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  • 1. Andersen, Gorm
    et al.
    Björnberg, Olof
    Polakova, Silvia
    Pynyaha, Yuriy
    Rasmussen, Anna
    Møller, Kasper
    Hofer, Anders
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Moritz, Thomas
    Sandrini, Michael Paolo Bastner
    Merico, Anna-Maria
    Compagno, Concetta
    Akerlund, Hans-Erik
    Gojković, Zoran
    Piskur, Jure
    A second pathway to degrade pyrimidine nucleic acid precursors in eukaryotes.2008In: Journal of Molecular Biology, ISSN 1089-8638, Vol. 380, no 4, p. 656-66Article in journal (Other academic)
  • 2. Andersson, J
    et al.
    Westman, M
    Hofer, Anders
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Sjoberg, B M
    Allosteric regulation of the class III anaerobic ribonucleotide reductase from bacteriophage T4.2000In: Journal of Biological Chemistry, ISSN 0021-9258, Vol. 275, no 26, p. 19443-8Article in journal (Refereed)
  • 3. Benelli, Giovanni
    et al.
    Maggi, Filippo
    Pavela, Roman
    Murugan, Kadarkarai
    Govindarajan, Marimuthu
    Vaseeharan, Baskaralingam
    Petrelli, Riccardo
    Cappellacci, Loredana
    Kumar, Suresh
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Youssefi, Mohammad Reza
    Alarfaj, Abdullah A.
    Hwang, Jiang-Shiou
    Higuchi, Akon
    Mosquito control with green nanopesticides: towards the One Health approach? A review of non-target effects2018In: Environmental science and pollution research international, ISSN 0944-1344, E-ISSN 1614-7499, Vol. 25, no 11, p. 10184-10206Article, review/survey (Refereed)
    Abstract [en]

    The rapid spread of highly aggressive arboviruses, parasites, and bacteria along with the development of resistance in the pathogens and parasites, as well as in their arthropod vectors, represents a huge challenge in modern parasitology and tropical medicine. Eco-friendly vector control programs are crucial to fight, besides malaria, the spread of dengue, West Nile, chikungunya, and Zika virus, as well as other arboviruses such as St. Louis encephalitis and Japanese encephalitis. However, research efforts on the control of mosquito vectors are experiencing a serious lack of eco-friendly and highly effective pesticides, as well as the limited success of most biocontrol tools currently applied. Most importantly, a cooperative interface between the two disciplines is still lacking. To face this challenge, we have reviewed a wide number of promising results in the field of green-fabricated pesticides tested against mosquito vectors, outlining several examples of synergy with classic biological control tools. The non-target effects of green-fabricated nanopesticides, including acute toxicity, genotoxicity, and impact on behavioral traits of mosquito predators, have been critically discussed. In the final section, we have identified several key challenges at the interface between "green" nanotechnology and classic biological control, which deserve further research attention.

  • 4. Björnberg, O
    et al.
    Vodnala, Munender
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Domkin, Vladimir
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Rasmussen, A
    Andersen, G
    Piskur, J
    Ribosylurea accumulates in yeast urc4 mutants2010In: Nucleosides, Nucleotides & Nucleic Acids, ISSN 1525-7770, E-ISSN 1532-2335, Vol. 29, no 4-6, p. 433-437Article in journal (Refereed)
    Abstract [en]

    Yeast Saccharomyces (Lachancea) kluyveri urc4 mutants, unable to grow on uracil, biotransformed (14)C(2)-uracil into two labeled compounds, as detected by high performance liquid chromatography (HPLC). These two compounds could also be obtained following organic synthesis of ribosylurea. This finding demonstrates that in the URC pyrimidine degradation pathway, the opening of the uracil ring takes place when uracil is attached to the ribose moiety. Ribosylurea has not been reported in the cell metabolism before and the two observed compounds likely represent an equilibrium mixture of the pyranosyl and furanosyl forms.

  • 5.
    Bugaytsova, Jeanna A.
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Björnham, Oscar
    Umeå University, Faculty of Science and Technology, Department of Applied Physics and Electronics. Swedish Defence Research Agency, 906 21 Umeå, Sweden.
    Chernov, Yevgen A.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Gideonsson, Pär
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Henriksson, Sara
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Mendez, Melissa
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Sjöström, Rolf
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Mahdavi, Jafar
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. School of Life Sciences, CBS, University of Nottingham, NG7 2RD Nottingham, UK.
    Shevtsova, Anna
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Ilver, Dag
    Moonens, Kristof
    Quintana-Hayashi, Macarena P.
    Moskalenko, Roman
    Aisenbrey, Christopher
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Bylund, Göran
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Schmidt, Alexej
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Åberg, Anna
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Brännström, Kristoffer
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Koeniger, Verena
    Vikström, Susanne
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Rakhimova, Lena
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Ögren, Johan
    Umeå University, Faculty of Medicine, Department of Public Health and Clinical Medicine, Section of Medicine.
    Liu, Hui
    Goldman, Matthew D.
    Whitmire, Jeannette M.
    Åden, Jörgen
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Younson, Justine
    Kelly, Charles G.
    Gilman, Robert H.
    Chowdhury, Abhijit
    Mukhopadhyay, Asish K.
    Nair, G. Balakrish
    Papadakos, Konstantinos S.
    Martinez-Gonzalez, Beatriz
    Sgouras, Dionyssios N.
    Engstrand, Lars
    Unemo, Magnus
    Danielsson, Dan
    Suerbaum, Sebastian
    Oscarson, Stefan
    Morozova-Roche, Ludmilla A.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Olofsson, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Gröbner, Gerhard
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Holgersson, Jan
    Esberg, Anders
    Umeå University, Faculty of Medicine, Department of Odontology.
    Strömberg, Nicklas
    Umeå University, Faculty of Medicine, Department of Odontology.
    Landström, Maréne
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Eldridge, Angela M.
    Chromy, Brett A.
    Hansen, Lori M.
    Solnick, Jay V.
    Linden, Sara K.
    Haas, Rainer
    Dubois, Andre
    Merrell, D. Scott
    Schedin, Staffan
    Umeå University, Faculty of Science and Technology, Department of Applied Physics and Electronics.
    Remaut, Han
    Arnqvist, Anna
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Berg, Douglas E.
    Boren, Thomas
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Helicobacter pylori Adapts to Chronic Infection and Gastric Disease via pH-Responsive BabA-Mediated Adherence2017In: Cell Host and Microbe, ISSN 1931-3128, E-ISSN 1934-6069, Vol. 21, no 3, p. 376-389Article in journal (Refereed)
    Abstract [en]

    The BabA adhesin mediates high-affinity binding of Helicobacter pylori to the ABO blood group antigen-glycosylated gastric mucosa. Here we show that BabA is acid responsive-binding is reduced at low pH and restored by acid neutralization. Acid responsiveness differs among strains; often correlates with different intragastric regions and evolves during chronic infection and disease progression; and depends on pH sensor sequences in BabA and on pH reversible formation of high-affinity binding BabA multimers. We propose that BabA's extraordinary reversible acid responsiveness enables tight mucosal bacterial adherence while also allowing an effective escape from epithelial cells and mucus that are shed into the acidic bactericidal lumen and that bio-selection and changes in BabA binding properties through mutation and recombination with babA-related genes are selected by differences among individuals and by changes in gastric acidity over time. These processes generate diverse H. pylori subpopulations, in which BabA's adaptive evolution contributes to H. pylori persistence and overt gastric disease.

