
    Persson, Bengt L.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Carlenor, E
    Transhydrogenase linked to pyridine nucleotides1987In: Pyridine Nucleotide Coenzymes: Chemical, Biochemical, and Medical Aspects / [ed] Dolphin, D, New York: John Wiley & Sons, 1987, p. 433-460Chapter in book (Other academic)
  • 211. Rydström, J
    et al.
    Persson, Bengt L.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Tang, H-L
    Mitochondrial nicotinamide nucleotide transhydrogenase1984In: Bioenergetics / [ed] L. Ernster, Amsterdam: Elsevier, 1984, p. 207-219Chapter in book (Other academic)
  • 212.
    Samyn, Dieter R.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Structure/function relationships of inorganic phosphate transporters2011Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Of the many nutrients that make the metabolic clock tick, inorganic phosphate fulfills an essential role in all yeast (and other organisms), being necessary for both structural and metabolic purposes. In order to transport phosphate into the cell interior, Saccharomyces cerevisiae makes use of two systems, comprising high- or low-affinity transporters, which are responsible for the cellular accumulation of inorganic phosphate. Depending on the extracellular concentration of inorganic phosphate, one of the two systems will be responsible for scavenging phosphate from the surroundings.The present thesis focuses on the high-affinity transport system in S. cerevisiae, i.e. Pho84, which is induced under low phosphate conditions. The expression and degradation of Pho84 is dependent on the availability of phosphate. When confronted with ample amounts of external phosphate, the Pho84 undergoes phosphorylation, prior to ubiquitylation. These events will eventually lead to the removal of Pho84 from the plasma membrane, followed by vacuolar degradation. The Pho84, together with the ANTR1 high-affinity inorganic phosphate transporter of Arabidopsis thaliana, are integral membrane proteins belonging to the major facilitator superfamily. Both are predicted to have 12 transmembrane helices, which have been confirmed by in silico modeling of both proteins, using the glycerol-3-phosphate transporter as template. The obtained models served as a rational start point for the study of the molecular transport mechanisms by means of site-directed mutagenesis and consequently functional and biochemical characterization.The other high-affinity transporter, Pho89, has been studied less because of its lower activity (as compared to the Pho84) and its preferred alkaline operational conditions. In order to study Pho89 in more detail, a quadruple deletions strain (Pho84Δ Pho87Δ Pho90Δ Pho91Δ) was used.In conclusion, this thesis has contributed to broaden the knowledge of structural and functional aspects of inorganic phosphate transporters.

  • 213.
    Samyn, Dieter R.
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Andersson, M.
    Ruiz-Pavon, Lorena
    Popova, Y.
    Persson, Bengt L.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Thevelein, J.
    The high-affinity inorganic phosphate transport system of Saccharomyces cerevisiae: a tale of two proteins2013In: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 280, p. 152-152Article in journal (Other academic)
  • 214.
    Samyn, Dieter R.
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persson, Bengt L.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Inorganic phosphate and sulfate transport in S. cerevisiae2016In: Yeast Membrane Transport / [ed] José Ramos, Hana Sychrova, Maik Kschischo, Springer, 2016, p. 253-269Chapter in book (Refereed)
    Abstract [en]

    Inorganic ions such as phosphate and sulfate are essential macronutrients required for a broad spectrum of cellular functions and their regulation. In a constantly fluctuating environment microorganisms have for their survival developed specific nutrient sensing and transport systems ensuring that the cellular nutrient needs are met. This chapter focuses on the S. cerevisiae plasma membrane localized transporters, of which some are strongly induced under conditions of nutrient scarcity and facilitate the active uptake of inorganic phosphate and sulfate. Recent advances in studying the properties of the high-affinity phosphate and sulfate transporters by means of site-directed mutagenesis have provided further insight into the molecular mechanisms contributing to substrate selectivity and transporter functionality of this important class of membrane transporters.

  • 215.
    Samyn, Dieter R.
    et al.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Ruiz-Pavon, Lorena
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Andersson, Michael R.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Popova, Yulia
    Katholieke Universiteit Leuven.
    Thevelein, Johan
    Katholieke Universiteit Leuven.
    Persson, Bengt L.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Mutational analysis of putative phosphate- and proton-binding sites in the Saccharomyces cerevisiae Pho84 phosphate:H+ transceptor and its effect on signalling to the PKA and PHO pathways2012In: Biochemical Journal, ISSN 0264-6021, E-ISSN 1470-8728, Vol. 445, p. 413-422Article in journal (Refereed)
    Abstract [en]

    In Saccharomyces cerevisiae, the Pho84 phosphate transporter acts as the main provider of phosphate to the cell using a proton symport mechanism, but also mediates rapid activation of the PKA (protein kinase A) pathway. These two features led to recognition of Pho84 as a transceptor. Although the physiological role of Pho84 has been studied in depth, the mechanisms underlying the transport and sensor functions are unclear. To obtain more insight into the structure–function relationships of Pho84, we have rationally designed and analysed site-directed mutants. Using a three-dimensional model of Pho84 created on the basis of the GlpT permease, complemented with multiple sequence alignments, we selected Arg168 and Lys492, and Asp178, Asp358 and Glu473 as residues potentially involved in phosphate or proton binding respectively, during transport. We found that Asp358 (helix 7) and Lys492 (helix 11) are critical for the transport function, and might be part of the putative substrate-binding pocket of Pho84. Moreover, we show that alleles mutated in the putative proton-binding site Asp358 are still capable of strongly activating PKA pathway targets, despite their severely reduced transport activity. This indicates that signalling does not require transport and suggests that mutagenesis of amino acid residues involved in binding of the co-transported ion may constitute a promising general approach to separate the transport and signalling functions in transceptors.

  • 216.
    Samyn, Dieter R.
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Cocordia University, Canada.
    Van der Veken, Jeroen
    Inst Agr & Fisheries Res ILVO, Belgium.
    Van Zeebroeck, Griet
    VIB, Belgium;Katholieke Univ Leuven, Belgium.
    Persson, Bengt L.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Karlsson, Björn C. G.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Key Residues and Phosphate Release Routes in the Saccharomyces cerevisae Pho84 Transceptor - The Role of Tyr179 in Functional Regulation2016In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 291, no 51, p. 26388-26398Article in journal (Refereed)
    Abstract [en]

    Pho84, a major facilitator superfamily (MFS) protein, is the main high-affinity Pi transceptor in Saccharomyces cerevisiae. Although transport mechanisms have been suggested for other MFS members, the key residues and molecular events driving transport by Pi: H+ symporters are unclear. The current Pho84 transport model is based on the inward-facing occluded crystal structure of the Pho84 homologue PiPT in the fungus Piriformospora indica. However, this model is limited by the lack of experimental data on the regulatory residues for each stage of the transport cycle. In this study, an open, inward-facing conformation of Pho84 was used to study the release of Pi. A comparison of this conformation with the model for Pi release in PiPT revealed that Tyr(179) in Pho84 (Tyr150 in PiPT) is not part of the Pi binding site. This difference may be due to a lack of detailed information on the Pi release step in PiPT. Molecular dynamics simulations of Pho84 in which a residue adjacent to Tyr(179), Asp(178), is protonated revealed a conformational change in Pho84 from an open, inward-facing state to an occluded state. Tyr(179) then became part of the binding site as was observed in the PiPT crystal structure. The importance of Tyr(179) in regulating Pi release was supported by site-directed mutagenesis and transport assays. Using trehalase activity measurements, we demonstrated that the release of Pi is a critical step for transceptor signaling. Our results add to previous studies on PiPT, creating a more complete picture of the proton-coupled Pi transport cycle of a transceptor.

