Digitala Vetenskapliga Arkivet

Endre søk
Begrens søket
1234567 101 - 150 of 6026
RefereraExporteraLink til resultatlisten
Permanent link
Referera
Referensformat
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Annet format
Fler format
Språk
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Annet språk
Fler språk
Utmatningsformat
  • html
  • text
  • asciidoc
  • rtf
Treff pr side
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sortering
  • Standard (Relevans)
  • Forfatter A-Ø
  • Forfatter Ø-A
  • Tittel A-Ø
  • Tittel Ø-A
  • Type publikasjon A-Ø
  • Type publikasjon Ø-A
  • Eldste først
  • Nyeste først
  • Skapad (Eldste først)
  • Skapad (Nyeste først)
  • Senast uppdaterad (Eldste først)
  • Senast uppdaterad (Nyeste først)
  • Disputationsdatum (tidligste først)
  • Disputationsdatum (siste først)
  • Standard (Relevans)
  • Forfatter A-Ø
  • Forfatter Ø-A
  • Tittel A-Ø
  • Tittel Ø-A
  • Type publikasjon A-Ø
  • Type publikasjon Ø-A
  • Eldste først
  • Nyeste først
  • Skapad (Eldste først)
  • Skapad (Nyeste først)
  • Senast uppdaterad (Eldste først)
  • Senast uppdaterad (Nyeste først)
  • Disputationsdatum (tidligste først)
  • Disputationsdatum (siste først)
Merk
Maxantalet träffar du kan exportera från sökgränssnittet är 250. Vid större uttag använd dig av utsökningar.
  • 101.
    Almstedt, Karin
    et al.
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biokemi. Linköpings universitet, Tekniska högskolan.
    Lundqvist, Martin
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Molekylär Bioteknik. Linköpings universitet, Tekniska högskolan.
    Carlsson, Jonas
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Bioinformatik. Linköpings universitet, Tekniska högskolan.
    Karlsson, Martin
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biokemi. Linköpings universitet, Tekniska högskolan.
    Persson, Bengt
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Bioinformatik. Linköpings universitet, Tekniska högskolan.
    Jonsson, Bengt-Harald
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Molekylär Bioteknik. Linköpings universitet, Tekniska högskolan.
    Carlsson, Uno
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biokemi. Linköpings universitet, Tekniska högskolan.
    Hammarström, Per
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biokemi. Linköpings universitet, Tekniska högskolan.
    Unfolding a folding disease: folding, misfolding and aggregation of the marble brain syndrome-associated mutant H107Y of human carbonic anhydrase II2004Inngår i: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 342, nr 2, s. 619-633Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

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

  • 102.
    Almstedt, Karin
    et al.
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biokemi. Linköpings universitet, Tekniska högskolan.
    Mårtensson, Lars-Göran
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biokemi. Linköpings universitet, Tekniska högskolan.
    Carlsson, Uno
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biokemi. Linköpings universitet, Tekniska högskolan.
    Hammarström, Per
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biokemi. Linköpings universitet, Tekniska högskolan.
    Thermodynamic interrogation of a folding disease. Mutant mapping of position 107 in human carbonic anhydrase II linked to marble brain disease.2008Inngår i: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 47, nr 5, s. 1288-1298Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Marble brain disease (MBD) also known as Guibaud−Vainsel syndrome is caused by autosomal recessive mutations in the human carbonic anhydrase II (HCA II) gene. HCA II is a 259 amino acid single domain enzyme and is dominated by a 10-stranded β-sheet. One mutation associated with MBD entails the H107Y substitution where H107 is a highly conserved residue in the carbonic anhydrase protein family. We have previously demonstrated that the H107Y mutation is a remarkably destabilizing folding mutation [Almstedt et al. (2004) J. Mol. Biol. 342, 619−633]. Here, the exceptional destabilization by the H107Y mutation has been further investigated. A mutational survey of position H107 and a neighboring conserved position E117 has been performed entailing the mutants H107A, H107F, H107N, E117A and the double mutants H107A/E117A and H107N/E117A. All mutants were severely destabilized versus GuHCl and heat denaturation. Thermal denaturation and GuHCl phase diagram and ANS analyses showed that the mutants shifted HCA II toward populating ensembles of intermediates of molten globule type under physiological conditions. The native state stability of the mutants was in the following order:  wt > H107N > E117A > H107A > H107F > H107Y > H107N/E117A > H107A/E117A. In conclusion:  (i) H107N is least destabilizing likely due to compensatory H-bonding ability of the introduced Asn residue. (ii) Double mutant cycles surprisingly reveal additive destabilization of H107N and E117A showing that H107 and E117 are independently stabilizing the folded protein. (iii) H107Y and H107F are exceptionally destabilizing due to bulkiness of the side chains whereas H107A is more accommodating, indicating long-range destabilizing effects of the natural pathogenic H107Y mutation.

  • 103.
    Almén, Markus Sällman
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Lamichhaney, Sangeet
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Berglund, Jonas
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Grant, B. Rosemary
    Princeton Univ, Dept Ecol & Evolutionary Biol, Princeton, NJ 08544 USA..
    Grant, Peter R.
    Princeton Univ, Dept Ecol & Evolutionary Biol, Princeton, NJ 08544 USA..
    Webster, Matthew T.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Andersson, Leif
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi. Swedish Univ Agr Sci, Dept Anim Breeding & Genet, Uppsala, Sweden.;Texas A&M Univ, Dept Vet Integrat Biosci, College Stn, TX USA..
    Adaptive radiation of Darwin's finches revisited using whole genome sequencing2016Inngår i: Bioessays, ISSN 0265-9247, E-ISSN 1521-1878, Vol. 38, nr 1, s. 14-20Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    We recently used genome sequencing to study the evolutionary history of the Darwin's finches. A prominent feature of our data was that different polymorphic sites in the genome tended to indicate different genetic relationships among these closely related species. Such patterns are expected in recently diverged genomes as a result of incomplete lineage sorting. However, we uncovered conclusive evidence that these patterns have also been influenced by interspecies hybridisation, a process that has likely played an important role in the radiation of Darwin's finches. A major discovery was that segregation of two haplotypes at the ALX1 locus underlies variation in beak shape among the Darwin's finches, and that differences between the two haplotypes in a 240 kb region in blunt and pointed beaked birds involve both coding and regulatory changes. As we review herein, the evolution of such adaptive haplotypes comprising multiple causal changes appears to be an important mechanism contributing to the evolution of biodiversity.

  • 104.
    Alneberg, Johannes
    et al.
    KTH, Skolan för bioteknologi (BIO), Genteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Bjarnason, Brynjar Smári
    KTH, Skolan för bioteknologi (BIO), Genteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    de Bruijn, Ino
    KTH, Skolan för bioteknologi (BIO), Genteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab. Bioinformatics Infrastructure for Life Sciences (BILS), Sweden.
    Schirmer, Melanie
    Quick, Joshua
    Ijaz, Umer Z.
    Lahti, Leo
    Loman, Nicholas J.
    Andersson, Anders F.
    KTH, Skolan för bioteknologi (BIO), Genteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Quince, Christopher
    Binning metagenomic contigs by coverage and composition2014Inngår i: Nature Methods, ISSN 1548-7091, E-ISSN 1548-7105, Vol. 11, nr 11, s. 1144-1146Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Shotgun sequencing enables the reconstruction of genomes from complex microbial communities, but because assembly does not reconstruct entire genomes, it is necessary to bin genome fragments. Here we present CONCOCT, a new algorithm that combines sequence composition and coverage across multiple samples, to automatically cluster contigs into genomes. We demonstrate high recall and precision on artificial as well as real human gut metagenome data sets.

  • 105. Alriksson, Björn
    et al.
    Horváth, Ilona Sárvári
    Jönsson, Leif J
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Department of Chemistry and Biomedical Sciences, Karlstad University.
    Overexpression of Saccharomyces cerevisiae transcription factor and multidrug resistance genes conveys enhanced resistance to lignocellulose-derived fermentation inhibitors2010Inngår i: Process Biochemistry, ISSN 1359-5113, E-ISSN 1873-3298, Vol. 45, nr 2, s. 264-271Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The presence of fermentation inhibitors in lignocellulose hydrolysates is an obstacle for achieving efficient fermentation of lignocellulose hydrolysates to ethanol and other commodities. In this investigation, the possibility of generating more inhibitor-resistant Saccharomyces cerevisiae by genetic engineering was explored. Based on previous results from studies of deletion mutants, three S. cerevisiae genes (ATR1, FLR1, YAP1) involved in multidrug resistance and stress response of yeast were selected for overexpression in three S. cerevisiae strains. The resistance of the transformed strains to lignocellulose-derived fermentation inhibitors and a dilute-acid spruce hydrolysate was evaluated in fermentation experiments. Overexpression of FLR1 resulted in enhanced resistance to the phenolic inhibitor coniferyl aldehyde and the furan aldehyde HMF (5-hydroxymethyl-2-furaldehyde). Overexpression of ATR1 conferred increased resistance to coniferyl aldehyde. Strains overexpressing YAP1, which encodes a transcription factor, displayed increased resistance to coniferyl aldehyde, HMF, and the spruce hydrolysate. An ethanol productivity of 0.17 g ethanol × l−1 × h−1 was achieved for a YAP1-overexpressing transformant cultivated in spruce hydrolysate, whereas a control transformant, which did not overexpress YAP1, only reached a productivity of 0.05 g ethanol × l−1 × h−1

  • 106.
    Al-Shakargi, Bilall
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för farmaceutisk biovetenskap.
    Variationer i allelfrekvens hos cytokrom-generna;CYP3A4*1B, CYP3A5*3 och CYP2B6*6 mellan Uganda och Tanzania.2019Independent thesis Advanced level (degree of Master (Two Years)), 20 poäng / 30 hpOppgave
    Abstract [sv]

    1. Abstrakt:Bakgrund: Malaria är en av världens viktigaste infektionssjukdomar och det beräknas vara ca300 miljoner drabbade varje år, därför är metabolismen av läkemedel som används för attbehandla malaria såsom kinin och artemisinin värt att lägga fokus på. Cytokrom P450enzymerna har en viktig roll i metabolism av malarialäkemedel och dessa uppvisar variationinterindividuellt samt mellan olika populationer på grund av polymorfism.Syfte: Denna studie har fokuserat på att undersöka tre polymorfa gener (CYP3A4*1B,CYP3A5*3 och CYP2B6*6) hos både friska och malariasmittade barn i Uganda för attjämföra resultaten med en population i Mwanza, Tanzania. Dessa polymorfa alleler påverkarmetabolismen av artemisinin och kinin, vilket i sin tur kan förorsaka minskad/ökad ellermisslyckad klinisk effekt.Metod: Genotypning av individernas blodprov gällande dessa genvarianter undersöktesgenom laboratoriestudier med PCR som huvudsaklig metod. DNA sekvensering utfördes vidUppsala Genome Center.Resultat: Resultaten visade att allelfrekvensen i Mwanza för CYP3A4*1B var 78%respektive 16% för CYP3A5*3, medan de var 72 % respektive 50% hos populationen iUganda. Allelfrekvensen för CYP2B6*6 i Uganda var 72 % och 36 % i Mwanza, Tanzania.Sammanfattning: De flesta malariamediciner uppvisar skillnader i kinetik och dynamikvilket kräver terapeutisk läkemedelsmonitorering. Sammanfattningsvis finns det skillnaderoch variationer bland de studerade polymorfa CYP enzymer interindividuellt och mellanolikaetniska grupper.Resultatet av denna studie visade både små skillnader mellan de två studerade populationernamen även skillnader mellan individer i den studerade gruppen i Uganda.