  • 6.
    Bugaytsova, Jeanna
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Chernov, Yevgen A
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Gideonsson, Pär
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Mendez, Melissa
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Henriksson, Sara
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Mahdavi, Jafar
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. School of Life Sciences, CBS, University of Nottingham, Nottingham, UK..
    Quintana-Hayashi, Macarena
    Department of Biochemistry and Cell biology, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden..
    Shevtsova, Anna
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Sjöström, Rolf
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Moskalenko, Roman
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Department of Pathology, Medical Institute, State University, Sumy, Ukraine.
    Aisenbrey, Christopher
    Umeå University, Faculty of Science and Technology, Department of Chemistry. Université de Strasbourg, Institut de Chimie, Strasbourg, France.
    Moonens, Kristof
    Structural and Molecular Microbiology, VIB Department of Structural Biology, Belgium.
    Björnham, Oscar
    Umeå University, Faculty of Science and Technology, Department of Applied Physics and Electronics. FOI Totalförsvarets Forskningsinstitut, Umeå, Sweden..
    Brännström, Kristoffer
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Bylund, Göran
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Königer, Verena
    Max von Pettenkofer Institute of Hygiene and Medical Microbiology, LMU, Munich, Germany.
    Vikström, Susanne
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Schmidt, Alexej
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Rakhimova, Lena
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Ögren, Johan
    Umeå University, Faculty of Medicine, Department of Odontology.
    Ilver, Dag
    Department of Biochemistry and Cell biology, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
    Liu, Hui
    Department of Medicine, USUHS, Bethesda, MD, USA.
    Goldman, Matthew
    Department of Pediatrics, USUHS, Bethesda, MD, USA.
    Whitmire, Jeannette M
    Department of Microbiology and Immunology, USUHS, Bethesda, MD USA.
    Kelly, Charles G
    King's College London, Dental Institute, London, UK.
    Gilman, Robert H
    Department of International Health, John Hopkins School of Public Health, Baltimore, MD, USA.
    Chowdhury, Abhijit
    Centre for Liver Research, School of Digestive and Liver Diseases, Institute of Post Graduate Medical Education & Research, Kolkata, India.
    Mukhopadhyay, Asish K
    Division of Bacteriology, National Institute of Cholera and Enteric Diseases, Kolkata, India.
    Nair, Balakrish G
    Translational Health Science and Technology Institute, Haryana, India.
    Papadakos, Konstantinos S
    Hellenic Pasteur Institute, Athens, Greece.
    Martinez-Gonzalez, Beatriz
    Hellenic Pasteur Institute, Athens, Greece.
    Sgouras, Dionyssios N
    Hellenic Pasteur Institute, Athens, Greece.
    Engstrand, Lars
    Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden.
    Unemo, Magnus
    Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Danielsson, Dan
    Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Sebastian, Suerbaum
    Institute for Medical Microbiology and Hospital Epidemiology Hannover Medical School, Hannover, Germany.
    Oscarson, Stefan
    Centre for Synthesis and Chemical Biology, School of Chemistry, University College Dublin, Dublin, Ireland.
    Morozova-Roche, Ludmilla
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Gröbner, Gerhard
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Holgersson, Jan
    Department of Clinical Chemistry and Transfusion Medicine, Sahlgrenska Academy, University of Gothenburg, Sahlgrenska University Hospital, Gothenburg, Sweden.
    Strömberg, Nicklas
    Umeå University, Faculty of Medicine, Department of Odontology.
    Esberg, Anders
    Umeå University, Faculty of Medicine, Department of Odontology.
    Eldridge, Angela
    Department of Pathology and Laboratory Medicine, University of California Davis School of Medicine, Sacramento, CA, USA.
    Chromy, Brett A
    Department of Pathology and Laboratory Medicine, University of California Davis School of Medicine, Sacramento, CA, USA.
    Hansen, Lori
    Departments of Medical Microbiology and Immunology, Center for Comparative Medicine, University of California Davis, Davis, CA, USA.
    Solnick, Jay
    Departments of Medical Microbiology and Immunology, Center for Comparative Medicine, University of California Davis, Davis, CA, USA.
    Haas, Rainer
    Max von Pettenkofer Institute of Hygiene and Medical Microbiology, LMU Munich, Munich, Germany.
    Schedin, Staffan
    Umeå University, Faculty of Science and Technology, Department of Applied Physics and Electronics.
    Lindén, Sara K
    Department of Biochemistry and Cell biology, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
    Dubois, Andre
    Department of Medicine, USUHS, Bethesda, MD, USA.
    Merrell, D. Scott
    Department of Microbiology and Immunology, USUHS, Bethesda, MD, USA.
    Remaut, Han
    Structural and Molecular Microbiology, VIB Department of Structural Biology, VIB, Brussels, Belgium.
    Arnqvist, Anna
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Berg, Douglas E
    Department of Medicine, University of California San Diego, La Jolla, CA, USA.
    Borén, Thomas
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Acid Responsive Helicobacter pylori Adherence: Implications for Chronic Infection and DiseaseManuscript (preprint) (Other academic)
  • 7. Chiruvella, Kishore K
    et al.
    Rajaei, Naghmeh
    Jonna, Venkateswara Rao
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Åstrom, Stefan U
    Biochemical Characterization of Kat1: a Domesticated hAT-Transposase that Induces DNA Hairpin Formation and MAT-Switching2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, article id 21671Article in journal (Refereed)
    Abstract [en]

    Kluyveromyces lactis hAT-transposase 1 (Kat1) generates hairpin-capped DNA double strand breaks leading to MAT-switching (MATa to MAT alpha). Using purified Kat1, we demonstrate the importance of terminal inverted repeats and subterminal repeats for its endonuclease activity. Kat1 promoted joining of the transposon end into a target DNA molecule in vitro, a biochemical feature that ties Kat1 to transposases. Gas-phase Electrophoretic Mobility Macromolecule analysis revealed that Kat1 can form hexamers when complexed with DNA. Kat1 point mutants were generated in conserved positions to explore structure-function relationships. Mutants of predicted catalytic residues abolished both DNA cleavage and strand-transfer. Interestingly, W576A predicted to be impaired for hairpin formation, was active for DNA cleavage and supported wild type levels of mating-type switching. In contrast, the conserved CXXH motif was critical for hairpin formation because Kat1 C402A/H405A completely blocked hairpinning and switching, but still generated nicks in the DNA. Mutations in the BED zinc-finger domain (C130A/C133A) resulted in an unspecific nuclease activity, presumably due to nonspecific DNA interaction. Kat1 mutants that were defective for cleavage in vitro were also defective for mating-type switching. Collectively, this study reveals Kat1 sharing extensive biochemical similarities with cut and paste transposons despite being domesticated and evolutionary diverged from active transposons.

  • 8. Crona, Mikael
    et al.
    Codo, Paula
    Jonna, Venkateswara Rao
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Fernandes, Aristi P.
    Tholander, Fredrik
    A ribonucleotide reductase inhibitor with deoxyribonucleoside-reversible cytotoxicity2016In: Molecular Oncology, ISSN 1574-7891, E-ISSN 1878-0261, Vol. 10, no 9, p. 1375-1386Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide Reductase (RNR) is the sole enzyme that catalyzes the reduction of ribonucleotides into deoxyribonucleotides. Even though RNR is a recognized target for antiproliferative molecules, and the main target of the approved drug hydroxyurea, few new leads targeted to this enzyme have been developed. We have evaluated a recently identified set of RNR inhibitors with respect to inhibition of the human enzyme and cellular toxicity. One compound, NSC73735, is particularly interesting; it is specific for leukemia cells and is the first identified compound that hinders oligomerization of the mammalian large RNR subunit. Similar to hydroxyurea, it caused a disruption of the cell cycle distribution of cultured HL-60 cells. In contrast to hydroxyurea, the disruption was reversible, indicating higher specificity. NSC73735 thus defines a potential lead candidate for RNR-targeted anticancer drugs, as well as a chemical probe with better selectivity for RNR inhibition than hydroxyurea. 

  • 9. Crona, Mikael
    et al.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Astorga-Wells, Juan
    Sjöberg, Britt-Marie
    Tholander, Fredrik
    Biochemical Characterization of the Split Class II Ribonucleotide Reductase from Pseudomonas aeruginosa2015In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, no 7, article id e0134293Article in journal (Refereed)
    Abstract [en]

    The opportunistic pathogen Pseudomonas aeruginosa can grow under both aerobic and anaerobic conditions. Its flexibility with respect to oxygen load is reflected by the fact that its genome encodes all three existing classes of ribonucleotides reductase (RNR): the oxygen-dependent class I RNR, the oxygen-indifferent class II RNR, and the oxygen-sensitive class III RNR. The P. aeruginosa class II RNR is expressed as two separate polypeptides (NrdJa and NrdJb), a unique example of a split RNR enzyme in a free-living organism. A split class II RNR is also found in a few closely related gamma-Proteobacteria. We have characterized the P. aeruginosa class II RNR and show that both subunits are required for formation of a biologically functional enzyme that can sustain vitamin B12-dependent growth. Binding of the B12 coenzyme as well as substrate and allosteric effectors resides in the NrdJa subunit, whereas the NrdJb subunit mediates efficient reductive dithiol exchange during catalysis. A combination of activity assays and activity-independent methods like surface plasmon resonance and gas phase electrophoretic macromolecule analysis suggests that the enzymatically active form of the enzyme is a (NrdJa-NrdJb) 2 homodimer of heterodimers, and a combination of hydrogen-deuterium exchange experiments and molecular modeling suggests a plausible region in NrdJa that interacts with NrdJb. Our detailed characterization of the split NrdJ from P. aeruginosa provides insight into the biochemical function of a unique enzyme known to have central roles in biofilm formation and anaerobic growth.