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  • 217.
    Schothorst, Joep
    et al.
    Katholieke Universiteit Leuven, Belgium.
    Nag Kankipati, Harish
    Katholieke Universiteit Leuven, Belgium.
    Conrad, Michaela
    Katholieke Universiteit Leuven, Belgium.
    Samyn, Dieter R.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Van Zeebroeck, Griet
    Katholieke Universiteit Leuven, Belgium.
    Popova, Yulia
    Katholieke Universiteit Leuven, Belgium.
    Rubio-Texeira, Marta
    Katholieke Universiteit Leuven, Belgium.
    Persson, Bengt L.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Thevelein, Johan
    Katholieke Universiteit Leuven, Belgium.
    Yeast nutrient transceptors provide novel insight in the functionality of membrane transporters.2013In: Current Genetics, ISSN 0172-8083, E-ISSN 1432-0983, Vol. 59, no 4, p. 197-206Article in journal (Refereed)
    Abstract [en]

    In the yeast Saccharomyces cerevisiae several nutrient transporters have been identified that possess an additional function as nutrient receptor. These transporters are induced when yeast cells are starved for their substrate, which triggers entry into stationary phase and acquirement of a low protein kinase A (PKA) phenotype. Re-addition of the lacking nutrient triggers exit from stationary phase and sudden activation of the PKA pathway, the latter being mediated by the nutrient transceptors. At the same time, the transceptors are ubiquitinated, endocytosed and sorted to the vacuole for breakdown. Investigation of the signaling function of the transceptors has provided a new read-out and new tools for gaining insight into the functionality of transporters. Identification of amino acid residues that bind co-transported ions in symporters has been challenging because the inactivation of transport by site-directed mutagenesis is not conclusive with respect to the cause of the inactivation. The discovery of nontransported agonists of the signaling function in transceptors has shown that transport is not required for signaling. Inactivation of transport with maintenance of signaling in transceptors supports that a true proton-binding residue was mutagenised. Determining the relationship between transport and induction of endocytosis has also been challenging, since inactivation of transport by mutagenesis easily causes loss of all affinity for the substrate. The use of analogues with different combinations of transport and signaling capacities has revealed that transport, ubiquitination and endocytosis can be uncoupled in several unexpected ways. The results obtained are consistent with transporters undergoing multiple substrate-induced conformational changes, which allow interaction with different accessory proteins to trigger specific downstream events.

  • 218.
    Seisenbaeva, Gulaim A.
    et al.
    Swedish University of Agricultural Sciences.
    Fromell, Karin
    Uppsala University.
    Vinogradov, Vasiliy V.
    ITMO Univ Kronverksky, Russia.
    Terekhov, Aleksey N.
    Ivanovo State Med Acad, Russia.
    Pakhomov, Andrey V.
    Ivanovo State Med Acad, Russia.
    Nilsson, Bo
    Uppsala University.
    Nilsson Ekdahl, Kristina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Uppsala University.
    Vinogradov, Vladimir V.
    ITMO Univ Kronverksky, Russia.
    Kessler, Vadim G.
    Swedish University of Agricultural Sciences.
    Dispersion of TiO2 nanoparticles improves burn wound healing and tissue regeneration through specific interaction with blood serum proteins2017In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, article id 15448Article in journal (Refereed)
    Abstract [en]

    Burn wounds are one of the most important causes of mortality and especially morbidity around the world. Burn wound healing and skin tissue regeneration remain thus one of the most important challenges facing the mankind. In the present study we have addressed this challenge, applying a solution-stabilized dispersion TiO2 nanoparticles, hypothesizing that their ability to adsorb proteins will render them a strong capacity in inducing body fluid coagulation and create a protective hybrid material coating. The in vitro study of interaction between human blood and titania resulted at enhanced TiO2 concentrations in formation of rather dense gel composite materials and even at lower content revealed specific adsorption pattern initiating the cascade response, promising to facilitate the regrowth of the skin. The subsequent in vivo study of the healing of burn wounds in rats demonstrated formation of a strongly adherent crust of a nanocomposite, preventing infection and inflammation with quicker reduction of wound area compared to untreated control. The most important result in applying the TiO2 dispersion was the apparently improved regeneration of damaged tissues with appreciable decrease in scar formation and skin color anomalies.

  • 219.
    Sengottaiyan, Palanivelu
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Petrlova, Jitka
    Lund university.
    Lagerstedt, Jens
    Lund university.
    Ruiz-Pavon, Lorena
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Budamagunta, Madhu
    University of California, USA.
    Voss, John
    University of California, USA.
    Persson, Bengt L.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Characterization of the biochemical and biophysical properties of the Saccharomyces cerevisiae phosphate transporter Pho892013In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 436, no 3, p. 551-556Article in journal (Refereed)
    Abstract [en]

    In Saccharomyces cerevisiae, Pho89 mediates a cation-dependent transport of Pi across the plasma membrane. This integral membrane protein belongs to the Inorganic Phosphate Transporter (PiT) family, a group that includes the mammalian Na+/Pi cotransporters Pit1 and Pit2. Here we report that the Pichia pastoris expressed recombinant Pho89 was purified in the presence of Foscholine-12 and functionally reconstituted into proteoliposomes with a similar substrate specificity as observed in an intact cell system. The alpha-helical content of the Pho89 protein was estimated to 44%. EPR analysis showed that purified Pho89 protein undergoes conformational change upon addition of substrate. 

  • 220.
    Sengottaiyan, Palanivelu
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Ruiz-Pavon, Lorena
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persson, Bengt L.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Functional expression, purification and reconstitution of the recombinant phosphate transporter Pho89 of Saccharomyces cerevisiae2013In: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 280, no 3, p. 965-975Article in journal (Refereed)
    Abstract [en]

    The Saccharomyces cerevisiae high-affinity phosphate transporter Pho89 is a member of the inorganic phosphate (Pi) transporter (PiT) family, and shares significant homology with the type III Na+/Pi symporters, hPit1 and hPit2. Currently, detailed biochemical and biophysical analyses of Pho89 to better understand its transport mechanisms are limited, owing to the lack of purified Pho89 in an active form. In the present study, we expressed functional Pho89 in the cell membrane of Pichia pastoris, solubilized it in Triton X-100 and foscholine-12, and purified it by immobilized nickel affinity chromatography combined with size exclusion chromatography. The protein eluted as an oligomer on the gel filtration column, and SDS/PAGE followed by western blotting analysis revealed that the protein appeared as bands of approximately 63, 140 and 520 kDa, corresponding to the monomeric, dimeric and oligomeric masses of the protein, respec- tively. Proteoliposomes containing purified and reconstituted Pho89 showed Na+-dependent Pi transport activity driven by an artificially imposed electrochemical Na+ gradient. This implies that Pho89 operates as a symporter. Moreover, its activity is sensitive to the Na+ ionophore monensin. To our knowledge, this study represents the first report on the functional reconstitution of a Pi-coupled PiT family member. 