    Fulltekst (pdf)
    fulltext
  • 107.
    Alstermark, Mirjam
    Linnéuniversitetet, Fakulteten för Hälso- och livsvetenskap (FHL), Institutionen för kemi och biomedicin (KOB).
    Fluorescens in situ hybridisering: Optimering och vidareutveckling av en kurslaboration på Biomedicinska analytikerprogrammet2019Independent thesis Basic level (degree of Bachelor), 10 poäng / 15 hpOppgave
    Abstract [sv]

    Fluorescens In Situ Hybridisering (FISH) är en cytogenetisk teknik som kan detektera genetiska sjukdomar och avvikelser i Deoxyribonukleinsyra (DNA) och Ribonukleinsyra (RNA). FISH börjar med kromosomutvinning av önskat analyspreparat, därefter får en direkt- eller indirekt fluorescensinmärkt probe (15-30 baspar lång) binda in till sin genetiska målsekvens via hybridisering. Preparatet kan sedan analyseras i fluorescensmikroskop för att bedöma om proben bundit in till sin målsekvens. I kursen ”Fördjupad laboratoriemetodik” för Biomedicinska analytikerprogrammet, Linnéuniversitetet utförs en FISH-laboration där resultaten varit otydliga och ej reproducerbara. Syftet med examensarbetet var att förbättra kurslaboration FISH som ges under kursen ”Fördjupad laboratoriemetodik”.

    Adherenta celler odlades till ≥ 3 x 106 i antal och till viss konfluens; primära endotelcell-linjen HCMEC till konfluenserna 60 % och 80 % och cancercell-linjerna VMM1 och H1915 till 80 % konfluens. Därefter skördades cellerna och dess respektive kromosomer utvanns. Kromosomerna undersöktes sedan med metoderna G-bandsfärgning, DNA-FISH och Multicolor-FISH (24X-probe) för tydliga och reproducerbara resultat.

    G-bandsfärgningen av HCMEC visade många hela interfasceller och få fria kromosomer för celler i 60 % konfluens. Både G-bandsfärgningen och DNA-FISH visade att HCMEC odlade till 80 % konfluens visade fria kromosomer från celler i metafas (celldelningsfas) där det fanns en svag signal för X-kromosomen. Multicolor-FISH-analys av VMM1 och H1915 gav tydliga resultat i fluorescensmikroskop där fria kromosomer var Multicolor-probeinmärkta; blå/aqua, röd och grön. Konklusionen är att vid kromosomutvinning från odlade adherenta celler bör dessa vara 80 % konfluenta. Detta för att ge tydliga och reproducerbara probeinfärgningar av kromosomer i metafas vid analys med Multicolor-FISH. Analys av 80 % konfluenta celler och användning av Multicolor-FISH-analys är en klar förbättring av kurslaborationen. 

    Fulltekst (pdf)
    fulltext
  • 108. Altai, M.
    et al.
    Wallberg, H.
    Honarvar, H.
    Strand, J.
    Orlova, A.
    Löfblom, John
    KTH, Skolan för bioteknologi (BIO), Proteinteknologi.
    Varasteh, Z.
    Sandström, M.
    Ståhl, Stefan
    KTH, Skolan för bioteknologi (BIO), Proteinteknologi.
    Tolmachev, V.
    Re-188-Z(HER2: V2), a promising targeting agent against HER2-expressing tumors: in vitro and in vivo assessment2013Inngår i: European Journal of Nuclear Medicine and Molecular Imaging, ISSN 1619-7070, E-ISSN 1619-7089, Vol. 40, s. S119-S119Artikkel i tidsskrift (Annet vitenskapelig)
  • 109.
    Altai, Mohamed
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Medicinsk strålningsvetenskap. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi.
    Leitao, Charles Dahlsson
    KTH Royal Inst Technol, Dept Prot Sci, Sch Engn Sci Chem Biotechnol & Hlth, S-10691 Stockholm, Sweden.
    Rinne, Sara S.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Theranostics.
    Vorobyeva, Anzhelika
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Medicinsk strålningsvetenskap.
    Atterby, Christina
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Medicinsk strålningsvetenskap.
    Ståhl, Stefan
    KTH Royal Inst Technol, Dept Prot Sci, Sch Engn Sci Chem Biotechnol & Hlth, S-10691 Stockholm, Sweden.
    Tolmachev, Vladimir
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Medicinsk strålningsvetenskap.
    Löfblom, John
    KTH Royal Inst Technol, Dept Prot Sci, Sch Engn Sci Chem Biotechnol & Hlth, S-10691 Stockholm, Sweden.
    Orlova, Anna
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Theranostics. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Influence of Molecular Design on the Targeting Properties of ABD-Fused Mono- and Bi-Valent Anti-HER3 Affibody Therapeutic Constructs2018Inngår i: CELLS, ISSN 2073-4409, Vol. 7, nr 10, artikkel-id 164Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Overexpression of human epidermal growth factor receptor type 3 (HER3) is associated with tumour cell resistance to HER-targeted therapies. Monoclonal antibodies (mAbs) targeting HER3 are currently being investigated for treatment of various types of cancers. Cumulative evidence suggests that affibody molecules may be appropriate alternatives to mAbs. We previously reported a fusion construct (3A3) containing two HER3-targeting affibody molecules flanking an engineered albumin-binding domain (ABD 035) included for the extension of half-life in circulation. The 3A3 fusion protein (19.7 kDa) was shown to delay tumour growth in mice bearing HER3-expressing xenografts and was equipotent to the mAb seribantumab. Here, we have designed and explored a series of novel formats of anti-HER3 affibody molecules fused to the ABD in different orientations. All constructs inhibited heregulin-induced phosphorylation in HER3-expressing BxPC-3 and DU-145 cell lines. Biodistribution studies demonstrated extended the half-life of all ABD-fused constructs, although at different levels. The capacity of our ABD-fused proteins to accumulate in HER3-expressing tumours was demonstrated in nude mice bearing BxPC-3 xenografts. Formats where the ABD was located on the C-terminus of affibody binding domains (3A, 33A, and 3A3) provided the best tumour targeting properties in vivo. Further development of these promising candidates for treatment of HER3-overexpressing tumours is therefore justified.

    Fulltekst (pdf)
    FULLTEXT01
  • 110.
    Altai, Mohamed
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Medicinsk strålningsvetenskap.
    Liu, Hao
    KTH Royal Inst Technol, Dept Prot Sci, Roslagstullsbacken 21, S-11417 Stockholm, Sweden.
    Ding, Haozhong
    KTH Royal Inst Technol, Dept Prot Sci, Roslagstullsbacken 21, S-11417 Stockholm, Sweden.
    Mitran, Bogdan
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Theranostics.
    Edqvist, Per-Henrik D
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Experimentell och klinisk onkologi.
    Tolmachev, Vladimir
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Medicinsk strålningsvetenskap.
    Orlova, Anna
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Medicinsk strålningsvetenskap.
    Gräslund, Torbjorn
    KTH Royal Inst Technol, Dept Prot Sci, Roslagstullsbacken 21, S-11417 Stockholm, Sweden.
    Affibody-derived drug conjugates: Potent cytotoxic molecules for treatment of HER2 over-expressing tumors2018Inngår i: Journal of Controlled Release, ISSN 0168-3659, E-ISSN 1873-4995, Vol. 288, s. 84-95Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Patients with HER2-positive tumors often suffer resistance to therapy, warranting development of novel treatment modalities. Affibody molecules are small affinity proteins which can be engineered to bind to desired targets. They have in recent years been found to allow precise targeting of cancer specific molecular signatures such as the HER2 receptor. In this study, we have investigated the potential of an affibody molecule targeting HER2, Z(HER2:2891), conjugated with the cytotoxic maytansine derivate MC-DM1, for targeted cancer therapy. Z(HER2:2891) was expressed as a monomer (Z(HER2:2891)), dimer ((Z(HER2:2891)) 2) and dimer with an albumin binding domain (ABD) for half-life extension ((Z(HER2:2891)) 2-ABD). All proteins had a unique C-terminal cysteine that could be used for efficient and site-specific conjugation with MC-DM1. The resulting affibody drug conjugates were potent cytotoxic molecules for human cells over-expressing HER2, with sub-nanomolar IC50-values similar to trastuzumab emtansine, and did not affect cells with low HER2 expression. A biodistribution study of a radiolabeled version of (Z(HER2:2891))(2)-ABD-MC-DM1, showed that it was taken up by the tumor. The major site of off-target uptake was the kidneys and to some extent the liver. (Z(HER2:2891)) 2-ABD-MC-DM1 was found to have a half-life in circulation of 14 h. The compound was tolerated well by mice at 8.5 mg/kg and was shown to extend survival of mice bearing HER2 over-expressing tumors. The findings in this study show that affibody molecules are a promising class of engineered affinity proteins to specifically deliver small molecular drugs to cancer cells and that such conjugates are potential candidates for clinical evaluation on HER2-overexpressing cancers.