  • 10. Crona, Mikael
    et al.
    Moffatt, Connor
    Friedrich, Nancy C
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Sjöberg, Britt-Marie
    Edgell, David R
    Assembly of a fragmented ribonucleotide reductase by protein interaction domains derived from a mobile genetic element.2011In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 39, no 4, p. 1381-9Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductase (RNR) is a critical enzyme of nucleotide metabolism, synthesizing precursors for DNA replication and repair. In prokaryotic genomes, RNR genes are commonly targeted by mobile genetic elements, including free standing and intron-encoded homing endonucleases and inteins. Here, we describe a unique molecular solution to assemble a functional product from the RNR large subunit gene, nrdA that has been fragmented into two smaller genes by the insertion of mobE, a mobile endonuclease. We show that unique sequences that originated during the mobE insertion and that are present as C- and N-terminal tails on the split NrdA-a and NrdA-b polypeptides, are absolutely essential for enzymatic activity. Our data are consistent with the tails functioning as protein interaction domains to assemble the tetrameric (NrdA-a/NrdA-b)(2) large subunit necessary for a functional RNR holoenzyme. The tails represent a solution distinct from RNA and protein splicing or programmed DNA rearrangements to restore function from a fragmented coding region and may represent a general mechanism to neutralize fragmentation of essential genes by mobile genetic elements.

  • 11. Crona, Mikael
    et al.
    Torrents, Eduard
    Rohr, Asmund K.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Furrer, Ernst
    Tomter, Ane B.
    Andersson, K. Kristoffer
    Sahlin, Margareta
    Sjoberg, Britt-Marie
    NrdH-Redoxin Protein Mediates High Enzyme Activity in Manganese-reconstituted Ribonucleotide Reductase from Bacillus anthracis2011In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 286, no 38, p. 33053-33060Article in journal (Refereed)
    Abstract [en]

    Bacillus anthracis is a severe mammalian pathogen encoding a class Ib ribonucleotide reductase (RNR). RNR is a universal enzyme that provides the four essential deoxyribonucleotides needed for DNA replication and repair. Almost all Bacillus spp. encode both class Ib and class III RNR operons, but the B. anthracis class III operon was reported to encode a pseudogene, and conceivably class Ib RNR is necessary for spore germination and proliferation of B. anthracis upon infection. The class Ib RNR operon in B. anthracis encodes genes for the catalytic NrdE protein, the tyrosyl radical metalloprotein NrdF, and the flavodoxin protein NrdI. The tyrosyl radical in NrdF is stabilized by an adjacent Mn(2)(III) site (Mn-NrdF) formed by the action of the NrdI protein or by a Fe(2)(III) site (Fe-NrdF) formed spontaneously from Fe(2+) and O(2). In this study, we show that the properties of B. anthracis Mn-NrdF and Fe-NrdF are in general similar for interaction with NrdE and NrdI. Intriguingly, the enzyme activity of Mn-NrdF was approximately an order of magnitude higher than that of Fe-NrdF in the presence of the class Ib-specific physiological reductant NrdH, strongly suggesting that the Mn-NrdF form is important in the life cycle of B. anthracis. Whether the Fe-NrdF form only exists in vitro or whether the NrdF protein in B. anthracis is a true cambialistic enzyme that can work with either manganese or iron remains to be established.

  • 12.
    Decker, Daniel
    et al.
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology. Umeå University, Faculty of Science and Technology, Umeå Plant Science Centre (UPSC).
    Meng, Meng
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology. Umeå University, Faculty of Science and Technology, Umeå Plant Science Centre (UPSC).
    Gornicka, Agnieszka
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology. Umeå University, Faculty of Science and Technology, Umeå Plant Science Centre (UPSC).
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Wilczynska, Malgorzata
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Kleczkowski, Leszek A
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology. Umeå University, Faculty of Science and Technology, Umeå Plant Science Centre (UPSC).
    Substrate kinetics and substrate effects on the quaternary structure of barley UDP-glucose pyrophosphorylase2012In: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 79, p. 39-45Article in journal (Refereed)
    Abstract [en]

    UDP-Glc pyrophosphorylase (UGPase) is an essential enzyme responsible for production of UDP-Glc, which is used in hundreds of glycosylation reactions involving addition of Glc to a variety of compounds. In this study, barley UGPase was characterized with respect to effects of its substrates on activity and quaternary structure of the protein. Its K(m) values with Glc-1-P and UTP were 0.33 and 0.25 mM, respectively. Besides using Glc-1-P as a substrate, the enzyme had also considerable activity with Gal-1-P; however, the K(m) for Gal-1-P was very high (>10 mM), rendering this reaction unlikely under physiological conditions. UGPase had a relatively broad pH optimum of 6.5-8.5, regardless of the direction of reaction. The enzyme equilibrium constant was 0.4, suggesting slight preference for the Glc-1-P synthesis direction of the reaction. The quaternary structure of the enzyme, studied by Gas-phase Electrophoretic Mobility Macromolecule Analysis (GEMMA), was affected by addition of either single or both substrates in either direction of the reaction, resulting in a shift from UGPase dimers toward monomers, the active form of the enzyme. The substrate-induced changes in quaternary structure of the enzyme may have a regulatory role to assure maximal activity. Kinetics and factors affecting the oligomerization status of UGPase are discussed.

  • 13. Elmlund, Hans
    et al.
    Baraznenok, Vera
    Linder, Tomas
    Szilagyi, Zsolt
    Rofougaran, Reza
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hebert, Hans
    Lindahl, Martin
    Gustafsson, Claes M
    Cryo-EM reveals promoter DNA binding and conformational flexibility of the general transcription factor TFIID2009In: Structure, ISSN 0969-2126, E-ISSN 1878-4186, Vol. 17, no 11, p. 1442-1452Article in journal (Refereed)
    Abstract [en]

    The general transcription factor IID (TFIID) is required for initiation of RNA polymerase II-dependent transcription at many eukaryotic promoters. TFIID comprises the TATA-binding protein (TBP) and several conserved TBP-associated factors (TAFs). Recognition of the core promoter by TFIID assists assembly of the preinitiation complex. Using cryo-electron microscopy in combination with methods for ab initio single-particle reconstruction and heterogeneity analysis, we have produced density maps of two conformational states of Schizosaccharomyces pombe TFIID, containing and lacking TBP. We report that TBP-binding is coupled to a massive histone-fold domain rearrangement. Moreover, docking of the TBP-TAF1(N-terminus) atomic structure to the TFIID map and reconstruction of a TAF-promoter DNA complex helps to account for TAF-dependent regulation of promoter-TBP and promoter-TAF interactions.

  • 14. Falahati, Hanieh
    et al.
    Pazhang, Mohammad
    Zareian, Shekufeh
    Ghaemi, Nasser
    Rofougaran, Reza
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Rezaie, Alireza R
    Khajeh, Khosro
    Transmitting the allosteric signal in methylglyoxal synthase2013In: Protein Engineering Design & Selection, ISSN 1741-0126, E-ISSN 1741-0134, Vol. 26, no 7, p. 445-452Article in journal (Refereed)
    Abstract [en]

    The homohexameric enzyme methylglyoxal synthase (MGS) converts dihydroxyacetone phosphate (DHAP) to methylglyoxal and phosphate. This enzyme is allosterically inhibited by phosphate. The allosteric signal induced by phosphate in MGS from Thermus sp. GH5 (TMGS) has been tracked by site-directed mutagenesis, from the binding site of phosphate to the pathways that transmit the signal, and finally to the active site which is the receiver of the signal. In TMGS, Ser-55 distinguishes the inhibitory phosphate from the phosphoryl group of the substrate, DHAP, and transmits the allosteric signal through Pro-82, Arg-97 and Val-101 to the active site. Furthermore, the addition of a C-terminal tail to TMGS reinforces the allosteric signal by introducing a new salt bridge between Asp-10 and an Arg in this tail. Lastly, the active site amino acid, Gly-56, is shown to be involved in both allostery and phosphate elimination step from DHAP by TMGS. Interestingly, some of the mutations also trigger homotropic allostery, supporting the hypothesis that allostery is an intrinsic property of all dynamic proteins. The details of the TMGS allosteric network discussed in this study can serve as a model system for understanding the enigmatic allosteric mechanism of other proteins.