  • 221.
    Sengottaiyan, Palanivelu
    et al.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Spetea, Cornelia
    University of Gothenburg.
    Lagerstedt, Jens O.
    Lund University.
    Samyn, Dieter R.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Andersson, Michael R.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Ruiz-Pavon, Lorena
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Persson, Bengt L.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences. Katholieke Universiteit Leuven, Belgium ; Flanders Institute of Biotechnology, Belgium.
    The intrinsic GTPase activity of the Gtr1 protein from Saccharomyces cerevisiae2012In: BMC Biochemistry, ISSN 1471-2091, E-ISSN 1471-2091, Vol. 13, article id 11Article in journal (Refereed)
    Abstract [en]

    Background

    The Gtr1 protein of Saccharomyces cerevisiae is a member of the RagA subfamily of the Ras-like small GTPase superfamily. Gtr1 has been implicated in various cellular processes. Particularly, the Switch regions in the GTPase domain of Gtr1 are essential for TORC1 activation and amino acid signaling [R. Gong, L. Li, Y. Liu, P. Wang, H. Yang, L. Wang, J. Cheng, K.L. Guan, Y. Xu, Genes Dev. 25 (2011) 1668–1673]. Therefore, knowledge about the biochemical activity of Gtr1 is required to understand its mode of action and regulation.

    Results

    By employing tryptophan fluorescence analysis and radioactive GTPase assays, we demonstrate that Gtr1 can adopt two distinct GDP- and GTP-bound conformations, and that it hydrolyses GTP much slower than Ras proteins. Using cysteine mutagenesis of Arginine-37 and Valine-67, residues at the Switch I and II regions, respectively, we show altered GTPase activity and associated conformational changes as compared to the wild type protein and the cysteine-less mutant.

    Conclusions

    The extremely low intrinsic GTPase activity of Gtr1 implies requirement for interaction with activating proteins to support its physiological function. These findings as well as the altered properties obtained by mutagenesis in the Switch regions provide insights into the function of Gtr1 and its homologues in yeast and mammals.

  • 222.
    Shahini, Negar
    et al.
    Oslo University Hospital, Norway;University of Oslo, Norway.
    Schjalm, Camilla
    Oslo University Hospital, Norway;University of Oslo, Norway.
    Nilsson, Per H.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Oslo University Hospital, Norway;University of Oslo, Norway.
    Holt, Margrethe Flesvig
    Oslo University Hospital, Norway;University of Oslo, Norway.
    Ogaard, Jonas D. S.
    Oslo University Hospital, Norway;University of Oslo, Norway.
    Lien, Egil
    UMass Med Sch, USA;Norwegian Univ Sci & Technol, Norway.
    Ahmed, Muhammad S.
    Oslo University Hospital, Norway;University of Oslo, Norway.
    Attramadal, Havard
    Oslo University Hospital, Norway;University of Oslo, Norway.
    Aukrust, Pal
    Oslo University Hospital, Norway;University of Oslo, Norway;Norwegian Univ Sci & Technol, Norway.
    Yndestad, Arne
    Oslo University Hospital, Norway;University of Oslo, Norway.
    Mollnes, Tom Eirik
    Oslo University Hospital, Norway;University of Oslo, Norway;Norwegian Univ Sci & Technol, Norway;Nordland Hosp, Norway;Univ Tromsö, Norway.
    Louwe, Mieke C.
    Oslo University Hospital, Norway;University of Oslo, Norway.
    Complement component C3 and the TLR co-receptor CD14 are not involved in angiotensin II induced cardiac remodelling2020In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 523, no 4, p. 867-873Article in journal (Refereed)
    Abstract [en]

    Inflammation is centrally involved in the development of cardiac hypertrophy and the processes of remodelling. The complement system and Toll-like receptor (TLR) family, two upstream arms of the innate immune system, have previously been reported to be involved in cardiac remodelling. However, the role of complement component 3 (C3), TLR co-receptor CD14 and the synergy between them have not been addressed during pressure overload-induced cardiac remodelling. Here, we examined angiotensin II-induced cardiac hypertrophy and remodelling for 7 days in male C57Bl/6 J mice deficient in C3, CD14, or both (C3CD14), and WT controls. Angiotensin II infusion induced a mild concentric hypertrophic phenotype in WT mice with increased left ventricle weight, wall thicknesses and reduced ventricular internal diameter, associated with increased cardiac fibrosis. However, there were no differences between WT mice and mice deficient for C3, CD14 or C3CD14, as systolic blood pressure, cardiac function and structure and levels of fibrosis were comparable between WT mice and the three other genotypes. C5a did not change in angiotensin II treated mice, whereas Mac2 levels were increased in angiotensin II treated mice, but did not differ between genotypes. The inflammatory IL-6 response was comparable between WT and C3 deficient mice, however, it was decreased in CD14 and C3CD14 deficient mice. We conclude that deficiency in C3, CD14 or C3CD14 had no effect on cardiac remodelling following angiotensin II-induced pressure overload. This suggests that C3 and CD14 are not involved in angiotensin II-induced adverse cardiac remodelling. (C) 2020 Published by Elsevier Inc.

  • 223. Shand, Jessica C
    et al.
    Jansson, Johan
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Hsu, Yu-Chiao
    Campbell, Andrew
    Mullen, Craig A
    Differential gene expression in acute lymphoblastic leukemia cells surviving allogeneic transplant.2010In: Cancer Immunology and Immunotherapy, ISSN 0340-7004, E-ISSN 1432-0851, Vol. 59, no 11, p. 1633-1644Article in journal (Refereed)
    Abstract [en]

    The effectiveness of allogeneic graft-versus-leukemia (GVL) activity in control of acute lymphoblastic leukemia is generally regarded as poor. One possible factor is dynamic adaptation of the leukemia cell to the allogeneic environment. This work tested the hypothesis that the pattern of gene expression in acute lymphoblastic leukemia cells in an allogeneic environment would differ from that in a non-allogeneic environment. Expression microarray studies were performed in murine B lineage acute lymphoblastic leukemia cells recovered from mice that had undergone allogeneic MHC-matched but minor histocompatibility antigen mismatched transplants. A limited number of genes were found to be differentially expressed in ALL cells surviving in the allogeneic environment. Functional analysis demonstrated that genes related to immune processes, antigen presentation, ubiquitination and GTPase function were significantly enriched. Several genes with known immune activities potentially relevant to leukemia survival (Ly6a/Sca-1, TRAIL and H2-T23) were examined in independent validation experiments. Increased expression in vivo in allogeneic hosts was observed, and could be mimicked in vitro with soluble supernatants of mixed lymphocyte reactions or interferon-gamma. The changes in gene expression were reversible when the leukemia cells were removed from the allogeneic environment. These findings suggest that acute lymphoblastic leukemia cells respond to cytokines present after allogeneic transplantation and that these changes may reduce the effectiveness of GVL activity.