  • 111. Altai, Mohamed
    et al.
    Strand, Joanna
    Rosik, Daniel
    Selvaraju, Ram Kumar
    Eriksson Karlström, Amelie
    KTH, Skolan för bioteknologi (BIO), Molekylär Bioteknologi. KTH, Skolan för bioteknologi (BIO), Centra, Albanova VinnExcellence Center for Protein Technology, ProNova.
    Orlova, Anna
    Tolmachev, Vladimir
    Influence of Nuclides and Chelators on Imaging Using Affibody Molecules: Comparative Evaluation of Recombinant Affibody Molecules Site-Specifically Labeled with Ga-68 and In-111 via Maleimido Derivatives of DOTA and NODAGA2013Inngår i: Bioconjugate chemistry, ISSN 1043-1802, E-ISSN 1520-4812, Vol. 24, nr 6, s. 1102-1109Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Accurate detection of cancer-associated molecular abnormalities in tumors could make cancer treatment more of personalized. Affibody molecules enable high contrast imaging of tumor-associated protein expression shortly after injection. The use should increase sensitivity of HER2 imaging. The chemical nature of the generator-produced positron-emitting radionuclide Ga-68 of radionuclides and chelators influences the biodistribution of Affibody molecules, providing an opportunity to further increase the imaging contrast. The aim of the study was to compare maleimido derivatives of DOTA and NODAGA for site-specific labeling of a recombinant Z(HER2:2395) HER2-binding Affibody molecule with Ga-68. DOTA and NODAGA were site-specifically conjugated to the Z(HER2:2395) Affibody molecule having a C-terminal cysteine and labeled with Ga-68 and In-111. All labeled conjugates retained specificity to HER2 in vitro. Most of the cell-associated activity was membrane-bound with a minor difference in internalization rate. All variants demonstrated specific targeting of xenografts and a high tumor uptake. The xenografts were dearly visualized using all conjugates. The influence of chelator on the biodistribution and targeting properties was much less pronounced for Ga-68 than for In-111. The tumor uptake of Ga-68-NODAGA-Z(HER2:2395) and Ga-68-NODAGA-Z(HER2:2395) and tumor-to-blood ratios at 2 h p.i. did not differ significantly. However, the tumor-to-liver ratio was significantly higher for Ga-68-NODAGA- Z(HER2:2395) (8 +/- 2 vs 5.0 +/- 0.3) offering the advantage of better liver metastases visualization. In conclusion, influence of chelators on biodistribution of Affibody molecules depends on the radionuclides and reoptimization of labeling chemistry is required when a radionuclide label is changed.

  • 112. Altai, Mohamed
    et al.
    Wållberg, Helena
    KTH, Skolan för bioteknologi (BIO), Molekylär Bioteknologi.
    Orlova, Anna
    Rosestedt, Maria
    Hosseinimehr, Seyed Jalal
    Tolmachev, Vladimir
    Ståhl, Stefan
    KTH, Skolan för bioteknologi (BIO), Molekylär Bioteknologi.
    Order of amino acids in C-terminal cysteine-containing peptide-based chelators influences cellular processing and biodistribution of Tc-99m-labeled recombinant Affibody molecules2012Inngår i: Amino Acids, ISSN 0939-4451, E-ISSN 1438-2199, Vol. 42, nr 5, s. 1975-1985Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Affibody molecules constitute a novel class of molecular display selected affinity proteins based on non-immunoglobulin scaffold. Preclinical investigations and pilot clinical data have demonstrated that Affibody molecules provide high contrast imaging of tumor-associated molecular targets shortly after injection. The use of cysteine-containing peptide-based chelators at the C-terminus of recombinant Affibody molecules enabled site-specific labeling with the radionuclide Tc-99m. Earlier studies have demonstrated that position, composition and the order of amino acids in peptide-based chelators influence labeling stability, cellular processing and biodistribution of Affibody molecules. To investigate the influence of the amino acid order, a series of anti-HER2 Affibody molecules, containing GSGC, GEGC and GKGC chelators have been prepared and characterized. The affinity to HER2, cellular processing of Tc-99m-labeled Affibody molecules and their biodistribution were investigated. These properties were compared with that of the previously studied Tc-99m-labeled Affibody molecules containing GGSC, GGEC and GGKC chelators. All variants displayed picomolar affinities to HER2. The substitution of a single amino acid in the chelator had an appreciable influence on the cellular processing of Tc-99m. The biodistribution of all Tc-99m-labeled Affibody molecules was in general comparable, with the main difference in uptake and retention of radioactivity in excretory organs. The hepatic accumulation of radioactivity was higher for the lysine-containing chelators and the renal retention of Tc-99m was significantly affected by the amino acid composition of chelators. The order of amino acids influenced renal uptake of some conjugates at 1 h after injection, but the difference decreased at later time points. Such information can be helpful for the development of other scaffold protein-based imaging and therapeutic radiolabeled conjugates.

  • 113.
    Altuvia, S.
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Teknisk-naturvetenskapliga fakulteten, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Wagner, E.G.H.
    Switching on and off with RNA.2000Inngår i: Proc. Natl. Acad. Sci. USA, Vol. 97, nr 18, s. 9824-9826Artikkel i tidsskrift (Fagfellevurdert)
  • 114.
    Alvarez-Rodriguez, Manuel
    et al.
    Linköpings universitet, Institutionen för biomedicinska och kliniska vetenskaper, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Martinez-Serrano, Cristina
    Linköpings universitet, Institutionen för biomedicinska och kliniska vetenskaper, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    Wright, Dominic
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Biologi. Linköpings universitet, Tekniska fakulteten.
    Barranco, Isabel
    Univ Girona, Spain.
    Roca, Jordi
    Univ Murcia, Spain.
    Rodriguez-Martinez, Heriberto
    Linköpings universitet, Institutionen för biomedicinska och kliniska vetenskaper, Avdelningen för barns och kvinnors hälsa. Linköpings universitet, Medicinska fakulteten.
    The Transcriptome of Pig Spermatozoa, and Its Role in Fertility2020Inngår i: International Journal of Molecular Sciences, ISSN 1422-0067, E-ISSN 1422-0067, INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES, Vol. 21, nr 5, artikkel-id 1572Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    In the study presented here we identified transcriptomic markers for fertility in the cargo of pig ejaculated spermatozoa using porcine-specific micro-arrays (GeneChip((R)) miRNA 4.0 and GeneChip((R)) Porcine Gene 1.0 ST). We report (i) the relative abundance of the ssc-miR-1285, miR-16, miR-4332, miR-92a, miR-671-5p, miR-4334-5p, miR-425-5p, miR-191, miR-92b-5p and miR-15b miRNAs, and (ii) the presence of 347 up-regulated and 174 down-regulated RNA transcripts in high-fertility breeding boars, based on differences of farrowing rate (FS) and litter size (LS), relative to low-fertility boars in the (Artificial Insemination) AI program. An overrepresentation analysis of the protein class (PANTHER) identified significant fold-increases for C-C chemokine binding (GO:0019957): CCR7, which activates B- and T-lymphocytes, 8-fold increase), XCR1 and CXCR4 (with ubiquitin as a natural ligand, 1.24-fold increase), cytokine receptor activity (GO:0005126): IL23R receptor of the IL23 protein, associated to JAK2 and STAT3, 3.4-fold increase), the TGF-receptor (PC00035) genes ACVR1C and ACVR2B (12-fold increase). Moreover, two micro-RNAs (miR-221 and mir-621) were down- and up-regulated, respectively, in high-fertility males. In conclusion, boars with different fertility performance possess a wide variety of differentially expressed RNA present in spermatozoa that would be attractive targets as non-invasive molecular markers for predicting fertility.

    Fulltekst (pdf)
    fulltext
  • 115.
    Al-walai, Somar
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för biologisk grundutbildning. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för neurovetenskap.
    Characterization of solutecarrier SLC38A62012Student paper other, 10 poäng / 15 hpOppgave
    Abstract [en]

    Transport across the membrane of a cell is of crucial importance for cellular functions. The solute carrier family,SLC38 is a family of membrane proteins that transports various substances through the membrane and thusperforms many physiologically important functions, for example, transport of glutamine from astrocyte toneurons in the central nervous system. In this paper, we demonstrate that one of the transporters in this familynamed SLC38A6 forms several protein complexes with a variety of proteins in the membrane and in synapticvesicles, suggesting that SLC38A6 is involved in the synaptic release of neurotransmitters in synapses. Weperformed sensitive protein interaction analysis between the protein of interest and a variety of proteinsexpressed at different sites in the neuronal cell. We showed that SLC38A6 interacts with proteins in the cellmembrane as well as in the membrane of synaptic vesicles. The current theory is that SLC38A6 interact withthese proteins when the synaptic vesicles are in close proximity with the cell membrane during the release of theneurotransmitters.

    Fulltekst (pdf)
    Report_SLC38A6_solute_carrier_Somar_Al-walai_Uppsala_University_Sweden
  • 116.
    Amelina, Hanna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Proteomics in biomarker research: Insights into the effects of aging and environment on biological systems2011Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Proteomics is the global analysis of proteins that covers a broad range of technologies aimed at determining the identity and quantity of proteins expressed in the cell, their three-dimensional structure and interaction partners. In contrast to genome, proteome reflects more accurately on the dynamic state of the cell, tissue, or an organism. Therefore much is expected from proteomics to yield better disease markers for early diagnosis and therapy monitoring, as well as biomarkers that would indicate environmental exposure or provide prediction of biological age.

    In this thesis, I have developed and applied robust and sensitive subproteomic approaches to study the effect of aging as well as and environmental pollution using different animal models.

    In the first part, a high-throughput proteomic method based on liquid chromatography coupled to 2-dimensional gel electrophoresis (LC/2-DE) was developed. The usefulness of this method has been demonstrated by applying it to the assessment of marine pollution in a field experiment.

    Next, I have utilized this subproteomic approach to study the effect of aging in mouse kidney of both genders. As a result, a protein expression signature of aging kidney was obtained, revealing gender-dependent alterations in proteome profiles of aging mouse kidney.

    In order to further reduce the dynamic range of protein expression and increase the sensitivity of proteomic analysis, I have applied a shotgun mass spectrometry-based proteomic approach using isobaric tags for relative and absolute quantification (iTRAQ) coupled to liquid chromatography and tandem mass spectrometry (LC-MS/MS) to study age-related differences in peroxisome-enriched fractions from mouse liver. Only eight proteins showed statistically significant difference in expression (p<0.05) with moderate folds. This study indicates that age-depended changes in the liver proteome are minimal, suggesting that its proteome is efficiently maintained until certain age.

    Finally, in the context of aging studies and the role of peroxisomes in aging, I have tested the utility of cell-penetrating peptides (CPPs) as agents for protein delivery into acatalasemic peroxisomes using yeast as a model. The results obtained suggest that CPPs may be suitable for the delivery of antioxidants to peroxisomes and in future could provide a tool for the protein therapy of age-related diseases.

    Fulltekst (pdf)
    FULLTEXT01
  • 117.
    Amelina, Hanna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Apraiz, Itxaso
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Sun, Wei
    Cristobal, Susana
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Proteomics-based method for the assessment of marine pollution using liquid chromatography coupled with two-dimensional electrophoresis2007Inngår i: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 6, nr 6, s. 2094-2104Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Using a proteomic approach, we have developed a new method for the assessment of marine pollution that generates highly reproducible protein expression patterns and it is simple and scalable. The protocol is based on applying liquid chromatography (LC) coupled with two-dimensional electrophoresis (2-DE) to analyze changes in the protein expression pattern after exposure to marine pollution. The digestive gland of the sentinel “blue mussel” (Mytilus edulis) was batch-processed through a simple cell fractionation followed by ion-exchange chromatography and 2-DE. The selection of ligands, elution method, and small volume design was carefully considered to define a protocol that could be mainly robotized. A pilot study with samples collected from different Gothenburg harbor areas indicated that the clean area could be distinguished from the polluted ones based on a protein expression pattern (PES) composed of 13 proteins. Principal component analysis (PCA) and hierarchical clustering confirmed that the PES was sufficient to discriminate polluted and unpolluted areas and to provide a spatial gradient from the polluted source. Several proteins from the PES were identified by electrospray ionization tandem mass spectrometry (ESI−MS/MS), and they are involved in β-oxidation, amino acid metabolism, detoxification, protein degradation, organelle biogenesis, and protein folding. In the near future, this methodology could show potential advantages to assess marine pollution and could become a stable platform to elucidate ecotoxicological questions.