  • 15. Farge, Géraldine
    et al.
    Holmlund, Teresa
    Khvorostova, Julia
    Rofougaran, Reza
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Falkenberg, Maria
    The N-terminal domain of TWINKLE contributes to single-stranded DNA binding and DNA helicase activities.2008In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 36, no 2, p. 393-403Article in journal (Refereed)
  • 16.
    Fijolek, Artur
    et al.
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Thelander, Lars
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Expression, purification, characterization, and in vivo targeting of trypanosome CTP synthetase for treatment of African sleeping sickness.2007In: Journal of biological chemistry, ISSN 0021-9258, Vol. 282, no 16, p. 11858-11865Article in journal (Refereed)
  • 17. Grinberg, Inna
    et al.
    McGann, Matthew
    Lundin, Daniel
    Crona, Mikael
    Hasan, Mahmudal
    Jonna, Venkateswara Rao
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Loderer, Christoph
    Sahlin, Margareta
    Markova, Natalia
    Stenson, John
    Borovok, Ilya
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Logan, Derek
    Sjöberg, Britt-Marie
    Novel ATP-Cone-Driven Allosteric Regulation of Ribonucleotide Reductase Via the Radical-Generating Subunit2018In: Protein Science, ISSN 0961-8368, E-ISSN 1469-896X, Vol. 27, p. 87-88Article in journal (Other academic)
  • 18.
    Gupta, Arun A.
    et al.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Reinartz, Ines
    Karunanithy, Gogulan
    Spilotros, Alessandro
    Jonna, Venkateswara Rao
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Svergun, Dmitri I.
    Baldwin, Andrew J.
    Schug, Alexander
    Wolf-Watz, Magnus
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Formation of a Secretion-Competent Protein Complex by a Dynamic Wrap-around Binding Mechanism2018In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 430, no 18, Part B, p. 3157-3169Article in journal (Refereed)
    Abstract [en]

    Bacterial virulence is typically initiated by translocation of effector or toxic proteins across host cell membranes. A class of gram-negative pathogenic bacteria including Yersinia pseudotuberculosis and Yersinia pestis accomplishes this objective with a protein assembly called the type III secretion system. Yersinia effector proteins (Yop) are presented to the translocation apparatus through formation of specific complexes with their cognate chaperones (Syc). In the complexes where the structure is available, the Yops are extended and wrap around their cognate chaperone. This structural architecture enables secretion of the Yop from the bacterium in early stages of translocation. It has been shown previously that the chaperone-binding domain of YopE is disordered in its isolation but becomes substantially more ordered in its wrap-around complex with its chaperone SycE. Here, by means of NMR spectroscopy, small-angle X-ray scattering and molecular modeling, we demonstrate that while the free chaperone-binding domain of YopH (YopHCBD) adopts a fully ordered and globular fold, it populates an elongated, wrap-around conformation when it engages in a specific complex with its chaperone SycH2. Hence, in contrast to YopE that is unstructured in its free state, YopH transits from a globular free state to an elongated chaperone-bound state. We demonstrate that a sparsely populated YopHCBD state has an elevated affinity for SycH2 and represents an intermediate in the formation of the protein complex. Our results suggest that Yersinia has evolved a binding mechanism where SycH2 passively stimulates an elongated YopH conformation that is presented to the type III secretion system in a secretion-competent conformation.

  • 19.
    Gupta, Arun
    et al.
    Umeå University.
    Reinartz, Ines
    Spilotros, Alessandro
    Jonna, Venkateswara R.
    Umeå University.
    Hofer, Anders
    Umeå University.
    Svergun, Dmitri I.
    Schug, Alexander
    Wolf-Watz, Magnus
    Umeå University.
    Global Disordering in Stereo-Specific Protein Association2017In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 112, no 3, p. 33A-33AArticle in journal (Refereed)
  • 20. Hayakawa, H
    et al.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Thelander, Lars
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Kitajima, S
    Cai, Y
    Oshiro, S
    Yakushiji, H
    Nakabeppu, Y
    Kuwano, M
    Sekiguchi, M
    Metabolic fate of oxidized guanine ribonucleotides in mammalian cells.1999In: Biochemistry, ISSN 0006-2960, Vol. 38, no 12, p. 3610-4Article in journal (Refereed)
  • 21.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Nucleotide Metabolism of the Human Pathogen Trypanosoma brucei2000Doctoral thesis, comprehensive summary (Other academic)
  • 22.
    Hofer, Anders
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Crona, Mikael
    Logan, Derek T
    Sjöberg, Britt-Marie
    DNA building blocks: keeping control of manufacture2012In: Critical reviews in biochemistry and molecular biology, ISSN 1040-9238, E-ISSN 1549-7798, Vol. 47, no 1, p. 50-63Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductase (RNR) is the only source for de novo production of the four deoxyribonucleoside triphosphate (dNTP) building blocks needed for DNA synthesis and repair. It is crucial that these dNTP pools are carefully balanced, since mutation rates increase when dNTP levels are either unbalanced or elevated. RNR is the major player in this homeostasis, and with its four different substrates, four different allosteric effectors and two different effector binding sites, it has one of the most sophisticated allosteric regulations known today. In the past few years, the structures of RNRs from several bacteria, yeast and man have been determined in the presence of allosteric effectors and substrates, revealing new information about the mechanisms behind the allosteric regulation. A common theme for all studied RNRs is a flexible loop that mediates modulatory effects from the allosteric specificity site (s-site) to the catalytic site for discrimination between the four substrates. Much less is known about the allosteric activity site (a-site), which functions as an on-off switch for the enzyme's overall activity by binding ATP (activator) or dATP (inhibitor). The two nucleotides induce formation of different enzyme oligomers, and a recent structure of a dATP-inhibited α(6)β(2) complex from yeast suggested how its subunits interacted non-productively. Interestingly, the oligomers formed and the details of their allosteric regulation differ between eukaryotes and Escherichia coli. Nevertheless, these differences serve a common purpose in an essential enzyme whose allosteric regulation might date back to the era when the molecular mechanisms behind the central dogma evolved.

  • 23.
    Hofer, Anders
    et al.
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Ekanem, J T
    Thelander, Lars
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Allosteric regulation of Trypanosoma brucei ribonucleotide reductase studied in vitro and in vivo.1998In: Journal of Biological Chemistry, ISSN 0021-9258, Vol. 273, no 51, p. 34098-104Article in journal (Refereed)
  • 24.
    Hofer, Anders
    et al.
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Schmidt, P P
    Gräslund, Astrid
    Thelander, Lars
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Cloning and characterization of the R1 and R2 subunits of ribonucleotide reductase from Trypanosoma brucei.1997In: Proceedings of the National Academy of Sciences of the U S A, ISSN 0027-8424, Vol. 94, no 13, p. 6959-64Article in journal (Refereed)
  • 25.
    Hofer, Anders
    et al.
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Steverding, D
    Chabes, Andrei
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Brun, R
    Thelander, Lars
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Trypanosoma brucei CTP synthetase: a target for the treatment of African sleeping sickness.2001In: Proceedings of the National Academy of Sciences of the U S A, ISSN 0027-8424, Vol. 98, no 11, p. 6412-6Article in journal (Refereed)
  • 26.
    Hosseinzadeh, Ava
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Rofougaran, Reza
    Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Clinical Neuroscience.
    Vodnala, Munender
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Kötemann, A
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Urban, Constantin F.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Adenosine is a drugable negative regulator of neutrophil activity during Candida albicans infectionManuscript (preprint) (Other academic)
  • 27.
    Håkansson, Pelle
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Thelander, Lars
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Regulation of mammalian ribonucleotide reduction and dNTP pools after DNA damage and in resting cells.2006In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 281, no 12, p. 7834-7841Article in journal (Refereed)
  • 28. Johansson, Renzo
    et al.
    Jonna, Venkateswara Rao
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Kumar, Rohit
    Nayeri, Niloofar
    Lundin, Daniel
    Sjöberg, Britt-Marie
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Logan, Derek T
    Structural Mechanism of Allosteric Activity Regulation in a Ribonucleotide Reductase with Double ATP Cones2016In: Structure, ISSN 0969-2126, E-ISSN 1878-4186, Vol. 24, no 6, p. 906-917Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductases (RNRs) reduce ribonucleotides to deoxyribonucleotides. Their overall activity is stimulated by ATP and downregulated by dATP via a genetically mobile ATP cone domain mediating the formation of oligomeric complexes with varying quaternary structures. The crystal structure and solution X-ray scattering data of a novel dATP-induced homotetramer of the Pseudomonas aeruginosa class I RNR reveal the structural bases for its unique properties, namely one ATP cone that binds two dATP molecules and a second one that is non-functional, binding no nucleotides. Mutations in the observed tetramer interface ablate oligomerization and dATP-induced inhibition but not the ability to bind dATP. Sequence analysis shows that the novel type of ATP cone may be widespread in RNRs. The present study supports a scenario in which diverse mechanisms for allosteric activity regulation are gained and lost through acquisition and evolutionary erosion of different types of ATP cone.