  • 224.
    Shen, Qian
    et al.
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Zhang, Lida
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Liao, Zhihua
    Southwest Univ, Peoples Republic of China.
    Wang, Shengyue
    Chinese Natl Human Genome Ctr Shanghai, Peoples Republic of China.
    Yan, Tingxiang
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Shi, Pu
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Liu, Meng
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Fu, Xueqing
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Pan, Qifang
    Shanghai Jiao Tong Univ,Peoples Republic of China.
    Wang, Yuliang
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Lv, Zongyou
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Lu, Xu
    Shanghai Jiao Tong Univ, Peoples Republic of China;China Pharmaceut Univ, Peoples Republic of China.
    Zhang, Fangyuan
    Shanghai Jiao Tong Univ, Peoples Republic of China;Southwest Univ, Peoples Republic of China.
    Jiang, Weimin
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Ma, Yanan
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Chen, Minghui
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Hao, Xiaolong
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Li, Ling
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Tang, Yueli
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Lv, Gang
    Chinese Natl Human Genome Ctr Shanghai, Peoples Republic of China.
    Zhou, Yan
    Chinese Natl Human Genome Ctr Shanghai, Peoples Republic of China.
    Sun, Xiaofen
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    Brodelius, Peter E.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Rose, Jocelyn K. C.
    Cornell Univ, USA.
    Tang, Kexuan
    Shanghai Jiao Tong Univ, Peoples Republic of China.
    The Genome of Artemisia annua Provides Insight into the Evolution of Asteraceae Family and Artemisinin Biosynthesis2018In: Molecular Plant, ISSN 1674-2052, E-ISSN 1752-9867, Vol. 11, no 6, p. 776-788Article in journal (Refereed)
    Abstract [en]

    Artemisia annua, commonly known as sweet wormwood or Qinghao, is a shrub native to China and has long been used for medicinal purposes. A. annua is now cultivated globally as the only natural source of a potent anti-malarial compound, artemisinin. Here, we report a high-quality draft assembly of the 1.74-gigabase genome of A. annua, which is highly heterozygous, rich in repetitive sequences, and contains 63 226 protein-coding genes, one of the largest numbers among the sequenced plant species. We found that, as one of a few sequenced genomes in the Asteraceae, the A. annua genome contains a large number of genes specific to this large angiosperm clade. Notably, the expansion and functional diversification of genes encoding enzymes involved in terpene biosynthesis are consistent with the evolution of the artemisinin biosynthetic pathway. We further revealed by transcriptome profiling that A. annua has evolved the sophisticated transcriptional regulatory networks underlying artemisinin biosynthesis. Based on comprehensive genomic and transcriptomic analyses we generated transgenic A. annua lines producing high levels of artemisinin, which are now ready for large-scale production and thereby will help meet the challenge of increasing global demand of artemisinin.

  • 225.
    Shneyer, Boris I.
    et al.
    Technion, Israel.
    Ušaj, Marko
    Technion, Israel.
    Henn, Arnon
    Technion, Israel.
    Myo19 is an outer mitochondrial membrane motor and effector of starvation-induced filopodia2016In: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 129, no 3, p. 543-556Article in journal (Refereed)
    Abstract [en]

    Mitochondria respond to environmental cues and stress conditions. Additionally, the disruption of the mitochondrial network dynamics and its distribution is implicated in a variety of neurodegenerative diseases. Here, we reveal a new function for Myo19 in mitochondrial dynamics and localization during the cellular response to glucose starvation. Ectopically expressed Myo19 localized with mitochondria to the tips of starvation-induced filopodia. Corollary to this, RNA interference (RNAi)-mediated knockdown of Myo19 diminished filopodia formation without evident effects on the mitochondrial network. We analyzed the Myo19–mitochondria interaction, and demonstrated that Myo19 is uniquely anchored to the outer mitochondrial membrane (OMM) through a 30–45-residue motif, indicating that Myo19 is a stably attached OMM molecular motor. Our work reveals a new function for Myo19 in mitochondrial positioning under stress.

  • 226.
    Shneyer, Boris I.
    et al.
    Technion, Israel.
    Ušaj, Marko
    Technion, Israel.
    Wiesel-Motiuk, Naama
    Technion, Israel.
    Regev, Ronit
    Technion, Israel.
    Henn, Arnon
    Technion, Israel.
    ROS induced distribution of mitochondria to filopodia by Myo19 depends on a class specific tryptophan in the motor domain2017In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, no 1, article id 11577Article in journal (Refereed)
    Abstract [en]

    The role of the actin cytoskeleton in relation to mitochondria function and dynamics is only recently beginning to be recognized. Myo19 is an actin-based motor that is bound to the outer mitochondrial membrane and promotes the localization of mitochondria to filopodia in response to glucose starvation. However, how glucose starvation induces mitochondria localization to filopodia, what are the dynamics of this process and which enzymatic adaptation allows the translocation of mitochondria to filopodia are not known. Here we show that reactive oxygen species (ROS) mimic and mediate the glucose starvation induced phenotype. In addition, time-lapse fluorescent microscopy reveals that ROS-induced Myo19 motility is a highly dynamic process which is coupled to filopodia elongation and retraction. Interestingly, Myo19 motility is inhibited by back-to-consensus-mutation of a unique residue of class XIX myosins in the motor domain. Kinetic analysis of the purified mutant Myo19 motor domain reveals that the duty ratio (time spent strongly bound to actin) is highly compromised in comparison to that of the WT motor domain, indicating that Myo19 unique motor properties are necessary to propel mitochondria to filopodia tips. In summary, our study demonstrates the contribution of actin-based motility to the mitochondrial localization to filopodia by specific cellular cues.

  • 227.
    Shneyer, Boris
    et al.
    Technion, Israel.
    Ušaj, Marko
    Technion, Israel.
    Henn, Arnon
    Technion, Israel.
    Myosin 19 is Anchored to the Mitochondria, Affecting its Localization and Morphology2015In: Biophysical Journal supplement 1, 2015, Vol. 108, p. 303a-, article id 1517-PosConference paper (Refereed)
    Abstract [en]

    Mitochondria undergo continuous cycles of fission and fusion creating a highly dynamic network, which is essential for its proper functions in apoptosis, ATP generation and calcium homeostasis. Mitochondria long-range motility relies on the microtubule motors kinesin and dynein. Recently, actin and myosin 19 have been implicated in mitochondrial motility in vertebrates. However, the interaction of endogenous myosin 19 with the mitochondria remains unknown. Here, we show using multiple complementary approaches that endogenous myosin 19 is anchored directly to the outer mitochondrial membrane (OMM) in a monotopic fashion. We have identified a region of 30 residues at the tail domain of myosin 19, which is both essential and sufficient for myosin 19-OMM interaction. Furthermore, we have purified to near homogeneity a 45 long peptide comprised of this region to study its biochemical and biophysical properties. We performed in-vitro binding assay by fluorescence anisotropy of this specific purified peptide to vesicles with different phospholipid compositions. Our results revealed that that this peptide binds to vesicles mimicking the OMM with the highest affinity. To relate this tight binding to the mitochondria to myosin 19 ATPase activity, we have purified myosin 19-3IQ construct and measured its actin-dependent steady state ATPase activity. Interestingly, we found that it is completely inhibited by very low calcium concentration, suggesting that myosin 19 activity may be regulated by local calcium concentration. The interaction between a motor protein and an organelle, and the calcium dependence implicates that myosin 19 plays a role in mitochondria network dynamics.