  • 118.
    Amelina, Hanna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Cristobal, Susana
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Proteomic study on gender differences in aging kidney of mice2009Inngår i: Proteome Science, ISSN 1477-5956, E-ISSN 1477-5956, Vol. 7, nr 16Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    BACKGROUND: This study aims to analyze sex differences in mice aging kidney. We applied a proteomic technique based on subfractionation, and liquid chromatography coupled with 2-DE. Samples from male and female CD1-Swiss outbred mice from 28 weeks, 52 weeks, and 76 weeks were analysed by 2-DE, and selected proteins were identified by matrix assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS).

    RESULTS: This proteomic analysis detected age-related changes in protein expression in 55 protein-spots, corresponding to 22 spots in males and 33 spots in females. We found a protein expression signature (PES) of aging composed by 8 spots, common for both genders. The identified proteins indicated increases in oxidative and proteolytic proteins and decreases in glycolytic proteins, and antioxidant enzymes.

    CONCLUSION: Our results provide insights into the gender differences associated to the decline of kidney function in aging. Thus, we show that proteomics can provide valuable information on age-related changes in expression levels of proteins and related modifications. This pilot study is still far from providing candidates for aging-biomarkers. However, we suggest that the analysis of these proteins could suggest mechanisms of cellular aging in kidney, and improve the kidney selection for transplantation.

  • 119.
    Amelina, Hanna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Sjodin, Marcus O. D.
    Bergquist, Jonas
    Cristobal, Susana
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Quantitative subproteomic analysis of age-related changes in mouse liver peroxisomes by iTRAQ LC-MS/MS2011Inngår i: Journal of chromatography. B, ISSN 1570-0232, E-ISSN 1873-376X, Vol. 879, nr 30, s. 3393-3400Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Aging is a complex multifactorial phenomenon, which is believed to result from the accumulation of cellular damage to biological macromolecules. Peroxisomes recently emerged as another important source of reactive oxygen species (ROS) production in addition to mitochondria. However, the role of these organelles in the process of aging is still not clear. The aim of this study was to characterize the changes in protein expression profiles of young (10 weeks old) versus old (18 months old) mouse liver peroxisome-enriched fractions. We have applied shotgun proteomic approach based on liquid chromatography and tandem mass spectrometry (LC-MS/MS) combined with iTRAQ (isobaric tags for relative and absolute quantitation) labeling that allows comparative quantitative multiplex analysis. Our analysis led to identification and quantification of 150 proteins, 8 out of which were differentially expressed between two age groups at a statistically significant level (p < 0.05), with folds ranging from 1.2 to 4.1. These proteins involved in peroxisornal beta-oxidation, detoxification of xenobiotics and production of ROS. Noteworthy, differences in liver proteome have been observed between as well as within different age groups. In conclusion, our subproteomic quantitative study suggests that mouse liver proteome is sufficiently maintained until certain age.

  • 120.
    Ament-Velasquez, Sandra Lorena
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för organismbiologi, Systematisk biologi. Univ Montpellier, Inst Evolutionary Sci, CNRS, IRD,EPHE, Pl Eugene Bataillon, F-34095 Montpellier, France..
    Figuet, E.
    Univ Montpellier, Inst Evolutionary Sci, CNRS, IRD,EPHE, Pl Eugene Bataillon, F-34095 Montpellier, France..
    Ballenghien, M.
    Univ Montpellier, Inst Evolutionary Sci, CNRS, IRD,EPHE, Pl Eugene Bataillon, F-34095 Montpellier, France..
    Zattara, E. E.
    Indiana Univ, Dept Biol, 107 S Indiana Ave, Bloomington, IN 47405 USA.;Smithsonian Inst, Natl Museum Nat Hist, Dept Invertebrate Zool, 10th St & Constitut Ave NW, Washington, DC 20560 USA..
    Norenburg, J. L.
    Smithsonian Inst, Natl Museum Nat Hist, Dept Invertebrate Zool, 10th St & Constitut Ave NW, Washington, DC 20560 USA..
    Fernandez-Alvarez, F. A.
    CSIC Barcelona, Inst Ciencies Mar, Barcelona 08003, Spain..
    Bierne, J.
    Univ Reims, Lab Biol Cellulaire & Mol, 9 Blvd Paix, F-51100 Reims, France..
    Bierne, N.
    Univ Montpellier, Inst Evolutionary Sci, CNRS, IRD,EPHE, Pl Eugene Bataillon, F-34095 Montpellier, France..
    Galtier, N.
    Univ Montpellier, Inst Evolutionary Sci, CNRS, IRD,EPHE, Pl Eugene Bataillon, F-34095 Montpellier, France..
    Population genomics of sexual and asexual lineages in fissiparous ribbon worms (Lineus, Nemertea): hybridization, polyploidy and the Meselson effect2016Inngår i: Molecular Ecology, ISSN 0962-1083, E-ISSN 1365-294X, Vol. 25, nr 14, s. 3356-3369Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Comparative population genetics in asexual vs. sexual species offers the opportunity to investigate the impact of asexuality on genome evolution. Here, we analyse coding sequence polymorphism and divergence patterns in the fascinating Lineus ribbon worms, a group of marine, carnivorous nemerteans with unusual regeneration abilities, and in which asexual reproduction by fissiparity is documented. The population genomics of the fissiparous L. pseudolacteus is characterized by an extremely high level of heterozygosity and unexpectedly elevated pi(N)/pi(S) ratio, in apparent agreement with theoretical expectations under clonal evolution. Analysis of among-species allele sharing and read-count distribution, however, reveals that L. pseudolacteus is a triploid hybrid between Atlantic populations of L. sanguineus and L. lacteus. We model and quantify the relative impact of hybridity, polyploidy and asexuality on molecular variation patterns in L. pseudolacteus and conclude that (i) the peculiarities of L. pseudolacteus population genomics result in the first place from hybridization and (ii) the accumulation of new mutations through the Meselson effect is more than compensated by processes of heterozygosity erosion, such as gene conversion or gene copy loss. This study illustrates the complexity of the evolutionary processes associated with asexuality and identifies L. pseudolacteus as a promising model to study the first steps of polyploid genome evolution in an asexual context.

  • 121.
    Amer, Ayad
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Costa, Tiago
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Farag, Salah
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Avican, Ummehan
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Forsberg, Åke
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Francis, Matthew
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Genetically engineered frameshifted YopN-TyeA chimeras influence type III secretion system function in Yersinia pseudotuberculosis2013Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, nr 10, artikkel-id e77767Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Type III secretion is a tightly controlled virulence mechanism utilized by many gram negative bacteria to colonize their eukaryotic hosts. To infect their host, human pathogenic Yersinia spp. translocate protein toxins into the host cell cytosol through a preassembled Ysc-Yop type III secretion device. Several of the Ysc-Yop components are known for their roles in controlling substrate secretion and translocation. Particularly important in this role is the YopN and TyeA heterodimer. In this study, we confirm that Y. pseudotuberculosis naturally produce a 42 kDa YopN-TyeA hybrid protein as a result of a +1 frame shift near the 3 prime of yopN mRNA, as has been previously reported for the closely related Y. pestis. To assess the biological role of this YopN-TyeA hybrid in T3SS by Y. pseudotuberculosis, we used in cis site-directed mutagenesis to engineer bacteria to either produce predominately the YopN-TyeA hybrid by introducing +1 frame shifts to yopN after codon 278 or 287, or to produce only singular YopN and TyeA polypeptides by introducing yopN sequence from Y. enterocolitica, which is known not to produce the hybrid. Significantly, the engineered 42 kDa YopN-TyeA fusions were abundantly produced, stable, and were efficiently secreted by bacteria in vitro. Moreover, these bacteria could all maintain functionally competent needle structures and controlled Yops secretion in vitro. In the presence of host cells however, bacteria producing the most genetically altered hybrids (+1 frameshift after 278 codon) had diminished control of polarized Yop translocation. This corresponded to significant attenuation in competitive survival assays in orally infected mice, although not at all to the same extent as Yersinia lacking both YopN and TyeA proteins. Based on these studies with engineered polypeptides, most likely a naturally occurring YopN-TyeA hybrid protein has the potential to influence T3S control and activity when produced during Yersinia-host cell contact.

    Fulltekst (pdf)
    fulltext
  • 122.
    Amer, Ayad
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Gurung, Jyoti
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Costa, Tiago
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Ruuth, Kristina
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Zavialov, Anton
    Joint Biotechnology Laboratory, Department of Chemistry, University of Turku, Turku, Finland.
    Forsberg, Åke
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Francis, Matthew S
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    YopN and TyeA Hydrophobic Contacts Required for Regulating Ysc-Yop Type III Secretion Activity by Yersinia pseudotuberculosis2016Inngår i: Frontiers in Cellular and Infection Microbiology, E-ISSN 2235-2988, Vol. 6, artikkel-id 66Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Yersinia bacteria target Yop effector toxins to the interior of host immune cells by the Ysc-Yop type III secretion system. A YopN-TyeA heterodimer is central to controlling Ysc-Yop targeting activity. A + 1 frameshift event in the 3-prime end of yopN can also produce a singular secreted YopN-TyeA polypeptide that retains some regulatory function even though the C-terminal coding sequence of this YopN differs greatly from wild type. Thus, this YopN C-terminal segment was analyzed for its role in type III secretion control. Bacteria producing YopN truncated after residue 278, or with altered sequence between residues 279 and 287, had lost type III secretion control and function. In contrast, YopN variants with manipulated sequence beyond residue 287 maintained full control and function. Scrutiny of the YopN-TyeA complex structure revealed that residue W279 functioned as a likely hydrophobic contact site with TyeA. Indeed, a YopNW279G mutant lost all ability to bind TyeA. The TyeA residue F8 was also critical for reciprocal YopN binding. Thus, we conclude that specific hydrophobic contacts between opposing YopN and TyeA termini establishes a complex needed for regulating Ysc-Yop activity.