  • 29.
    Jonna, Venkateswara Rao
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Crona, Mikael
    Rofougaran, Reza
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Lundin, Daniel
    Johansson, Samuel
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Brännström, Kristoffer
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Sjöberg, Britt-Marie
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Diversity in Overall Activity Regulation of Ribonucleotide Reductase2015In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 290, no 28, p. 17339-17348Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductase (RNR) catalyzes the reduction of ribonucleotides to the corresponding deoxyribonucleotides, which are used as building blocks for DNA replication and repair. This process is tightly regulated via two allosteric sites, the specificity site (s-site) and the overall activity site (a-site). The a-site resides in an N-terminal ATP cone domain that binds dATP or ATP and functions as an on/off switch, whereas the composite s-site binds ATP, dATP, dTTP, or dGTP and determines which substrate to reduce. There are three classes of RNRs, and class I RNRs consist of different combinations of α and β subunits. In eukaryotic and Escherichia coli class I RNRs, dATP inhibits enzyme activity through the formation of inactive α6 and α4β4 complexes, respectively. Here we show that the Pseudomonas aeruginosa class I RNR has a duplicated ATP cone domain and represents a third mechanism of overall activity regulation. Each α polypeptide binds three dATP molecules, and the N-terminal ATP cone is critical for binding two of the dATPs because a truncated protein lacking this cone could only bind dATP to its s-site. ATP activates the enzyme solely by preventing dATP from binding. The dATP-induced inactive form is an α4 complex, which can interact with β2 to form a non-productive α4β2 complex. Other allosteric effectors induce a mixture of α2 and α4 forms, with the former being able to interact with β2 to form active α2β2 complexes. The unique features of the P. aeruginosa RNR are interesting both from evolutionary and drug discovery perspectives.

  • 30. Kamte, Stephane L. Ngahang
    et al.
    Ranjbarian, Farahnaz
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Campagnaro, Gustavo Daniel
    Nya, Prosper C. Biapa
    Mbuntcha, Hélène
    Woguem, Verlaine
    Womeni, Hilaire Macaire
    Tapondjou, Léon Azefack
    Giordani, Cristiano
    Barboni, Luciano
    Benelli, Giovanni
    Cappellacci, Loredana
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Petrelli, Riccardo
    Maggi, Filippo
    Trypanosoma brucei Inhibition by Essential Oils from Medicinal and Aromatic Plants Traditionally Used in Cameroon (Azadirachta indica, Aframomum melegueta, Aframomum daniellii, Clausena anisata, Dichrostachys cinerea and Echinops giganteus)2017In: International Journal of Environmental Research and Public Health, ISSN 1661-7827, E-ISSN 1660-4601, Vol. 14, no 7, article id 737Article in journal (Refereed)
    Abstract [en]

    Essential oils are complex mixtures of volatile components produced by the plant secondary metabolism and consist mainly of monoterpenes and sesquiterpenes and, to a minor extent, of aromatic and aliphatic compounds. They are exploited in several fields such as perfumery, food, pharmaceutics, and cosmetics. Essential oils have long-standing uses in the treatment of infectious diseases and parasitosis in humans and animals. In this regard, their therapeutic potential against human African trypanosomiasis (HAT) has not been fully explored. In the present work, we have selected six medicinal and aromatic plants (Azadirachta indica, Aframomum melegueta, Aframomum daniellii, Clausena anisata, Dichrostachys cinerea, and Echinops giganteus) traditionally used in Cameroon to treat several disorders, including infections and parasitic diseases, and evaluated the activity of their essential oils against Trypanosma brucei TC221. Their selectivity was also determined with Balb/3T3 (mouse embryonic fibroblast cell line) cells as a reference. The results showed that the essential oils from A. indica, A. daniellii, and E. giganteus were the most active ones, with half maximal inhibitory concentration (IC50) values of 15.21, 7.65, and 10.50 mu g/mL, respectively. These essential oils were characterized by different chemical compounds such as sesquiterpene hydrocarbons, monoterpene hydrocarbons, and oxygenated sesquiterpenes. Some of their main components were assayed as well on T. brucei TC221, and their effects were linked to those of essential oils.

  • 31. Kamte, Stephane L. Ngahang
    et al.
    Ranjbarian, Farahnaz
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Cianfaglione, Kevin
    Sut, Stefania
    Dall'Acqua, Stefano
    Bruno, Maurizio
    Afshar, Fariba Heshmati
    Iannarelli, Romilde
    Benelli, Giovanni
    Cappellacci, Loredana
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Maggi, Filippo
    Petrelli, Riccardo
    Identification of highly effective antitrypanosomal compounds in essential oils from the Apiaceae family2018In: Ecotoxicology and Environmental Safety, ISSN 0147-6513, E-ISSN 1090-2414, Vol. 156, p. 154-165Article in journal (Refereed)
    Abstract [en]

    The Apiaceae family encompasses aromatic plants of economic importance employed in foodstuffs, beverages, perfumery, pharmaceuticals and cosmetics. Apiaceae are rich sources of essential oils because of the wealth of secretory structures (ducts and vittae) they are endowed with. The Apiaceae essential oils are available on an industrial level because of the wide cultivation and disposability of the bulky material from which they are extracted as well as their relatively cheap price. In the fight against protozoal infections, essential oils may represent new therapeutic options. In the present work, we focused on a panel of nine Apiaceae species (Siler montamon, Sison amomum, Echinophora spinosa, Kundmannia sicula, Crithmum maritimum, Helosciadium nodiforum, Pimpinella anisum, Heracleum sphondylium and Trachyspermum cunmi) and their essential oils as a model for the identification of trypanocidal compounds to be used as alternative/integrative therapies in the treatment of Human African trypanosomiasis (HAT) and as starting material for drug design. The evaluation of inhibitory effects of the Apiaceae essential oils against Trypanosoma brucei showed that some of them (E. spinosa, S. amomum, C. maritimwn and H. nodifloruin) were active, with EC50 in the range 2.7-10.7 mu g/mL. Most of these oils were selective against T. brucei, except the one from C. maritimum that was highly selective against the BALB/3T3 mammalian cells. Testing nine characteristic individual components (a-pinene, sabinene, alpha-phellandrene, p-cymene, limonene, beta-ocimene, gamma-terpinene, terpinolene, and myristicin) of these oils, we showed that some of them had much higher selectivity than the oils themselves. Terpinolene was particularly active with an EC50 value of 0.035 mu g/rnL (0.26 mu M) and a selectivity index (SI) of 180. Four other compounds with EC50 in the range 1.0-6.0 mu g/mL (7.4-44 mu M) had also good SI: a-pinene (> 100), beta-ocimene (> 91), limonene (> 18) and sabinene ( > 17). In conclusion, these results highlight that the essential oils from the Apiaceae family are a reservoir of substances to be used as leading compounds for the development of natural drugs for the treatment of HAT.

  • 32. Lembo, D
    et al.
    Gribaudo, G
    Hofer, Anders
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Riera, L
    Cornaglia, M
    Mondo, A
    Angeretti, A
    Gariglio, M
    Thelander, Lars
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Landolfo, S
    Expression of an altered ribonucleotide reductase activity associated with the replication of murine cytomegalovirus in quiescent fibroblasts.2000In: Journal of Virology, ISSN 0022-538X, Vol. 74, no 24, p. 11557-65Article in journal (Refereed)
  • 33. Lembo, David
    et al.
    Donalisio, Manuela
    Hofer, Anders
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Cornaglia, Maura
    Brune, Wolfram
    Koszinowski, Ulrich
    Thelander, Lars
    Umeå University, Faculty of Medicine, Medical Biochemistry and Biophsyics.
    Landolfo, Santo
    The ribonucleotide reductase R1 homolog of murine cytomegalovirus is not a functional enzyme subunit but is required for pathogenesis.2004In: Journal of virology, ISSN 0022-538X, Vol. 78, no 8, p. 4278-88Article in journal (Refereed)
  • 34. Loderer, Christoph
    et al.
    Jonna, Venkateswara Rao
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Crona, Mikael
    Grinberg, Inna Rozman
    Sahlin, Margareta
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Lundin, Daniel
    Sjoeberg, Britt-Marie
    A unique cysteine-rich zinc finger domain present in a majority of class II ribonucleotide reductases mediates catalytic turnover2017In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 292, no 46, p. 19044-19054Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductases (RNRs) catalyze the reduction of ribonucleotides to the corresponding deoxyribonucleotides, used in DNA synthesis and repair. Two different mechanisms help deliver the required electrons to the RNR active site. Formate can be used as reductant directly in the active site, or glutaredoxins or thioredoxins reduce a C-terminal cysteine pair, which then delivers the electrons to the active site. Here, we characterized a novel cysteine-rich C-terminal domain (CRD), which is present in most class II RNRs found in microbes. The NrdJd-type RNR from the bacterium Stackebrandtia nassauensis was used as a model enzyme. We show that the CRD is involved in both higher oligomeric state formation and electron transfer to the active site. The CRD-dependent formation of high oligomers, such as tetramers and hexamers, was induced by addition of dATP or dGTP, but not of dTTP or dCTP. The electron transfer was mediated by an array of six cysteine residues at the very C-terminal end, which also coordinated a zinc atom. The electron transfer can also occur between subunits, depending on the enzyme's oligomeric state. An investigation of the native reductant of the system revealed no interaction with glutaredoxins or thioredoxins, indicating that this class II RNR uses a different electron source. Our results indicate that the CRD has a crucial role in catalytic turnover and a potentially new terminal reduction mechanism and suggest that the CRD is important for the activities of many class II RNRs.