  • 228. Sode, K
    et al.
    Brodelius, Peter
    Institute of Biotechnology, Swiss Federal Institute of Technology, Hönggerberg, CH-8093 Zfirich, Switzerland.
    Meussdoerffer, F
    Mosbach, K
    Ernst, J
    Continuous Production of Somatomedin C with Immobilized Transformed Yeast Cells1988In: Applied Microbiology and Biotechnology, ISSN 0175-7598, E-ISSN 1432-0614, Vol. 28, no 3, p. 215-221Article in journal (Refereed)
    Abstract [en]

    Yeast cells producing the growth hormonesomatomedin C (SMC) were constructedand applied in the immobilized form continuouslyfor a period of over 10 days in a flowthroughbioreactor. The construction of theMF~I-SMC fusion vector p336/l is given as wellas the results of the influence of various nutrientseffecting hormone production. Immobilization ofthe transformed yeast cells is described and theirapplication in a continuous bioreactor system.This study demonstrates the feasibility of a longtermand high-level hormone production by immobilizedtransformed yeast. The SMC productivitiesof free cells in batch and immobilized cellsunder continuous conditions were 0.2--0.3 and0.5--0.6 mg per g wet cells and day, respectively. 

  • 229.
    Solomon, Mellat
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Undersökning av genuttrycket av proteorhodopsin i Dokdonia donghaensis MED134 med qPCR-analyser2019Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Abstract [en]

    Dokdonia donghaensis MED134 is a marine bacterium that uses carbon and nitrogen for its energy source. These bacteria also have the ability to generate energy from light by using a membrane-bound proton pump called proteorhodopsin. In this work, qPCR analysis (quantitative polymerase chain reaction) was used in two different experiments, colony and evolution experiment. The aim of the colony experiment was to compare the proteorhodopsin expression in D. donghaensis MED134 in colonies from primary- and tertiary streak on a plate that has been allowed to grow in light and dark for three and seven days. The purpose of the evolution experiment was to investigate the bacterial growth and compare the expression of proteorhodopsin in generation 130 in D. donghaensis MED134 with the wild type sample (generation 0) which has grown in light and dark in a liquid medium. The results of the gene expression analysis showed that the colonies that grew at the primary streak (light), had tendency to higher proteorhodopsin expression compared to the tertiary streak (light) at day 3. The proteorhodopsin expression in light increased in the tertiary streak at day 7. In the evolution experiment, generation 0 and 130 (light) had higher growth than those that grew in the dark. Generation 130 (light) had tendency to higher proteorhodopsin expression compared to all others, and generation 130 in the dark had less proteorhodopsin expression than generation 0 in the dark. Statistical tests showed no significant difference between the groups in both experiments. The result in this study indicate that the expression of proteorhodopsin varies depending on time and where on the plate the bacteria have grown since competition plays a major role. Evolution may also have contributed to better survival for generation 130 (light) and worse for generation 130 (dark).

  • 230.
    Srinivas, Vivek
    et al.
    Stockholm University.
    Lebrette, Hugo
    Stockholm University.
    Lundin, Daniel
    Stockholm University.
    Kutin, Yuri
    Max Planck Inst Chem Energy Convers, Germany.
    Sahlin, Margareta
    Stockholm University.
    Lerche, Michael
    Stockholm University.
    Eirich, Jürgen
    Karolinska Institutet.
    Branca, Rui M. M.
    Karolinska Institutet.
    Cox, Nicholas
    Australian Natl Univ, Australia.
    Sjöberg, Britt-Marie
    Stockholm University.
    Högbom, Martin
    Stockholm University;Stanford Univ, USA.
    Metal-free ribonucleotide reduction powered by a DOPA radical in Mycoplasma pathogens2018In: Nature, ISSN 0028-0836, E-ISSN 1476-4687, Vol. 563, no 7731, p. 416-420Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductase (RNR) catalyses the only known de novo pathway for the production of all four deoxyribonucleotides that are required for DNA synthesis(1,2). It is essential for all organisms that use DNA as their genetic material and is a current drug target(3,4). Since the discovery that iron is required for function in the aerobic, class I RNR found in all eukaryotes and many bacteria, a dinuclear metal site has been viewed as necessary to generate and stabilize the catalytic radical that is essential for RNR activity(5-7). Here we describe a group of RNR proteins in Mollicutes-including Mycoplasma pathogens-that possess a metal-independent stable radical residing on a modified tyrosyl residue. Structural, biochemical and spectroscopic characterization reveal a stable 3,4-dihydroxyphenylalanine (DOPA) radical species that directly supports ribonucleotide reduction in vitro and in vivo. This observation overturns the presumed requirement for a dinuclear metal site in aerobic ribonucleotide reductase. The metal-independent radical requires new mechanisms for radical generation and stabilization, processes that are targeted by RNR inhibitors. It is possible that this RNR variant provides an advantage under metal starvation induced by the immune system. Organisms that encode this type of RNR-some of which are developing resistance to antibiotics-are involved in diseases of the respiratory, urinary and genital tracts. Further characterization of this RNR family and its mechanism of cofactor generation will provide insight into new enzymatic chemistry and be of value in devising strategies to combat the pathogens that utilize it. We propose that this RNR subclass is denoted class Ie.

  • 231.
    Strand, Malin
    et al.
    Swedish University of Agricultural Sciences.
    Andersson, Håkan S.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Slemmaskens hemlighet2016In: Forskning & Framsteg, ISSN 0015-7937, no 2, p. 26-33Article in journal (Other (popular science, discussion, etc.))
    Abstract [sv]

    Nyckeln till framtidens mediciner kan gömma sig hos slemmiga, giftiga maskar som lever på havets botten. Här berättar marinbiologen Malin Strand och biokemisten Håkan Andersson om jakten som ledde till en oväntad upptäckt – och som tog slemmasken från det marinbiologiska laboratoriet på Tjärnö och Sveriges västkust till kemilaboratoriet i Uppsala.

  • 232.
    Svensson, P. Andreas
    et al.
    Monash University, Australia.
    Wong, B. B. M.
    Monash University, Australia.
    Carotenoid-based signals in behavioural ecology: a review2011In: Behaviour, ISSN 0005-7959, E-ISSN 1568-539X, Vol. 148, no 2, p. 131-189Article in journal (Refereed)
    Abstract [en]

    Carotenoids are among the most prevalent pigments used in animal signals and are also important for a range of physiological functions. These concomitant roles havemade carotenoidbased signals a popular topic in behavioural ecology while also causing confusion and controversy. After a thorough background, we review the many pitfalls, caveats and seemingly contradictory conclusions that can result from not fully appreciating the complex nature of carotenoid function. Current controversies may be resolved through a more careful regard of this complexity, and of the immense taxonomic variability of carotenoid metabolism. Studies investigating the physiological trade-offs between ornamental and physiological uses of carotenoids have yielded inconsistent results. However, in many studies, homeostatic regulation of immune and antioxidant systems may have obscured the effects of carotenoid supplementation. We highlight how carefully designed experiments can overcome such complications. There is also a need to investigate factors other than physiological trade-offs (such as predation risk and social interactions) as these, too, may shape the expression of carotenoidbased signals.Moreover, the processes limiting signal expression individuals are likely different from those operating over evolutionary time-scales. Future research should give greater attention to carotenoid pigmentation outside the area of sexual selection, and to taxa other than fishes and birds.