    Fulltekst (pdf)
    fulltext
  • 123.
    Ameur, Adam
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab. Monash Univ, Sch Publ Hlth & Prevent Med, Melbourne, Vic, Australia.
    Kloosterman, Wigard P.
    UMC Utrecht, Dept Genet, Ctr Mol Med, NL-3584 CG Utrecht, Netherlands.
    Hestand, Matthew S.
    Cincinnati Childrens Hosp Med Ctr, Div Human Genet, Cincinnati, OH 45229 USA.
    Single-Molecule Sequencing: Towards Clinical Applications2019Inngår i: Trends in Biotechnology, ISSN 0167-7799, E-ISSN 1879-3096, Vol. 37, nr 1, s. 72-85Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    In the past several years, single-molecule sequencing platforms, such as those by Pacific Biosciences and Oxford Nanopore Technologies, have become available to researchers and are currently being tested for clinical applications. They offer exceptionally long reads that permit direct sequencing through regions of the genome inaccessible or difficult to analyze by short-read platforms. This includes disease-causing long repetitive elements, extreme GC content regions, and complex gene loci. Similarly, these platforms enable structural variation characterization at previously unparalleled resolution and direct detection of epigenetic marks in native DNA. Here, we review how these technologies are opening up new clinical avenues that are being applied to pathogenic microorganisms and viruses, constitutional disorders, pharmaco-genomics, cancer, and more.

  • 124.
    Amir, Mohd.
    et al.
    Jamia Millia Islamia, Ctr Interdisciplinary Res Basic Sci, New Delhi 110025, India.
    Kumar, Vijay
    Amity Univ Noida, Amity Inst Neuropsychol & Neurosci, Noida, UP, India.
    Dohare, Ravins
    Jamia Millia Islamia, Ctr Interdisciplinary Res Basic Sci, New Delhi 110025, India;King Saud Univ, Dept Pharmacogmosy, Coll Pharm, Riyadh 11451, Saudi Arabia.
    Rehman, Md. Tabish
    King Saud Univ, Dept Pharmacogmosy, Coll Pharm, Riyadh 11451, Saudi Arabia.
    Hussain, Afzal
    King Saud Univ, Dept Pharmacogmosy, Coll Pharm, Riyadh 11451, Saudi Arabia.
    Alajmi, Mohamed F.
    King Saud Univ, Dept Pharmacogmosy, Coll Pharm, Riyadh 11451, Saudi Arabia.
    El-Seedi, Hesham
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Farmakognosi.
    Hassan, Hani Mutlak A.
    King Abdulaziz Univ, King Fand Med Res Ctr, POB 80216, Jeddah 21589, Saudi Arabia;King Abdulaziz Univ, Fac Appl Med Sci, Dept Med Lab Technol, POB 80216, Jeddah 21589, Saudi Arabia.
    Islam, Asimul
    Jamia Millia Islamia, Ctr Interdisciplinary Res Basic Sci, New Delhi 110025, India.
    Ahmad, Faizan
    Jamia Millia Islamia, Ctr Interdisciplinary Res Basic Sci, New Delhi 110025, India.
    Hassan, Md. Imtaiyaz
    Jamia Millia Islamia, Ctr Interdisciplinary Res Basic Sci, New Delhi 110025, India.
    Investigating architecture and structure-function relationships in cold shock DNA-binding domain family using structural genomics-based approach2019Inngår i: International Journal of Biological Macromolecules, ISSN 0141-8130, E-ISSN 1879-0003, Vol. 133, s. 484-494Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Oligonucleotide/oligosaccharide-binding fold (OB-fold) plays a major role in the regulation of central dogma of life via binding though DNA and RNA. The OB-fold domains are diverse in nature and present in large number of proteins with verities of molecular functions. Here, we have investigated the distribution of sequence, structure and repeats of cold shock DNA-binding proteins (CSDB), a member of OB-fold, in all three kingdoms to establish functional relationships. The CSDB is consists of 30 domains with a major contribution of S1 (>110,601 sequences), S12 (>23,760 sequences), S17 (>14,833 sequences) and S28e (>1615 sequence) domains. These domains are largely found in bacteria (70-90%). The number of S1 domain repeats in eukaryota varies from 1 to 15 and are well-correlated with the protein size. The molecular function analysis suggests that a large number of repeats in the S1 domain are involved in diverse molecular functions in bacteria and eukaryotes. In-depth structure analysis of Si, S12, S17 and S28e domain-containing proteins of the OB-fold family provides a reasonable basis to understand the relationship of size and number of repeats with the corresponding molecular functions.

  • 125.
    Amiri, H.
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för evolutionsbiologi, Molekylär evolution.
    Alsmark, U. C. M.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för evolutionsbiologi, Molekylär evolution.
    Andersson, S. G. E.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för evolutionsbiologi, Molekylär evolution.
    Proliferation and deterioration of Rickettsia palindromic elements2002Inngår i: Molecular biology and evolution, ISSN 0737-4038, E-ISSN 1537-1719, Vol. 19, nr 8, s. 1234-1243Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    It has been suggested that Rickettsia Palindromic Elements (RPEs) have evolved as selfish DNA that mediate protein sequence evolution by being targeted to genes that code for RNA and proteins. Here, we have examined the phylogenetic depth of two RPEs that are located close to the genes encoding elongation factors Tu (tuf) and G (fus) in Rickettsia. An exceptional organization of the elongation factor genes was found in all 11 species examined, with complete or partial RPEs identified downstream of the tuf gene (RPE-tuf) in six species and of the fus gene (RPE-fus) in 10 species. A phylogenetic reconstruction shows that both RPE-tuf and RPE-fus have evolved in a manner that is consistent with the expected species divergence. The analysis provides evidence for independent loss of RPE-tuf in several species, possibly mediated by short repetitive sequences flanking the site of excision. The remaining RPE-tuf sequences evolve as neutral sequences in different stages of deterioration. Likewise, highly fragmented remnants of the RPE-fus sequence were identified in two species. This suggests that genome-specific differences in the content of RPEs are the result of recent loss rather than recent proliferation.

  • 126.
    Amiri, H.
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Teknisk-naturvetenskapliga fakulteten, Biologiska sektionen, Institutionen för evolutionsbiologi. MOLECULAR EVOLUTION.
    Andersson, S. G. E.
    Rickettsia: The highly rearranged cousin of mitochondria.2001Inngår i: Recent Res. Devel. Microbiology, Vol. 5, s. 321-329Artikkel i tidsskrift (Fagfellevurdert)
  • 127.
    Amiri, H.
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Teknisk-naturvetenskapliga fakulteten, Biologiska sektionen, Institutionen för evolutionsbiologi. MOLECULAR EVOLUTION.
    Davids, W.
    Andersson, S.G.E.
    Birth and death of orphan genes in Rickettsia.2003Inngår i: Mol. Biol. Evol., Vol. 20, s. 1575-1587Artikkel i tidsskrift (Fagfellevurdert)
  • 128.
    Amiri, H.
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Teknisk-naturvetenskapliga fakulteten, Biologiska sektionen, Institutionen för evolutionsbiologi. MOLECULAR EVOLUTION.
    Karlberg, O.
    Andersson, S.G.E.
    Deep origin of plastid/parasite ATP/ADP translocases.2003Inngår i: J Mol Evol, Vol. 56, s. 137-150Artikkel i tidsskrift (Fagfellevurdert)
  • 129.
    Ampomah, Osei Yaw
    et al.
    University of Tromso.
    Jensen, John Beck
    University of Tromso.
    Bhuvaneswari, T V
    University of Tromso.
    Lack of trehalose catabolism in Sinorhizobium species increases their nodulation competitiveness on certain host genotypes2008Inngår i: New Phytologist, ISSN 0028-646X, E-ISSN 1469-8137, Vol. 179, nr 2, s. 495-504Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The role of host and bacterial genotypes in determining the competitiveness of trehalose utilization mutants of Sinorhizobium meliloti and Sinorhizobium medicae was investigated here. Trehalose utilization mutants of S. meliloti and S. medicae were obtained by mutagenesis of their trehalose utilization gene thuB. The mutant strains and the wild type were coinoculated on three cultivars of alfalfa (Medicago sativa) and two cultivars of Medicago truncatula and assessed for competitiveness in root colonization, and nodule occupancy. The thuB mutants formed more nodules than their parent strains on two of the three alfalfa lines tested and on one of the two M. truncatula lines tested. They were not more competitive on the other alfalfa and M. truncatula lines. Their competitiveness for nodule occupancy did not correlate positively with their ability to colonize these roots but correlated with the extent of thuB induction in the infection threads. Induction of thuB was shown to be dependent on the concentration of trehalose in the environment. These results suggest a direct role for host trehalose metabolism in early plant-symbiont interactions and show that the ability to manage host-induced stresses during infection, rather than the ability to colonize the root, is critical for competitive nodulation.

  • 130.
    Amrein, Beat A.
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Bauer, Paul
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Duarte, Fernanda
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Janfalk Carlsson, Åsa
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Kemiska sektionen, Institutionen för kemi - BMC, Biokemi.
    Naworyta, Agata
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Mowbray, Sherry L.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Widersten, Mikael
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Kemiska sektionen, Institutionen för kemi - BMC, Biokemi.
    Kamerlin, Shina C. L.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Expanding the catalytic triad in epoxide hydrolases and related enzymes2015Inngår i: ACS Catalysis, ISSN 2155-5435, E-ISSN 2155-5435, Vol. 5, nr 10, s. 5702-5713Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Potato epoxide hydrolase 1 exhibits rich enantio- and regioselectivity in the hydrolysis of a broadrange of substrates. The enzyme can be engineered to increase the yield of optically pureproducts, as a result of changes in both enantio- and regioselectivity. It is thus highly attractive inbiocatalysis, particularly for the generation of enantiopure fine chemicals and pharmaceuticals.The present work aims to establish the principles underlying the activity and selectivity of theenzyme through a combined computational, structural, and kinetic study, using the substratetrans-stilbene oxide as a model system. Extensive empirical valence bond simulations have beenperformed on the wild-type enzyme together with several experimentally characterized mutants.We are able to computationally reproduce the differences in activities between differentstereoisomers of the substrate, and the effects of mutations in several active-site residues. Inaddition, our results indicate the involvement of a previously neglected residue, H104, which iselectrostatically linked to the general base, H300. We find that this residue, which is highlyconserved in epoxide hydrolases and related hydrolytic enzymes, needs to be in its protonatedform in order to provide charge balance in an otherwise negatively-charged active site. Our datashow that unless the active-site charge balance is correctly treated in simulations, it is notpossible to generate a physically meaningful model for the enzyme that can accurately reproduceactivity and selectivity trends. We also expand our understanding of other catalytic residues,demonstrating in particular the role of a non-canonical residue, E35, as a “backup-base” in theabsence of H300. Our results provide a detailed view of the main factors driving catalysis andregioselectivity in this enzyme, and identify targets for subsequent enzyme design efforts.