  • 35. Petrelli, Riccardo
    et al.
    Meli, Maria
    Vita, Patrizia
    Torquati, Ilaria
    Ferro, Arianna
    Vodnala, Munender
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    D'Alessandro, Natale
    Tolomeo, Manlio
    Del Bello, Fabio
    Kusumanchi, Praveen
    Franchetti, Palmarisa
    Grifantini, Mario
    Jayaram, Hiremagalur N
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Cappellacci, Loredana
    From the covalent linkage of drugs to novel inhibitors of ribonucleotide reductase: Synthesis and biological evaluation of valproic esters of 3'-C-methyladenosine2014In: Bioorganic & Medicinal Chemistry Letters, ISSN 0960-894X, E-ISSN 1090-2120, Vol. 24, no 22, p. 5304-5309Article in journal (Refereed)
    Abstract [en]

    We synthesized a series of serum-stable covalently linked drugs derived from 3'-C-methyladenosine (3'-Me-Ado) and valproic acid (VPA), which are ribonucleotide reductase (RR) and histone deacetylase (HDAC) inhibitors, respectively. While the combination of free VPA and 3'-Me-Ado resulted in a clear synergistic apoptotic effect, the conjugates had lost their HDAC inhibitory effect as well as the corresponding apoptotic activity. Two of the analogs, 2',5'-bis-O-valproyl-3'-C-methyladenosine (A160) and 5'-O-valproyl-3'-C-methyladenosine (A167), showed promising cytotoxic activities against human hematological and solid cancer cell lines. A167 was less potent than A160 but had interesting features as an RR inhibitor. It inhibited RR activity by competing with ATP as an allosteric effector and concomitantly reduced the intracellular deoxyribonucleoside triphosphate (dNTP) pools. A167 represents a novel lead compound, which in contrast to previously used RR nucleoside analogs does not require intracellular kinases for its activity and therefore holds promise against drug resistant tumors with downregulated nucleoside kinases.

  • 36. Petrelli, Riccardo
    et al.
    Orsomando, Giuseppe
    Sorci, Leonardo
    Maggi, Filippo
    Ranjbarian, Farahnaz
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Nya, Prosper C. Biapa
    Petrelli, Dezemona
    Vitali, Luca A.
    Lupidi, Giulio
    Quassinti, Luana
    Bramucci, Massimo
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Cappellacci, Loredana
    Biological Activities of the Essential Oil from Erigeron floribundus2016In: Molecules, ISSN 1420-3049, E-ISSN 1420-3049, Vol. 21, no 8, article id 1065Article in journal (Refereed)
    Abstract [en]

    Erigeron floribundus (Asteraceae) is an herbaceous plant widely used in Cameroonian traditional medicine to treat various diseases of microbial and non-microbial origin. In the present study, we evaluated the in vitro biological activities displayed by the essential oil obtained from the aerial parts of E. floribundus, namely the antioxidant, antimicrobial and antiproliferative activities. Moreover, we investigated the inhibitory effects of E. floribundus essential oil on nicotinate mononucleotide adenylyltransferase (NadD), a promising new target for developing novel antibiotics, and Trypanosoma brucei, the protozoan parasite responsible for Human African trypanosomiasis. The essential oil composition was dominated by spathulenol (12.2%), caryophyllene oxide (12.4%) and limonene (8.8%). The E. floribundus oil showed a good activity against Staphylococcus aureus (inhibition zone diameter, IZD of 14 mm, minimum inhibitory concentration, MIC of 512 mu g/mL). Interestingly, it inhibited the NadD enzyme from S. aureus (IC50 of 98 mu g/mL), with no effects on mammalian orthologue enzymes. In addition, T. brucei proliferation was inhibited with IC50 values of 33.5 mu g/mL with the essential oil and 5.6 mu g/mL with the active component limonene. The essential oil exhibited strong cytotoxicity on HCT 116 colon carcinoma cells with an IC50 value of 14.89 mu g/mL, and remarkable ferric reducing antioxidant power (tocopherol-equivalent antioxidant capacity, TEAC = 411.9 mu mol.TE/g).

  • 37. Petrelli, Riccardo
    et al.
    Ranjbarian, Farahnaz
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Dall'Acqua, Stefano
    Papa, Fabrizio
    Iannarelli, Romilde
    Ngahang Kamte, Stephane L.
    Vittori, Sauro
    Benelli, Giovanni
    Maggi, Filippo
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Cappellacci, Loredana
    An overlooked horticultural crop, Smyrnium olusatrum, as a potential source of compounds effective against African trypanosomiasis2017In: Parasitology international, ISSN 1383-5769, E-ISSN 1873-0329, Vol. 66, no 2, p. 146-151Article in journal (Refereed)
    Abstract [en]

    Among natural products, sesquiterpenes have shown promising inhibitory effects against bloodstream forms of Trypanosoma brucei, the protozoan parasite causing human African trypanosomiasis (HAT). Smyrnium olusatrum (Apiaceae), also known as Alexanders or wild celery, is a neglected horticultural crop characterized by oxygenated sesquiterpenes containing a furan ring. In the present work we explored the potential of its essential oils obtained from different organs and the main oxygenated sesquiterpenes, namely isofuranodiene, germacrone and β-acetoxyfuranoeudesm-4(15)-ene, as inhibitors of Trypanosoma brucei. All essential oils effectively inhibited the growth of parasite showing IC50 values of 1.9–4.0 μg/ml. Among the main essential oil constituents, isofuranodiene exhibited a significant and selective inhibitory activity against T. brucei (IC50 of 0.6 μg/ml, SI = 30), with β-acetoxyfuranoeudesm-4(15)-ene giving a moderate potentiating effect. These results shed light on the possible application of isofuranodiene as an antiprotozoal agent to be included in combination treatments aimed not only at curing patients but also at preventing the diffusion of HAT.

  • 38. Ranjbarian, Farahnaz
    et al.
    Vodnala, Munender
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Alzahrani, K. H.
    de Koning, H. P.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    9-(2-Deoxy-2-fluoro-ß-D-arabinofuranosyl) adenine: a potent antitrypanosomal adenosine analogue that circumvents transport-related drug resistanceManuscript (preprint) (Other academic)
  • 39.
    Ranjbarian, Farahnaz
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Vodnala, Munender
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Alzahrani, Khalid J. H.
    Ebiloma, Godwin U.
    de Koning, Harry P.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    9-(2 '-Deoxy-2 '-Fluoro-beta-D-Arabinofuranosyl) Adenine Is a Potent Antitrypanosomal Adenosine Analogue That Circumvents Transport-Related Drug Resistance2017In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 61, no 6, article id e02719-16Article in journal (Refereed)
    Abstract [en]

    Current chemotherapy against African sleeping sickness, a disease caused by the protozoan parasite Trypanosoma brucei, is limited by toxicity, inefficacy, and drug resistance. Nucleoside analogues have been successfully used to cure T. brucei-infected mice, but they have the limitation of mainly being taken up by the P2 nucleoside transporter, which, when mutated, is a common cause of multidrug resistance in T. brucei. We report here that adenine arabinoside (Ara-A) and the newly tested drug 9-(2'-deoxy-2'-fluoro-beta-D-arabinofuranosyl) adenine (FANA-A) are instead taken up by the P1 nucleoside transporter, which is not associated with drug resistance. Like Ara-A, FANA-A was found to be resistant to cleavage by methylthioadenosine phosphorylase, an enzyme that protects T. brucei against the antitrypanosomal effects of deoxyadenosine. Another important factor behind the selectivity of nucleoside analogues is how well they are phosphorylated within the cell. We found that the T. brucei adenosine kinase had a higher catalytic efficiency with FANA-A than the mammalian enzyme, and T. brucei cells treated with FANA-A accumulated high levels of FANA-A triphosphate, which even surpassed the level of ATP and led to cell cycle arrest, inhibition of DNA synthesis, and the accumulation of DNA breaks. FANA-A inhibited nucleic acid biosynthesis and parasite proliferation with 50% effective concentrations (EC(50)s) in the low nanomolar range, whereas mammalian cell proliferation was inhibited in the micromolar range. Both Ara-A and FANA-A, in combination with deoxycoformycin, cured T. brucei-infected mice, but FANA-A did so at a dose 100 times lower than that of Ara-A.