  • 233. Szwajcer, E
    et al.
    Brodelius, Peter
    Lunds Universitet.
    Mosbach, K
    Production of alfa-Keto Acids. Part II - Immobilized Cells of Providencia sp. PCM 1298 Containing L-Amino Acid Oxidase1982In: Enzyme and microbial technology, Vol. 4, p. 409-413Article in journal (Refereed)
  • 234.
    Sävneby, Anna
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Jonsson, Nina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Svensson, P. Andreas
    Linnaeus University, Faculty of Health and Life Sciences, Department of Biology and Environmental Science.
    Hafenstein, Susan
    The Pennsylvania State University College of Medicine, USA.
    Lindberg, A. Michael
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Virus derived from an Enterovirus B construct efficiently reverts from a frameshift mutation immediately beyond the translation initiation siteManuscript (preprint) (Other academic)
  • 235.
    Sävneby, Anna
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Luthman, Johannes
    Karolinska institutet.
    Nordenskjöld, Fabian
    Karolinska institutet.
    Andersson, Björn
    Karolinska institutet.
    Lindberg, A. Michael
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Gene expression in rhabdomyosarcoma cells infected with cytolytic and non-cytolytic variants of coxsackievirus B2 OhioManuscript (preprint) (Other academic)
  • 236.
    Sävneby, Anna
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Lysholm, Fredrik
    Karolinska institutet ; Linköpings universitet.
    Jonsson, Nina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Edman, Kjell
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Allander, Tobias
    Karolinska Institutet.
    Andersson, Björn
    Karolinska Institutet.
    Lindberg, A. Michael
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Restricted replication of the rhinovirus C34 prototype in standard cell linesManuscript (preprint) (Other academic)
  • 237.
    Takatsuki, Hideyo
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Bengtsson, Elina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Persistence length of fascin-cross-linked actin filament bundles in solution and the in vitro motility assay2014In: Biochimica et Biophysica Acta - General Subjects, ISSN 0304-4165, E-ISSN 1872-8006, Vol. 1840, no 6, p. 1933-1942Article in journal (Refereed)
    Abstract [en]

    Background: Bundles of unipolar actin filaments (F-actin), cross-linked via the actin-binding protein fascin, are important in filopodia of motile cells and stereocilia of inner ear sensory cells. However, such bundles are also useful as shuttles in myosin-driven nanotechnological applications. Therefore, and for elucidating aspects of biological function, we investigate if the bundle tendency to follow straight paths (quantified by path persistence length) when propelled by myosin motors is directly determined by material properties quantified by persistence length of thermally fluctuating bundles. Methods: Fluorescent bundles, labeled with rhodamine-phalloidin, were studied at fascin:actin molar ratios: 0:1 (F-actin), 1:7, 1:4 and 1:2. Persistence lengths (Lp) were obtained by fitting the cosine correlation function (CCF) to a single exponential function: <cos(theta(0) theta(s)) > = exp(-s / (2Lp)) where theta(s) is tangent angle; s: path or contour lengths. < > denotes averaging over filaments. Results: Bundle-Lp (bundles < 15 mu m long) increased from similar to 10 to 150 mu m with increased fascin:actin ratio. The increase was similar for path-Lp (path < 15 mu m), with highly linear correlation. For longer bundle paths, the CCF-decay deviated from a single exponential, consistent with superimposition of the random path with a circular path as suggested by theoretical analysis. Conclusions: Fascin-actin bundles have similar path-Lp and bundle-Lp, both increasing with fascin:actin ratio. Path-Lp is determined by the flexural rigidity of the bundle. General significance: The findings give general insight into mechanics of cytoskeletal polymers that interact with molecular motors, aid rational development of nanotechnological applications and have implications for structure and in vivo functions of fascin-actin bundles. (C) 2014 The Authors. Published by Elsevier B.V.

  • 238.
    Takatsuki, Hideyo
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Characterization and Stabilization of Fascin-Bundled Actin Filaments Transported by Heavy Meromyosin2015In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, no 2, Supplement 1, p. 299A-299A, article id 1496-PosArticle in journal (Other academic)
  • 239.
    Teixeira, M.C.
    et al.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Coelho, N.
    Olsson, Mikael E.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Brodelius, Peter E.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Carvalho, I.S.
    Brodelius, Maria
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Molecular cloning and expression analysis of three omega-6 desaturase genes from purslane (Portulaca oleracea L.)2009In: Biotechnology letters, ISSN 0141-5492, E-ISSN 1573-6776, Vol. 31, no 7, p. 1089-1101Article in journal (Refereed)
    Abstract [en]

    Two full-length cDNA clones of PoleFAD2 and one full-length cDNA clone of PoleFAD6, encoding omega-6 fatty acid desaturases, the key enzymes for the conversion of oleic into linoleic acid, were isolated from purslane (Portulaca oleracea L.) leaves and seeds. The deduced amino acid sequence of both isoforms of PoleFAD2 showed higher similarities to other microsomal omega-6 desaturases then to PoleFAD6 or other plastidial orthologues, and vice versa. Expression analysis by RT-PCR showed that all genes are expressed in all tissues of purslane tested, but higher levels of mRNA accumulation were detected in reproductive organs and cells that proliferate rapidly or store lipids. Wounding affected the levels of mRNA accumulation of both, FAD2 and FAD6 genes in purslane leaves, while chilling stress affected only FAD2 transcript level. The expression patterns observed reflect the discrete roles of these genes in membrane synthesis for cell division, thylakoid development, and lipid storage or in the biosynthetic pathway for the production of signaling molecules that influence plant development or defense.

  • 240.
    ten Siethoff, Lasse
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Towards Myosin Powered Lab-on-a-Chip Devices2013Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Myosins are protein motors that use chemical energy in the form of adenosinetriphosphate to produce force and motion. These molecular motors might be usedto power transportation in Lab-on-a-chip devices where a series of laboratory tasks(e.g. separation, concentration and detection) are performed in one sequence on asmall chip. Because of the small size, lab-on-a-chip devices are predicted to befaster and more sensitive than conventional systems. Further potential advantagesinclude cost efficiency and the possibility to perform many analyzes in parallel.Substituting microfluidics with myosin based transport would allow furtherminiaturization and make lab-on-a-chip devices more readily portable by reducingthe need for external power supplies. However, there are also limitations thathamper the development of such devices. Here we investigate several aspects of amyosin powered lab-on-a-chip device and present ways to overcome criticallimitations. First we demonstrate covalent attachment of antibodies to actinfilament shuttles with retained ability of the filaments to be propelled by myosinfragments, previously believed to be difficult. Secondly we develop a separationmethod to overcome the deleterious effects of body fluids on the actomyosinsystem. Thirdly, we explore the possibility to concentrate actin shuttles on ananostructured surface and achieve >20 times concentration in <1 min. Monte-Carlo simulations of the concentration process suggest further room forimprovement. Fourth, we develop novel techniques for fast and automaticdetection of fluorescence at certain check points which improves S/N ratio >20times. Finally, we take the first steps towards the development of threedimensional,nanowire-based transport systems, important both for lab-on-a-chipapplications and fundamental studies. Our results demonstrate the potential of amyosin based lab-on-a-chip device and lay the foundation for furtherdevelopments. Thus, we anticipate that this work will influence future studiestowards a complete diagnostic lab-on-a-chip work-up based on molecular motors.In addition, the work might also have implications for the development of futurebiocomputation and drug screening devices as well as novel biophysical studies ofthe actomyosin system.