    Fulltekst (pdf)
    fulltext
  • 131.
    Amrein, Beat Anton
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Extending the Reach of Computational Approaches to Model Enzyme Catalysis2017Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

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

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

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

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

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

    Fulltekst (pdf)
    fulltext
    Download (jpg)
    preview image
  • 132.
    An, Junxue
    et al.
    KTH, Skolan för kemivetenskap (CHE), Kemi, Yt- och korrosionsvetenskap.
    Dédinaité, Andra
    KTH, Skolan för kemivetenskap (CHE), Kemi, Yt- och korrosionsvetenskap.
    Nilsson, Anki
    Holgersson, Jan
    Claesson, Per M.
    KTH, Skolan för kemivetenskap (CHE), Kemi, Yt- och korrosionsvetenskap.
    Comparison of a Brush-with-Anchor and a Train-of-Brushes Mucin on Poly(methyl methacrylate) Surfaces: Adsorption, Surface Forces, and Friction2014Inngår i: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 15, nr 4, s. 1515-1525Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Interfacial properties of two types of mucins have been investigated at the aqueous solution/poly(methyl methacrylate) (PMMA) interface. One is commercially available bovine submaxillary mucin, BSM, which consists of alternating glycosylated and nonglycosylated regions. The other one is a recombinant mucin-type fusion protein, PSGL-1/mIgG(2b), consisting of a glycosylated mucin part fused to the Fc part of an immunoglobulin. PSGL-1/mIgG(2b) is mainly expressed as a (timer upon production. A quartz crystal microbalance with dissipation was used to study the adsorption of the mucins to PMMA surfaces. The mass of the adsorbed mucin layers, including the adsorbed mucin and water trapped in the layer, was found to be significantly higher for PSGL-1/mIgG(2b) than for BSM. Atomic force microscopy with colloidal probe was employed to study interactions and frictional forces between mucin-coated PMMA surfaces. Purely repulsive forces of steric origin were Observed between PSGL-1/mIgG(2b) mucin layers, whereas a small adhesion was detected between BSM layers and attributed to bridging. Both mucin layers reduced the friction force between PMMA surfaces in aqueous solution. The reduction was, however, significantly more pronounced for PSGL-1/mIgG(2b). The effective friction coefficient between PSGL-1/mIgG(2b)-coated PMMA surfaces is as low as 0.02 at low loads, increasing to 0.24 at the highest load explored, 50 nN. In contrast, a friction coefficient of around 0.7 was obtained between BSM-coated PMMA surfaces. The large differences in interfacial properties for the two mucins are discussed in relation to their structural differences.

  • 133.
    Anandapadamanaban, Madhanagopal
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Kemi. Linköpings universitet, Tekniska fakulteten.
    Structural insights into protein-protein interactions governing regulation in transcription initiation and ubiquitination2015Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Virtually every aspect of the cellular processes in eukaryotes requires that the interactions between protein molecules are well coordinated in different regulatory pathways. Any protein dysfunction involved in these regulatory pathways might lead to various pathological conditions. Understanding the structural and functional peculiarities of these proteins molecular machineries will help in formulating structure-based drug design.

    The first regulatory process studied here is the RNA polymerase-II mediated transcription of the eukaryotic protein-coding genes to produce mRNAs. This process requires the formation of the ‘transcription initiation’ by the assembly of Pre-Initiation Complex (PIC) formation at a core promoter region. Regulation at this initiation level is a key mechanism for the control of gene expression that governs cellular growth and differentiation. The transcription Factor IID (TFIID) is a conserved multiprotein general transcription factor with an essential role in  nucleating the PIC formation, composed of TATA Binding Protein (TBP) and about 14 TBP Associated Factors (TAFs). The here presented crystal structure (1.97Å) of TBP bound to TAND1 and TAND2 domains from TAF1 reveals a detailed molecular pattern of interactions involving both transcriptionally activating and repressing regions in TBP, thereby uncovering central principles for anchoring of TBP-binding motifs. Together with NMR and cellular analysis, this work provides the structural basis of competitive binding with TFIIA to modulate TBP in promoter recognition.

    In eukaryotes, another fundamental mechanism in the regulation of cellular physiology is the posttranslational modification of substrate proteins by ubiquitin, termed ‘ubiquitination’. Important actors in this mechanism are the ubiquitin-ligases (E3s) that culminate the transfer of ubiquitin to the substrate and govern the specificity of this system. One E3 ligase in particular, TRIM21, defines a subgroup of the Tripartite Motif (TRIM) family, which belongs to the major RING-type of E3 ubiquitin ligases, and plays an important role in pathogenesis of autoimmunity by mediating ubiquitination of transcription factors. The crystal structure (2.86Å) of the RING domain from TRIM21 in complex with UBE2E1, an E2 conjugating enzyme, together with the NMR and SAXS analysis as well as biochemical functional analysis, reveals the molecular basis for the dynamic binding interfaces. The TRIM21 mode of ubiquitin recognition and activation for catalytic transfer of ubiquitin can be modeled onto the entire TRIM family.

    Finally, we explored the concepts of conformational selection in proteins as a possible key component for protein-mediated transcriptional regulation. In this framework, MexR, a bacterial repressor of the MexAB-OprM efflux pump, and its mutant Arg21Trp were studied as an example for proteins presenting different conformations. The residue Arg21Trp mutation is clinically identified to cause of Multi-Drug Resistant (MDR) by attenuated DNA binding, and leads to the overexpression of the MexAB-OprM efflux pump. With the crystal structure (2.19Å) of MexR mutant Arg21Trp, in combination with MD-simulations and SAXS for both wild-type and mutant, we could unravel the atomic details of the wild-type conformations consisting in subsets of populations of DNA bound and unbound forms. Remarkably, the mutant Arg21Trp stabilize the DNA unbound state and shifts MexR in a pre-existing equilibrium, from a repressed to a derepressed state.

    Taken together, these studies substantially broaden our knowledge at a molecular level in protein interactions that are involved in transcriptional regulation and ubiquitination, studied by a carefully selected combination of complementary structural methods spanning different resolutions and time scales.

    Fulltekst (pdf)
    fulltext
    Download (pdf)
    omslag
    Download (jpg)
    presentationsbild
  • 134.
    Anandapadamanaban, Madhanagopal
    et al.
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Kemi. Linköpings universitet, Tekniska fakulteten.
    Kyriakidis, Nikolaos
    Rheumatology Unit, Department of Medicine, Center for Molecular Medicine L8:04, Karolinska Institutet, Stockholm, Sweden.
    Espinosa, Alexander
    Rheumatology Unit, Department of Medicine, Center for Molecular Medicine L8:04, Karolinska Institutet, Stockholm, Sweden.
    Round, Adam R.
    European Molecular Biology Laboratory, Grenoble Outstation, Grenoble, France.
    Trewhella, Jill
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Kemi. Linköpings universitet, Tekniska fakulteten. School of Molecular Bioscience, The University of Sydney, New South Wales, Australia.
    Wahren-Herlenius, Marie
    Rheumatology Unit, Department of Medicine, Center for Molecular Medicine L8:04, Karolinska Institutet, Stockholm, Sweden.
    Moche, Martin
    Department of Medical Biochemistry and Biophysics, Protein Science Facility, Karolinska Institutet, Stockholm, Sweden.
    Sunnerhagen, Maria
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Kemi. Linköpings universitet, Tekniska fakulteten.
    Structure of a TRIM21 - UBE2El complex reveals the specificity of E2 and ubiquitin recognition by TRIM E3 RINGsManuskript (preprint) (Annet vitenskapelig)
    Abstract [en]

    TRIM21, a RlNG-containing E3 ubiquitin-ligase of the TRIM   protein family, is a major autoantigen in SLE and Sjögren's syndrome as well as a modifier of interferon regulatory factors, thereby regulating innate immune signalling. We herein report the 2.86 Å crystal structure ofhuman TRIM211-91 comprising the RING domain (residues 16-55), in complex with the human E2 conjugating UBE2El enzyme (also denoted UbcH6). The crystal structure, joint with analysis by NMR and SAXS as well as structure-directed mutations and functional assays provides a detailed view of the specificity-determining contacts that support specific E2 recognition in the TRIM family. A detailed comparison of our structure with known E2 bound ubiquitin complexes, supported by biochemical analyses, reveals the molecular basis for TRIM21 interactions with donor ubiquitin that activates catalytic ubiquitin transfer. Finally, our structure convincingly demonstrates the placement of the Ub-targeted Lys61 of the adjacent TRIM211- 91 close to the catalytically active UBE2El cysteine, and how the Lys61 amide is activated fora nucleophilic attack by hydrogen-bondeffected deshielding by conserved acidic residues at the E2 active site. In all, our structural findings provide molecular details ofthe selectivity involved in TRIM21 interactions with its cognate UBE2E1 enzyme and how TRIM21 positions ubiquitin in a catalytic conformation for ubiquitin transfer, and presents a snapshot of the Ub ligation step on a specific target residue of TRIM211-91 as an auto-ubiquitinated pseudo-substrate at high concentration. Increased structural and functional understanding of the TRIM mediated ubiquitination will aid development ofnovel therapeutic approaches in the entire TRIM family ofproteins.

  • 135.
    Andaloussi, Mounir
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för organisk farmaceutisk kemi.
    Henriksson, Lena M.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi.
    Wieckowska, Anna
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för organisk farmaceutisk kemi.
    Lindh, Martin
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för organisk farmaceutisk kemi.
    Björkelid, Christofer
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi.
    Larsson, Anna M.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi.
    Suresh, Surisetti
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för organisk farmaceutisk kemi.
    Iyer, Harini
    Srinivasa, Bachally R.
    Bergfors, Terese
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi.
    Unge, Torsten
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi.
    Mowbray, Sherry L.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi.
    Larhed, Mats
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för organisk farmaceutisk kemi.
    Jones, T. Alwyn
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Struktur- och molekylärbiologi.
    Karlén, Anders
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för organisk farmaceutisk kemi.
    Design, Synthesis, and X-ray Crystallographic Studies of alpha-Aryl Substituted Fosmidomycin Analogues as Inhibitors of Mycobacterium tuberculosis 1-Deoxy-D-xylulose 5-Phosphate Reductoisomerase2011Inngår i: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 54, nr 14, s. 4964-4976Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The natural antibiotic fosmidomycin acts via inhibition of 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR), an essential enzyme in the non-mevalonate pathway of isoprenoid biosynthesis. Fosmidomycin is active on Mycobacterium tuberculosis DXR (MtDXR), but it lacks antibacterial activity probably because of poor uptake. alpha-Aryl substituted fosmidomycin analogues have more favorable physicochemical properties and are also more active in inhibiting malaria parasite growth. We have solved crystal structures of MtDXR in complex with 3,4-dichlorophenyl substituted fosmidomycin analogues; these show important differences compared to our previously described forsmidomycin-DXR complex. Our best inhibitor has an IC(50) = 0.15 mu M on MtDXR but still lacked activity in a mycobacterial growth assay (MIC > 32 mu g/mL). The combined results, however, provide insights into how DXR accommodates the new inhibitors and serve as an excellent starting point for the design of other novel and more potent inhibitors, particularly against pathogens where uptake is less of a problem, such as the malaria parasite.