  • 40.
    Ranjbarian, Farahnaz
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Vodnala, Munender
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Vodnala, Sharvani Munender
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Rofougaran, Reza
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Department of Biochemistry, Institute of Biochemistry and Biophysics, Tehran University, Tehran, Iran.
    Thelander, Lars
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Trypanosoma brucei thymidine kinase is tandem protein consisting of two homologous parts, which together enable efficient substrate binding2012In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 287, no 21, p. 17628-17636Article in journal (Refereed)
    Abstract [en]

    Trypanosoma brucei causes African sleeping sickness, a disease for which existing chemotherapies are limited by their toxicity or lack of efficacy. We have found that four parasites, including T. brucei, contain genes where two or four thymidine kinase (TK) sequences are fused into a single open reading frame. The T. brucei full-length enzyme as well as its two constituent parts, domain 1 and domain 2, were separately expressed and characterized. Of potential interest for nucleoside analog development, T. brucei TK was less discriminative against purines than human TK1 with the following order of catalytic efficiencies: thymidine > deoxyuridine ≫ deoxyinosine > deoxyguanosine. Proteins from the TK1 family are generally dimers or tetramers, and the quaternary structure is linked to substrate affinity. T. brucei TK was primarily monomeric but can be considered a two-domain pseudodimer. Independent kinetic analysis of the two domains showed that only domain 2 was active. It had a similar turnover number (k(cat)) as the full-length enzyme but could not self-dimerize efficiently and had a 5-fold reduced thymidine/deoxyuridine affinity. Domain 1, which lacks three conserved active site residues, can therefore be considered a covalently attached structural partner that enhances substrate binding to domain 2. A consequence of the non-catalytic role of domain 1 is that its active site residues are released from evolutionary pressure, which can be advantageous for developing new catalytic functions. In addition, nearly identical 89-bp sequences present in both domains suggest that the exchange of genetic material between them can further promote evolution.

  • 41.
    Rofougaran, Reza
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Crona, Mikael
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Vodnala, Munender
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Sjöberg, Britt-Marie
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Oligomerization status directs overall activity regulation of the Escherichia coli class Ia ribonucleotide reductase2008In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 283, no 51, p. 35310-35318Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductase (RNR) is a key enzyme for the synthesis of the four DNA building blocks. Class Ia RNRs contain two subunits, denoted R1 (α) and R2 (β). These enzymes are regulated via two nucleotide-binding allosteric sites on the R1 subunit, termed the specificity and overall activity sites. The specificity site binds ATP, dATP, dTTP, or dGTP and determines the substrate to be reduced, whereas the overall activity site binds dATP (inhibitor) or ATP. By using gas-phase electrophoretic mobility macromolecule analysis and enzyme assays, we found that the Escherichia coli class Ia RNR formed an inhibited α4β4 complex in the presence of dATP and an active α2β2 complex in the presence of ATP (main substrate: CDP), dTTP (substrate: GDP) or dGTP (substrate: ADP). The R1-R2 interaction was 30–50 times stronger in the α4β4 complex than in the α2β2complex, which was in equilibrium with free α2 and β2 subunits. Studies of a known E. coli R1 mutant (H59A) showed that deficient dATP inhibition correlated with reduced ability to form α4β4 complexes. ATP could also induce the formation of a generally inhibited α4β4 complex in the E. coli RNR but only when used in combination with high concentrations of the specificity site effectors, dTTP/dGTP. Both allosteric sites are therefore important for α4β4 formation and overall activity regulation. The E. coli RNR differs from the mammalian enzyme, which is stimulated by ATP also in combination with dGTP/dTTP and forms active and inactive α6β2 complexes.

  • 42.
    Rofougaran, Reza
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Vodnala, Munender
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Enzymatically active mammalian ribonucleotide reductase exists primarily as an α6β2 octamer2006In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 281, no 38, p. 27705-27711Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductase synthesizes deoxyribonucleotides, which are essential building blocks for DNA synthesis. The mammalian ribonucleotide reductase is described as an alpha(2)beta(2) complex consisting of R1 (alpha) and R2 (beta) proteins. ATP stimulates and dATP inhibits enzyme activity by binding to an allosteric site called the activity site on the R1 protein. Despite the opposite effects by ATP and dATP on enzyme activity, both nucleotides induce formation of R1 oligomers. By using a new technique termed Gas-phase Electrophoretic-Mobility Macromolecule Analysis (GEMMA), we have found that the ATP/dATP-induced R1 oligomers have a defined size (hexamers) and can interact with the R2 dimer to form an enzymatically active protein complex (alpha(6)beta(2)). The newly discovered alpha(6)beta(2) complex can either be in an active or an inhibited state depending on whether ATP or dATP is bound. Our results suggest that this protein complex is the major form of ribonucleotide reductase at physiological levels of R1-R2 protein and nucleotides.

  • 43. Rozman Grinberg, Inna
    et al.
    Lundin, Daniel
    Hasan, Mahmudul
    Crona, Mikael
    Rao Jonna, Venkateswara
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Loderer, Chrishtoph
    Sahlin, Margareta
    Markova, Natalia
    Borovok, Ilya
    Berggren, Gustav
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Logan, Derek T.
    Sjöberg, Britt-Marie
    Novel ATP-cone-driven allosteric regulation of ribonucleotide reductase via the radical-generating subunit2018In: eLIFE, E-ISSN 2050-084X, Vol. 7, article id e31529Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductases (RNRs) are key enzymes in DNA metabolism, with allosteric mechanisms controlling substrate specificity and overall activity. In RNRs, the activity master-switch, the ATP-cone, has been found exclusively in the catalytic subunit. In two class I RNR subclasses whose catalytic subunit lacks the ATP-cone, we discovered ATP-cones in the radical-generating subunit. The ATP-cone in the Leeuwenhoekiella blandensis radical-generating subunit regulates activity via quaternary structure induced by binding of nucleotides. ATP induces enzymatically competent dimers, whereas dATP induces non-productive tetramers, resulting in different holoenzymes. The tetramer forms by interactions between ATP-cones, shown by a 2.45 A crystal structure. We also present evidence for an (MnMnIV)-Mn-III metal center. In summary, lack of an ATP-cone domain in the catalytic subunit was compensated by transfer of the domain to the radical-generating subunit. To our knowledge, this represents the first observation of transfer of an allosteric domain between components of the same enzyme complex.

  • 44. Rozman Grinberg, Inna
    et al.
    Lundin, Daniel
    Sahlin, Margareta
    Crona, Mikael
    Berggren, Gustav
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Sjöberg, Britt-Marie
    A glutaredoxin domain fused to the radical-generating subunit of ribonucleotide reductase (RNR) functions as an efficient RNR reductant2018In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 293, no 41, p. 15889-15900Article in journal (Refereed)
    Abstract [en]

    Class I ribonucleotide reductase (RNR) consists of a catalytic subunit (NrdA) and a radical-generating subunit (NrdB) that together catalyze reduction of ribonucleotides to their corresponding deoxyribonucleotides. NrdB from the firmicute Facklamia ignava is a unique fusion protein with N-terminal add-ons of a glutaredoxin (Grx) domain followed by an ATP-binding domain, the ATP cone. Grx, usually encoded separately from the RNR operon, is a known RNR reductant. We show that the fused Grx domain functions as an efficient reductant of the F. ignava class I RNR via the common dithiol mechanism and, interestingly, also via a monothiol mechanism, although less efficiently. To our knowledge, a Grx that uses both of these two reaction mechanisms has not previously been observed with a native substrate. The ATP cone is in most RNRs an N-terminal domain of the catalytic subunit. It is an allosteric on/off switch promoting ribonucleotide reduction in the presence of ATP and inhibiting RNR activity in the presence of dATP. We found that dATP bound to the ATP cone of F. ignava NrdB promotes formation of tetramers that cannot form active complexes with NrdA. The ATP cone bound two dATP molecules but only one ATP molecule. F. ignava NrdB contains the recently identified radical-generating cofactor MnIII/MnIV. We show that NrdA from F. ignava can form a catalytically competent RNR with the MnIII/MnIV-containing NrdB from the flavobacterium Leeuwenhoekiella blandensis. In conclusion, F. ignava NrdB is fused with a Grx functioning as an RNR reductant and an ATP cone serving as an on/off switch.