  • 241.
    Teramura, Yuji
    et al.
    Univ Tokyo, Japan.
    Asif, Sana
    Uppsala University.
    Gustafson, Elisabet
    Univ Uppsala Hosp.
    Nilsson Ekdahl, Kristina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Uppsala University.
    Nilsson, Bo
    Uppsala University.
    Heparinzation of cell surfaces with a heparin-binding peptide-conjugated PEG-lipid for regulation of thromboinflammation in transplantation of human MSC and hepatocyte2015In: Xenotransplantation, ISSN 0908-665X, E-ISSN 1399-3089, Vol. 22, no Supplement 1, p. S70-S70, article id 561Article in journal (Other academic)
  • 242.
    Teramura, Yuji
    et al.
    University of Tokyo, Japan;Uppsala University.
    Asif, Sana
    Uppsala University.
    Nilsson Ekdahl, Kristina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Uppsala University.
    Gustafson, Elisabet
    Uppsala University Hospital.
    Nilsson, Bo
    Uppsala University.
    Cell Adhesion Induced Using Surface Modification with Cell-Penetrating Peptide-Conjugated Poly(ethylene glycol)-Lipid: A New Cell Glue for 3D Cell-Based Structures2017In: ACS Applied Materials and Interfaces, ISSN 1944-8244, E-ISSN 1944-8252, Vol. 9, no 1, p. 244-254Article in journal (Refereed)
    Abstract [en]

    We synthesized a novel material, cell-penetrating peptide conjugated poly(ethylene glycol)-lipid (CPP-PEG-lipid), that can induce the adhesion of floating cells. Firm cell adhesion with spreading could be induced by cell surface modification with the CPP-PEG-lipids. Cell adhesion was induced by CPPs but not by any other cationic short peptides we tested. Here, we demonstrated adherence using the floating cell line CCRF-CEM as well as primary human T cells, B cells, erythrocytes, and hepatocytes. As compared to cells grown in suspension, adherent cells were more rapidly induced to attach to substrates with the cell-surface modification. The critical factor for attachment was localization of CPPs at the cell membrane by PEG-lipids with PEG > 20 kDa. These cationic CPPs on PEG chains were able to interact with substrate surfaces such as polystyrene (PS) surfaces, glass surfaces, and PS microfibers that are negatively charged, inducing firm cell adhesion and cell spreading. Also, as opposed to normal cationic peptides that interact strongly with cell membranes, CPPs were less interactive with the cell surfaces because of their cell-penetrating property, making them more available for adhering cells to the substrate surface. No effects on cell viability or cell proliferation were observed after the induction of cell adhesion. With this technique, cells could be easily immobilized onto PS microfibers, an important step in fabricating 3D cell-based structures. Cells immobilized onto 3D PS microfibers were alive, and human hepatocytes showed normal production of urea and albumin on the microfibers. This method is novel in inducing firm cell adhesion-via a one-step treatment.

  • 243.
    Toda, Shota
    et al.
    Shibaura Inst Technol, Japan.
    Fattah, Artin
    Uppsala University, Sweden.
    Asawa, Kenta
    Univ Tokyo, Japan.
    Nakamura, Naoko
    Shibaura Inst Technol, Japan.
    Nilsson Ekdahl, Kristina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Uppsala University, Sweden.
    Nilsson, Bo
    Uppsala University, Sweden.
    Teramura, Yuji
    Uppsala University, Sweden;Univ Tokyo, Japan.
    Optimization of Islet Microencapsulation with Thin Polymer Membranes for Long-Term Stability2019In: Micromachines, ISSN 2072-666X, E-ISSN 2072-666X, Vol. 10, no 11, p. 1-10, article id 755Article in journal (Refereed)
    Abstract [en]

    Microencapsulation of islets can protect against immune reactions from the host immune system after transplantation. However, sufficient numbers of islets cannot be transplanted due to the increase of the size and total volume. Therefore, thin and stable polymer membranes are required for the microencapsulation. Here, we undertook the cell microencapsulation using poly(ethylene glycol)-conjugated phospholipid (PEG-lipid) and layer-by-layer membrane of multiple-arm PEG. In order to examine the membrane stability, we used different molecular weights of 4-arm PEG (10k, 20k and 40k)-Mal to examine the influence on the polymer membrane stability. We found that the polymer membrane made of 4-arm PEG(40k)-Mal showed the highest stability on the cell surface. Also, the polymer membrane did not disturb the insulin secretion from beta cells.

  • 244.
    Unelius, C. Rikard
    et al.
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Park, K. -C
    McNeill, M.
    Wee, S. L.
    Bohman, Björn
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Suckling, D. M.
    Identification and electrophysiological studies of (4S,5S)-5-hydroxy-4-methyl-3-heptanone and 4-methyl-3,5-heptanedione in male lucerne weevils2013In: Die Naturwissenschaften, ISSN 0028-1042, E-ISSN 1432-1904, Vol. 100, no 2, p. 135-143Article in journal (Refereed)
    Abstract [en]

    An investigation to identify a sex or aggregation pheromone of Sitona discoideus GyllenhAyenl (Coleoptera: Curculionidae) is presented. Antenna flicking and attraction behaviors evoked by conspecifics of both sexes were recorded in arena bioassays, where attraction of females to males was observed. Air entrainment of both males and females was conducted in separate chambers. Gas chromatographic-mass spectrometric analysis of headspace volatiles revealed that two male-specific compounds, 4-methyl-3,5-heptanedione (major) and (4S,5S)-5-hydroxy-4-methyl-3-heptanone (minor), were emitted during the autumnal post-aestivatory flight period. The stereoisomers of the minor component were separated by enantioselective gas chromatography and their absolute configurations assigned by NMR (diastereomers) and the known preference of enantioselective transesterification reactions catalyzed by Candida antarctica lipase B. Electroantennogram and single sensillum recording studies indicate that 4-methyl-3,5-heptanedione as well as all individual stereoisomers of 5-hydroxy-4-methyl-3-heptanone are detected by the antennae of male and female S. discoideus. Further, single sensillum recordings suggest that both sexes of S. discoideus have specialized olfactory receptor neurons (ORNs) for detecting 4-methyl-3,5-heptanedione and different populations of stereoselective ORNs for detecting the stereoisomers of 5-hydroxy-4-methyl-3-heptanone. Some of these stereoselective ORNs appear to be sex-specific in S. discoideus.

  • 245.
    Ušaj, Marko
    et al.
    Technion, Israel.
    Henn, Arnon
    Technion, Israel.
    Kinetic adaptation of human Myo19 for active mitochondrial transport to growing filopodia tips2017In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, no 1, article id 11596Article in journal (Refereed)
    Abstract [en]

    Myosins are actin-based molecular motors which are enzymatically adapted for their cellular functions such as transportation and membrane tethering. Human Myo19 affects mitochondrial motility, and promotes their localization to stress-induced filopodia. Therefore, studying Myo19 enzymology is essential to understand how this motor may facilitate mitochondrial motility. Towards this goal, we have purified Myo19 motor domain (Myo19-3IQ) from a human-cell expression system and utilized transient kinetics to study the Myo19-3IQ ATPase cycle. We found that Myo19-3IQ exhibits noticeable conformational changes (isomerization steps) preceding both ATP and ADP binding, which may contribute to nucleotide binding regulation. Notably, the ADP isomerization step and subsequent ADP release contribute significantly to the rate-limiting step of the Myo19-3IQ ATPase cycle. Both the slow ADP isomerization and ADP release prolong the time Myo19-3IQ spend in the strong actin binding state and hence contribute to its relatively high duty ratio. However, the predicted duty ratio is lower than required to support motility as a monomer. Therefore, it may be that several Myo19 motors are required to propel mitochondria movement on actin filaments efficiently. Finally, we provide a model explaining how Myo19 translocation may be regulated by the local ATP/ADP ratio, coupled to the mitochondria presence in the filopodia.