    Fulltekst (pdf)
    fulltext
  • 136. Anderbrant, Olle
    et al.
    Matteson, Donald S.
    Unelius, C. Rikard
    Pharazyn, Philip S.
    Santangelo, Ellen M.
    KTH, Skolan för kemivetenskap (CHE), Kemi, Organisk kemi.
    Schlyter, Fredrik
    Birgersson, Goran
    Pheromone of the elm bark beetle Scolytus laevis (Coleoptera Scolytidae): stereoisomers of 4-methyl-3-heptanol reduce interspecific competition2010Inngår i: Chemoecology, ISSN 0937-7409, E-ISSN 1423-0445, Vol. 20, nr 3, s. 179-187Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Stereoisomers of 4-methyl-3-heptanol (MH) are pheromone components of several Scolytus bark beetles. The elm bark beetle Scolytus laevis (Coleoptera: Scolytidae) has in previous studies been caught in traps baited with commercial MH containing all four stereoisomers, but the lure has been considered a weak attractant. In this study, we addressed the question whether stereospecific responses by S. laevis to stereoisomers of MH might contribute to its niche separation from other sympatric Scolytus species. Using GC-MS, we analyzed extracts of hindguts and abdomens from male and female S. laevis and the sympatric S. triarmatus. We also tested all four MH-stereoisomers individually and in combinations in the field to determine their role for S. laevis. All four stereoisomers were synthesized via a boronic ester method with 1,2-dicyclohexylethanediol as chiral director. In addition, the (3S,4R)-stereoisomer of MH was prepared through enantioselective, lipase-mediated transesterification of a mixture of the four stereoisomers of MH. Females of both species contained small amounts of syn-MH, and males contained trace amounts of anti-MH. The anti stereoisomer (3R,4S)-MH was attractive to male and female S. laevis, whereas the syn stereoisomer (3S,4S)-MH acted as an inhibitor or deterrent and reduced the catch when added to the attractive isomer. The syn isomer is the main aggregation pheromone component of the larger and sympatric S. scolytus and possibly also of S. triarmatus. The avoidance response of S. laevis to the (3S,4S)-stereoisomer may reduce interspecific competition for host trees.

  • 137. Andersen, Felicie F.
    et al.
    Knudsen, Bjarne
    Oliveira, Cristiano Luis Pinto
    Frøhlich, Rikke F.
    Krüger, Dinna
    Bungert, Jörg
    Agbandje-McKenna, Mavis
    McKenna, Robert
    Juul, Sissel
    Veigaard, Christopher
    Koch, Jørn
    Rubinstein, John L.
    Guldbrandtsen, Bernt
    Hede, Marianne S.
    Karlsson, Göran
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Kemiska sektionen, Institutionen för fysikalisk och analytisk kemi, Fysikalisk kemi.
    Andersen, Anni H.
    Pedersen, Jan Skov
    Knudsen, Birgitta R.
    Assembly and structural analysis of a covalently closed nano-scale DNA cage2008Inngår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 36, nr 4, s. 1113-1119Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The inherent properties of DNA as a stable polymer with unique affinity for partner mols. detd. by the specific Watson-Crick base pairing makes it an ideal component in self-assembling structures. This has been exploited for decades in the design of a variety of artificial substrates for investigations of DNA-interacting enzymes. More recently, strategies for synthesis of more complex two-dimensional (2D) and 3D DNA structures have emerged. However, the building of such structures is still in progress and more experiences from different research groups and different fields of expertise are necessary before complex DNA structures can be routinely designed for the use in basal science and/or biotechnol. Here we present the design, construction and structural anal. of a covalently closed and stable 3D DNA structure with the connectivity of an octahedron, as defined by the double-stranded DNA helixes that assembles from eight oligonucleotides with a yield of .apprx.30%. As demonstrated by Small Angle X-ray Scattering and cryo-Transmission Electron Microscopy analyses the eight-stranded DNA structure has a central cavity larger than the apertures in the surrounding DNA lattice and can be described as a nano-scale DNA cage, Hence, in theory it could hold proteins or other bio-mols. to enable their investigation in certain harmful environments or even allow their organization into higher order structures.

  • 138. Andersen, Gorm
    et al.
    Andersen, Birgit
    Dobritzsch, Doreen
    Karolinska Institutet.
    Schnackerz, Klaus D
    Piskur, Jure
    A gene duplication led to specialized gamma-aminobutyrate and beta-alanine aminotransferase in yeast2007Inngår i: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 274, nr 7, s. 1804-1017Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    In humans, beta-alanine (BAL) and the neurotransmitter gamma-aminobutyrate (GABA) are transaminated by a single aminotransferase enzyme. Apparently, yeast originally also had a single enzyme, but the corresponding gene was duplicated in the Saccharomyces kluyveri lineage. SkUGA1 encodes a homologue of Saccharomyces cerevisiae GABA aminotransferase, and SkPYD4 encodes an enzyme involved in both BAL and GABA transamination. SkPYD4 and SkUGA1 as well as S. cerevisiae UGA1 and Schizosaccharomyces pombe UGA1 were subcloned, over-expressed and purified. One discontinuous and two continuous coupled assays were used to characterize the substrate specificity and kinetic parameters of the four enzymes. It was found that the cofactor pyridoxal 5'-phosphate is needed for enzymatic activity and alpha-ketoglutarate, and not pyruvate, as the amino group acceptor. SkPyd4p preferentially uses BAL as the amino group donor (V(max)/K(m)=0.78 U x mg(-1) x mm(-1)), but can also use GABA (V(max)/K(m)=0.42 U x mg(-1) x mm(-1)), while SkUga1p only uses GABA (V(max)/K(m)=4.01 U x mg(-1) x mm(-1)). SpUga1p and ScUga1p transaminate only GABA and not BAL. While mammals degrade BAL and GABA with only one enzyme, but in different tissues, S. kluyveri and related yeasts have two different genes/enzymes to apparently 'distinguish' between the two reactions in a single cell. It is likely that upon duplication approximately 200 million years ago, a specialized Uga1p evolved into a 'novel' transaminase enzyme with broader substrate specificity.

  • 139.
    Andersen, Malin
    et al.
    KTH, Skolan för bioteknologi (BIO), Genteknologi.
    Engström, Pär
    Lithwick, Stuart
    Arenillas, David
    Eriksson, Per
    Lenhard, Boris
    Wasserman, Wyeth
    Odeberg, Jacob
    KTH, Skolan för bioteknologi (BIO), Genteknologi.
    In silico detection of sequence variations modifying transcriptional regulation2008Inngår i: PloS Computational Biology, ISSN 1553-734X, E-ISSN 1553-7358, Vol. 4, nr 1, s. e5-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Identification of functional genetic variation associated with increased susceptibility to complex diseases can elucidate genes and underlying biochemical mechanisms linked to disease onset and progression. For genes linked to genetic diseases, most identified causal mutations alter an encoded protein sequence. Technological advances for measuring RNA abundance suggest that a significant number of undiscovered causal mutations may alter the regulation of gene transcription. However, it remains a challenge to separate causal genetic variations from linked neutral variations. Here we present an in silico driven approach to identify possible genetic variation in regulatory sequences. The approach combines phylogenetic footprinting and transcription factor binding site prediction to identify variation in candidate cis-regulatory elements. The bioinformatics approach has been tested on a set of SNPs that are reported to have a regulatory function, as well as background SNPs. In the absence of additional information about an analyzed gene, the poor specificity of binding site prediction is prohibitive to its application. However, when additional data is available that can give guidance on which transcription factor is involved in the regulation of the gene, the in silico binding site prediction improves the selection of candidate regulatory polymorphisms for further analyses. The bioinformatics software generated for the analysis has been implemented as a Web-based application system entitled RAVEN ( regulatory analysis of variation in enhancers). The RAVEN system is available at http://www.cisreg.ca for all researchers interested in the detection and characterization of regulatory sequence variation.

  • 140.
    Andersen, Malin
    et al.
    KTH, Skolan för bioteknologi (BIO), Genteknologi.
    Lenhard, Boris
    Whatling, Carl
    Eriksson, Per
    Odeberg, Jacob
    KTH, Skolan för bioteknologi (BIO), Genteknologi.
    Alternative promoter usage of the membrane glycoprotein CD362006Inngår i: BMC Molecular Biology, ISSN 1471-2199, E-ISSN 1471-2199, Vol. 7, s. 8-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: CD36 is a membrane glycoprotein involved in a variety of cellular processes such as lipid transport, immune regulation, hemostasis, adhesion, angiogenesis and atherosclerosis. It is expressed in many tissues and cell types, with a tissue specific expression pattern that is a result of a complex regulation for which the molecular mechanisms are not yet fully understood. There are several alternative mRNA isoforms described for the gene. We have investigated the expression patterns of five alternative first exons of the CD36 gene in several human tissues and cell types, to better understand the molecular details behind its regulation.

    Results: We have identified one novel alternative first exon of the CD36 gene, and confirmed the expression of four previously known alternative first exons of the gene. The alternative transcripts are all expressed in more than one human tissue and their expression patterns vary highly in skeletal muscle, heart, liver, adipose tissue, placenta, spinal cord, cerebrum and monocytes. All alternative first exons are upregulated in THP-1 macrophages in response to oxidized low density lipoproteins. The alternative promoters lack TATA-boxes and CpG islands. The upstream region of exon 1b contains several features common for house keeping gene and monocyte specific gene promoters.

    Conclusion: Tissue-specific expression patterns of the alternative first exons of CD36 suggest that the alternative first exons of the gene are regulated individually and tissue specifically. At the same time, the fact that all first exons are upregulated in THP-1 macrophages in response to oxidized low density lipoproteins may suggest that the alternative first exons are coregulated in this cell type and environmental condition. The molecular mechanisms regulating CD36 thus appear to be unusually complex, which might reflect the multifunctional role of the gene in different tissues and cellular conditions.