  • 45.
    Sadanandan, Sajna Anand
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Ekström, Jens-Ola
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Institute of Biomedical Technology, University of Tampere, FI-33520 Tampere, Finland.
    Jonna, Venkateswara Rao
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hultmark, Dan
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Institute of Biomedical Technology, University of Tampere, FI-33520 Tampere, Finland.
    VP3 is crucial for the stability of Nora virus virions2016In: Virus Research, ISSN 0168-1702, E-ISSN 1872-7492, Vol. 223, p. 20-27Article in journal (Refereed)
    Abstract [en]

    Nora virus is an enteric virus that causes persistent, non-pathological infection in Drosophila melanogaster. It replicates in the fly gut and is transmitted via the fecal-oral route. Nora virus has a single-stranded positive-sense RNA genome, which is translated in four open reading frames. Reading frame three encodes the VP3 protein, the structure and function of which we have investigated in this work. We have shown that VP3 is a trimer that has an α-helical secondary structure, with a functionally important coiled-coil domain. In order to identify the role of VP3 in the Nora virus life cycle, we constructed VP3-mutants using the cDNA clone of the virus. Our results show that VP3 does not have a role in the actual assembly of the virus particles, but virions that lack VP3 or harbor VP3 with a disrupted coiled coil domain are incapable of transmission via the fecal-oral route. Removing the region downstream of the putative coiled coil appears to have an effect on the fitness of the virus but does not hamper its replication or transmission. We also found that the VP3 protein and particularly the coiled coil domain are crucial for the stability of Nora virus virions when exposed to heat or proteases. Hence, we propose that VP3 is imperative to Nora virus virions as it confers stability to the viral capsid.

  • 46.
    Schmitt, Andreas
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Jiang, Kai
    Camacho, Martha I.
    Jonna, Venkateswara Rao
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Westerlund, Fredrik
    Christie, Peter J.
    Berntsson, Ronnie P-A.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    PrgB promotes aggregation, biofilm formation, and conjugation through DNA binding and compaction2018In: Molecular Microbiology, ISSN 0950-382X, E-ISSN 1365-2958, Vol. 109, no 3, p. 291-305Article in journal (Refereed)
    Abstract [en]

    Gram-positive bacteria deploy type IV secretion systems (T4SSs) to facilitate horizontal gene transfer. The T4SSs of Gram-positive bacteria rely on surface adhesins as opposed to conjugative pili to facilitate mating. Enterococcus faecalis PrgB is a surface adhesin that promotes mating pair formation and robust biofilm development in an extracellular DNA (eDNA) dependent manner. Here, we report the structure of the adhesin domain of PrgB. The adhesin domain binds and compacts DNA in vitro. In vivo PrgB deleted of its adhesin domain does not support cellular aggregation, biofilm development and conjugative DNA transfer. PrgB also binds lipoteichoic acid (LTA), which competes with DNA binding. We propose that PrgB binding and compaction of eDNA facilitates cell aggregation and plays an important role in establishment of early biofilms in mono- or polyspecies settings. Within these biofilms, PrgB mediates formation and stabilization of direct cell-cell contacts through alternative binding of cell-bound LTA, which in turn promotes establishment of productive mating junctions and efficient intra- or inter-species T4SS-mediated gene transfer.

  • 47. Sut, Stefania
    et al.
    Dall'Acqua, Stefano
    Baldan, Valeria
    Ngahang Kamte, Stephane L.
    Ranjbarian, Farahnaz
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Biapa Nya, Prosper C.
    Vittori, Sauro
    Benelli, Giovanni
    Maggi, Filippo
    Cappellacci, Loredana
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Petrelli, Riccardo
    Identification of tagitinin C from Tithonia diversifolia as antitrypanosomal compound using bioactivity-guided fractionation2018In: Fitoterapia (Milano), ISSN 0367-326X, E-ISSN 1873-6971, Vol. 124, p. 145-151Article in journal (Refereed)
    Abstract [en]

    Tithonia diversifolia (Asteraceae), is used as traditional medicine in tropical countries for the treatment of various diseases, including malaria. Although numerous studies have assessed the antimalarial properties, nothing is known about the effect of T. diversifolia extracts on trypanosomiasis. In this study extracts of T. diversifolia aerial parts were evaluated for their bioactivity against Trypanosoma brucei. The activity was studied against bloodstream forms of T. brucei (TC221), as well as against mammalian cells (BALB/3T3 mouse fibroblasts), as a counter-screen for toxicity. Both methanolic and aqueous extracts showed significant effects with IC50 values of 1.1 and 2.2 mu g/mL against T. brucei (TC221) and 5.2 and 3.7 mu g/mL against BALB/3T3 cells, respectively. A bioassay-guided fractionation on the methanolic extract yielded in identification of active fractions (F8 and F9) with IC50 values of 0.41 and 0.43 mu g/mL, respectively, against T. brucei (TC221) and 1.4 and 1.5 mu g/mL, respectively, against BALB/3T3 cells,. The phytochemical composition of the extracts and the purified fractions were investigated using HPLC-ESI-MS/MS and 1D and 2D NMR spectra showing the presence of sesquiterpene lactones that in turn were subjected to the isolation procedure. Tagitinin A and C were rather active but the latter presented a very strong inhibition on T. brucei (TC221) with an IC50 value of 0.0042 mu g/mL. This activity was 4.5 times better than that of the reference drug suramin. The results of this study shed light on the antitrypanosomal effects of T. diversifolia extracts and highlighted tagitinin C as one of the possible responsible for this effect. Further structure activity relationships studies on tagitinins are needed to consider this sesquiterpenes as lead compounds for the development of new antitrypanosomal drugs.

  • 48. Vande Voorde, Johan
    et al.
    Sabuncuoglu, Suna
    Noppen, Sam
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Ranjbarian, Farahnaz
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Fieuws, Steffen
    Balzarini, Jan
    Liekens, Sandra
    Nucleoside-catabolizing Enzymes in Mycoplasma-infected Tumor Cell Cultures Compromise the Cytostatic Activity of the Anticancer Drug Gemcitabine2014In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 289, no 19, p. 13054-13065Article in journal (Refereed)
    Abstract [en]

    Background: Gemcitabine is used to treat solid tumors. Some mycoplasmas preferentially colonize tumors in patients. Results: Mycoplasma-encoded cytidine deaminase and pyrimidine nucleoside phosphorylase compromise the cytostatic/antitumor activity of gemcitabine in mycoplasma-infected tumor cell cultures and xenografts in mice. Conclusion: Tumor-associated mycoplasmas may decrease the therapeutic efficiency of gemcitabine. Significance: Current treatment of mycoplasma-infected tumors with gemcitabine may be suboptimal. The intracellular metabolism and cytostatic activity of the anticancer drug gemcitabine (2,2-difluoro-2-deoxycytidine; dFdC) was severely compromised in Mycoplasma hyorhinis-infected tumor cell cultures. Pronounced deamination of dFdC to its less cytostatic metabolite 2,2-difluoro-2-deoxyuridine was observed, both in cell extracts and spent culture medium (i.e. tumor cell-free but mycoplasma-containing) of mycoplasma-infected tumor cells. This indicates that the decreased antiproliferative activity of dFdC in such cells is attributed to a mycoplasma cytidine deaminase causing rapid drug catabolism. Indeed, the cytostatic activity of gemcitabine could be restored by the co-administration of tetrahydrouridine (a potent cytidine deaminase inhibitor). Additionally, mycoplasma-derived pyrimidine nucleoside phosphorylase (PyNP) activity indirectly potentiated deamination of dFdC: the natural pyrimidine nucleosides uridine, 2-deoxyuridine and thymidine inhibited mycoplasma-associated dFdC deamination but were efficiently catabolized (removed) by mycoplasma PyNP. The markedly lower anabolism and related cytostatic activity of dFdC in mycoplasma-infected tumor cells was therefore also (partially) restored by a specific TP/PyNP inhibitor (TPI), or by exogenous thymidine. Consequently, no effect on the cytostatic activity of dFdC was observed in tumor cell cultures infected with a PyNP-deficient Mycoplasma pneumoniae strain. Because it has been reported that some commensal mycoplasma species (including M. hyorhinis) preferentially colonize tumor tissue in cancer patients, our findings suggest that the presence of mycoplasmas in the tumor microenvironment could be a limiting factor for the anticancer efficiency of dFdC-based chemotherapy. Accordingly, a significantly decreased antitumor effect of dFdC was observed in mice bearing M. hyorhinis-infected murine mammary FM3A tumors compared with uninfected tumors.

  • 49.
    Vodnala, Munender
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Fijolek, Artur
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Rofougaran, Reza
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Mosimann, Marc
    Mäser, Pascal
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Adenosine kinase mediates high affinity adenosine salvage in Trypanosoma brucei2008In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 283, no 9, p. 5380-5388Article in journal (Refereed)
  • 50.
    Vodnala, Munender
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Ranjbarian, Farahnaz
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    The protective role of Trypanosoma brucei methylthioadenosine phosphorylase against deoxyadenosine and deoxyadenosine analogsManuscript (preprint) (Other academic)
12 1 - 50 of 51
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