  • 246.
    Ušaj, Marko
    et al.
    Technion - Israel Institute of Technology, Israel.
    Zattelman, Lilach
    Technion - Israel Institute of Technology, Israel.
    Regev, Ronit
    Technion - Israel Institute of Technology, Israel.
    Shneyer, Boris I.
    Technion - Israel Institute of Technology, Israel.
    Wiesel-Motiuk, Naama
    Technion - Israel Institute of Technology, Israel.
    Henn, Arnon
    Technion - Israel Institute of Technology, Israel.
    Overexpression and purification of human myosins from transiently and stably transfected suspension adapted HEK293SF-3F6 cells2018In: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 558, no 1, p. 19-27Article in journal (Refereed)
    Abstract [en]

    The myosin family of motor proteins is an attractive target of therapeutic small-molecule protein inhibitors and modulators. Milligrams of protein quantities are required to conduct proper biophysical and biochemical studies to understand myosin functions. Myosin protein expression and purification represent a critical starting point towards this goal. Established utilization of Dictyostelium discoideum, Drosophila melanogaster, insect and mouse cells for myosin expression and purification is limited, cost, labor and time inefficient particularly for (full-length) human myosins. Here we are presenting detailed protocols for production of several difficult-to-purify recombinant human myosins in efficient quantities up to 1 mg of protein per liter of cell culture. This is the first time that myosins have been purified in large scales from suspension adapted transiently and stably expressing human cells. The method is also useful for expressing other human proteins in quantities sufficient to perform extensive biochemical and biophysical characterization.

  • 247. van Zalinge, Harm
    et al.
    Aveyard, Jenny
    Hajne, Joanna
    Persson, Malin
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Nicolau, Dan V.
    Actin Filament Motility Induced Variation of Resonance Frequency and Rigidity of Polymer Surfaces Studied by Quartz Crystal Microbalance2012In: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 28, no 42, p. 15033-15037Article in journal (Refereed)
    Abstract [en]

    This contribution reports on the quantification of the parameters of the motility assays for actomyosin system using a quartz crystal microbalance (QCM). In particular, we report on the difference in the observed resonance frequency and dissipation of a quartz crystal when actin filaments are stationary as opposed to when they are motile. The changes in QCM measurements were studied for various polymer-coated surfaces functionalized with heavy meromyosin (HMM). The results of the QCM experiments show that the HMM-induced sliding velocity of actin filaments is modulated by a combination of the viscoelastic properties of the polymer layer including the HMM motors.

  • 248.
    van Zalinge, Harm
    et al.
    Univ Liverpool, UK.
    Ramsey, Laurence C.
    Univ Liverpool, UK.
    Aveyard, Jenny
    Univ Liverpool, UK.
    Persson, Malin
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Nicolau, Dan V.
    Univ Liverpool, UK ; McGill Univ, Canada.
    Surface-Controlled Properties of Myosin Studied by Electric Field Modulation2015In: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 31, no 30, p. 8354-8361Article in journal (Refereed)
    Abstract [en]

    The efficiency of dynamic nanodevices using surface-immobilized protein molecular motors, which have been proposed for diagnostics, drug discovery, and biocomputation, critically depends on the ability to precisely control the motion of motor-propelled, individual cytoskeletal filaments transporting cargo to designated locations. The efficiency of these devices also critically depends on the proper function of the propelling motors, which is controlled by their interaction with the surfaces they are immobilized on. Here we use a microfluidic device to study how the motion of the motile elements, i.e., actin filaments propelled by heavy mero-myosin (HMM) motor fragments immobilized on various surfaces, is altered by the application of electrical loads generated by an external electric field with strengths ranging from 0 to 8 kVm(-1). Because the motility is intimately linked to the function of surface-immobilized motors, the study also showed how the adsorption properties of HMM on various surfaces, such as nitrocellulose (NC), trimethylclorosilane (TMCS), poly(methyl methacrylate) (PMMA), poly(tert-butyl methacrylate) (PtBMA), and poly(butyl methacrylate) (PBMA), can be characterized using an external field. It was found that at an electric field of 5 kVm(-1) the force exerted on the filaments is sufficient to overcome the frictionlike resistive force of the inactive motors. It was also found that the effect of assisting electric fields on the relative increase in the sliding velocity was markedly higher for the TMCS-derivatized surface than for all other polymer-based surfaces. An explanation of this behavior, based on the molecular rigidity of the TMCS-on-glass surfaces as opposed to the flexibility of the polymer-based ones, is considered. To this end, the proposed microfluidic device could be used to select appropriate surfaces for future lab-on-a-chip applications as illustrated here for the almost ideal TMCS surface. Furthermore, the proposed methodology can be used to gain fundamental insights into the functioning of protein molecular motors, such as the force exerted by the motors under different operational conditions.

  • 249.
    Verardo, Damiano
    et al.
    Lund University, Sweden.
    Lindberg, Frida W.
    Lund University, Sweden.
    Anttu, Nicklas
    Lund University, Sweden;AstraZeneca, Sweden.
    Niman, Cassandra S.
    Lund University, Sweden;University of California San Diego, USA.
    Lard, Mercy
    Lund University, Sweden.
    Dabkowska, Aleksandra P.
    Lund University, Sweden;Aalto University, Finland.
    Nylander, Tommy
    Lund University, Sweden.
    Månsson, Alf
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences. Lund University, Sweden.
    Prinz, Christelle N.
    Lund University, Sweden.
    Linke, Heiner
    Lund University, Sweden.
    Nanowires for Biosensing: Lightguiding of Fluorescence as a Function of Diameter and Wavelength2018In: Nano letters (Print), ISSN 1530-6984, E-ISSN 1530-6992, Vol. 18, no 8, p. 4796-4802Article in journal (Refereed)
    Abstract [en]

    Semiconductor nanowires can act as nanoscaled optical fibers, enabling them to guide and concentrate light emitted by surface-bound fluorophores, potentially enhancing the sensitivity of optical biosensing. While parameters such as the nanowire geometry and the fluorophore wavelength can be expected to strongly influence this lightguiding effect, no detailed description of their effect on in-coupling of fluorescent emission is available to date. Here, we use confocal imaging to quantify the lightguiding effect in GaP nanowires as a function of nanowire geometry and light wavelength. Using a combination of finite-difference time-domain simulations and analytical approaches, we identify the role of multiple waveguide modes for the observed lightguiding. The normalized frequency parameter, based on the step-index approximation, predicts the lightguiding ability of the nanowires as a function of diameter and fluorophore wavelength, providing a useful guide for the design of optical biosensors based on nanowires.

  • 250. Villegas, Mirza
    et al.
    Brodelius, Peter
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Elicitor-Induced Hydroxycinnamoyl-CoA:tyramine Hydroxycinna-moyltransferase in Plant Cell Suspension Cultures1990In: Physiologia Plantarum: An International Journal for Plant Biology, ISSN 0031-9317, E-ISSN 1399-3054, Vol. 78, p. 414-420Article in journal (Refereed)
23456 201 - 250 of 264
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