  • 141.
    Anderson, Judy E.
    et al.
    Department of Human Anatomy and Cell Science, University of Manitoba, Winnipeg, Canada; Manitoba Institute of Child's Health (MICH), University of Manitoba, Winnipeg, Canada.
    Hansen, Lise Lotte
    Institute of Human Genetics, University of Aarhus, Denmark.
    Mooren, Frank C.
    Department of Sports Medicine, Institute of Sport Sciences, University Giessen, Germany.
    Post, Markus
    Department of Sports Medicine, Institute of Sport Sciences, University Giessen, Germany.
    Hug, Hubert
    DSM Nutritional Products Ltd, Research & Development, Kaiseraugst, Switzerland.
    Zuse, Anne
    Manitoba Institute of Cell Biology (MICB), CancerCare Manitoba, 675 McDermot Ave. Rm. ON6010, Winnipeg, Man. R3E 0V9, Canada.
    Los, Marek Jan
    Manitoba Institute of Cell Biology, Cancer Care Manitoba; Manitoba Institute of Child Health; Department of Biochemistry and Medical Genetics; Department of Human Anatomy and Cell Science, University Manitoba, Winnipeg, Canada, .
    Methods and biomarkers for the diagnosis and prognosis of cancer and other diseases: Towards personalized medicine2006Inngår i: Drug resistance updates, ISSN 1368-7646, E-ISSN 1532-2084, Vol. 9, nr 4-5, s. 198-210Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The rapid development of new diagnostic procedures, the mapping of the human genome, progress in mapping genetic polymorphisms, and recent advances in nucleic acid- and protein chip technologies are driving the development of personalized therapies. This breakthrough in medicine is expected to be achieved largely due to the implementation of "lab-on-the-chip" technology capable of performing hundreds, even thousands of biochemical, cellular and genetic tests on a single sample of blood or other body fluid. Focusing on a few disease-specific examples, this review discusses selected technologies and their combinations likely to be incorporated in the "lab-on-the-chip" and to provide rapid and versatile information about specific diseases entities. Focusing on breast cancer and after an overview of single-nucleofide polymorphism (SNP)-screening methodologies, we discuss the diagnostic and prognostic importance of SNPs. Next, using Duchenne muscular dystrophy (DMD) as an example, we provide a brief overview of powerful and innovative integration of traditional immuno-histochemistry techniques with advanced biophysical methods such as NMR-spectroscopy or Fourier-transformed infrared (FT-IR) spectroscopy. A brief overview of the challenges and opportunities provided by protein and aptamer microarrays follows. We conclude by highlighting novel and promising biochemical markers for the development of personalized treatment of cancer and other diseases: serum cytochrome c, cytokeratin-18 and -19 and their proteolytic fragments for the detection and quantitation of malignant tumor mass, tumor cell turn-over, inflammatory processes during hepatitis and Epstein-Barr virus (EBV)-induced hemophagocytic lymphohistiocytosis and apoptotic/necrotic cancer cell death. (c) 2006 Elsevier Ltd. All rights reserved.

    Fulltekst (pdf)
    fulltext
  • 142.
    Anderson, Mattias
    KTH, Skolan för bioteknologi (BIO), Industriell bioteknologi.
    Amine Transaminases in Multi-Step One-Pot Reactions2017Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Amine transaminases are enzymes that catalyze the mild and selective formation of primary amines, which are useful building blocks for biologically active compounds and natural products. In order to make the production of these kinds of compounds more efficient from both a practical and an environmental point of view, amine transaminases were incorporated into multi-step one-pot reactions. With this kind of methodology there is no need for isolation of intermediates, and thus unnecessary work-up steps can be omitted and formation of waste is prevented. Amine transaminases were successfully combined with other enzymes for multi-step synthesis of valuable products: With ketoreductases all four diastereomers of a 1,3-amino alcohol could be obtained, and the use of a lipase allowed for the synthesis of natural products in the form of capsaicinoids. Amine transaminases were also successfully combined with metal catalysts based on palladium or copper. This methodology allowed for the amination of alcohols and the synthesis of chiral amines such as the pharmaceutical compound Rivastigmine. These examples show that the use of amine transaminases in multi-step one-pot reactions is possible, and hopefully this concept can be further developed and applied to make industrial processes more sustainable and efficient in the future.

    Fulltekst (pdf)
    fulltext
  • 143.
    Andersson, Anders
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för evolution, genomik och systematik, Molekylär evolution.
    Bernander, Rolf
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för evolution, genomik och systematik, Molekylär evolution.
    Nilsson, Peter
    Dual-genome primer design for construction of DNA microarrays2005Inngår i: Bioinformatics, ISSN 1367-4803, E-ISSN 1367-4811, Vol. 21, nr 3, s. 325-332Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Motivation: Microarray experiments using probes covering a whole transcriptome are expensive to initiate, and a major part of the costs derives from synthesizing gene-specific PCR primers or hybridization probes. The high costs may force researchers to limit their studies to a single organism, although comparing gene expression in different species would yield valuable information.

    Results: We have developed a method, implemented in the software DualPrime, that reduces the number of primers required to amplify the genes of two different genomes. The software identifies regions of high sequence similarity, and from these regions selects PCR primers shared between the genomes, such that either one or, preferentially, both primers in a given PCR can be used for amplification from both genomes. To assure high microarray probe specificity, the software selects primer pairs that generate products of low sequence similarity to other genes within the same genome. We used the software to design PCR primers for 2182 and 1960 genes from the hyperthermophilic archaea Sulfolobus solfataricus and Sulfolobus acidocaldarius, respectively. Primer pairs were shared among 705 pairs of genes, and single primers were shared among 1184 pairs of genes, resulting in a saving of 31% compared to using only unique primers. We also present an alternative primer design method, in which each gene shares primers with two different genes of the other genome, enabling further savings.

  • 144.
    Andersson, Annika
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Kudva, Renuka
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Magoulopoulou, Anastasia
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Lejarre, Quentin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Lara, Patricia
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Xu, Peibo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Goel, Suchi
    Pissi, Jennifer
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Ru, Xing
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Hessa, Tara
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Wahlgren, Mats
    von Heijne, Gunnar
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Karolinska Institutet, Sweden.
    Nilsson, IngMarie
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Tellgren-Roth, Åsa
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Membrane integration and topology of RIFIN and STEVOR proteins of the Plasmodium falciparum parasite2019Inngår i: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The malarial parasite Plasmodium exports its own proteins to the cell surfaces of red blood cells (RBCs) during infection. Examples of exported proteins include members of the repetitive interspersed family (RIFIN) and subtelomeric variable open reading frame (STEVOR) family of proteins from Plasmodium falciparum. The presence of these parasite-derived proteins on surfaces of infected RBCs triggers the adhesion of infected cells to uninfected cells (rosetting) and to the vascular endothelium potentially obstructing blood flow. While there is a fair amount of information on the localization of these proteins on the cell surfaces of RBCs, less is known about how they can be exported to the membrane and the topologies they can adopt during the process. The first step of export is plausibly the cotranslational insertion of proteins into the endoplasmic reticulum (ER) of the parasite, and here, we investigate the insertion of three RIFIN and two STEVOR proteins into the ER membrane. We employ a well-established experimental system that uses N-linked glycosylation of sites within the protein as a measure to assess the extent of membrane insertion and the topology it assumes when inserted into the ER membrane. Our results indicate that for all the proteins tested, transmembranes (TMs) 1 and 3 integrate into the membrane, so that the protein assumes an overall topology of Ncyt-Ccyt. We also show that the segment predicted to be TM2 for each of the proteins likely does not reside in the membrane, but is translocated to the lumen.

  • 145.
    Andersson, August
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Mäler, Lena
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Magnetic resonance investigations of lipid motion in isotropic bicelles2005Inngår i: Journal of the American Chemical Society, ISSN 0002-7863, E-ISSN 1520-5126, Vol. 21, nr 7, s. 7702-7709Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The dynamics of DMPC in different isotropic bicelles have been investigated by NMR and EPR methods. The local dynamics were obtained by interpretation of 13C NMR relaxation measurements of DMPC in the bicelles, and these results were compared to EPR spectra of spin-labeled lipids. The overall size of the bicelles was investigated by PFG NMR translational diffusion measurements. The dynamics and relative sizes were compared among three different bicelles: [DMPC]/[DHPC] = 0.25, [DMPC]/[DHPC] = 0.5, and [DMPC]/[CHAPS] = 0.5. The local motion is found to depend much more strongly on the choice of the detergent, rather than the overall size of the bicelle. The results provide an explanation for differences in apparent dynamics for different peptides, which are bound to bicelles. This in turn determines under what conditions reasonable NMR spectra can be observed. A model is presented in which extensive local motion, in conjunction with the overall size, affects the spectral properties. An analytical expression for the size dependence of the bicelles, relating the radius of the bilayer region with physical properties of the detergent and the lipid, is also presented.

  • 146.
    Andersson, C. Evalena
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Teknisk-naturvetenskapliga fakulteten, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Structure-Function Studies of Enzymes from Ribose Metabolism2004Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    In the pentose phosphate pathway, carbohydrates such as glucose and ribose are degraded with production of reductive power and energy. Another important function is to produce essential pentoses, such as ribose 5-phosphate, which later can be used in biosynthesis of nucleic acids and cofactors.

    This thesis presents structural and functional studies on three enzymes involved in ribose metabolism in Escherichia coli.

    Ribokinase is an enzyme that phosphorylates ribose in the presence of ATP and magnesium, as the first step of exogenous ribose metabolism. Two important aspects of ribokinase function, not previously known, have been elucidated. Ribokinase was shown to be activated by monovalent cations, specifically potassium. Structural analysis of the monovalent ion binding site indicates that the ion has a structural rather than catalytic role; a mode of activation involving a conformational change has been suggested. Product inhibition studies suggest that ATP is the first substrate to bind the enzyme. Independent Kd measurements with the ATP analogue AMP-PCP support this. The results presented here will have implications for several enzymes in the protein family to which ribokinase belongs, in particular the medically interesting enzyme adenosine kinase.

    Ribose 5-phosphate isomerases convert ribose 5-phosphate into ribulose 5-phosphate or vice versa. Structural studies on the two genetically distinct isomerases in E. coli have shown them to be fundamentally different in many aspects, including active site architecture. However, a kinetic study has demonstrated both enzymes to be efficient in terms of catalysis. Sequence searches of completed genomes show ribose 5-phosphate isomerase B to be the sole isomerase in many bacteria, although ribose 5-phosphate isomerase A is a nearly universal enzyme. All genomes contain at least one of the two enzymes. These results confirm that both enzymes must be independently capable of supporting ribose metabolism, a fact that had not previously been established.

    Fulltekst (pdf)
    FULLTEXT01
  • 147.
    Andersson, Charlotta
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi.
    Li, Xingru
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi.
    Lorenz, Fryderyk
    Umeå universitet, Medicinska fakulteten, Institutionen för strålningsvetenskaper, Onkologi.
    Golovleva, Irina
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Medicinsk och klinisk genetik.
    Wahlin, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för strålningsvetenskaper, Onkologi.
    Li, Aihong
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi.
    Reduction in WT1 Gene Expression During Early Treatment Predicts the Outcome in Patients With Acute Myeloid Leukemia2012Inngår i: Diagnostic molecular pathology (Print), ISSN 1052-9551, E-ISSN 1533-4066, Vol. 21, nr 4, s. 225-233Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Wilms tumor gene 1 (WT1) expression has been suggested as an applicable minimal residual disease marker in acute myeloid leukemia (AML). We evaluated the use of this marker in 43 adult AML patients. Quantitative assessment of WT1 gene transcripts was performed using real-time quantitative-polymerase chain reaction assay. Samples from both the peripheral blood and the bone marrow were analyzed at diagnosis and during follow-up. A strong correlation was observed between WT1 normalized with 2 different control genes (beta-actin and ABL1, P < 0.001). WT1 mRNA level at diagnosis was of no prognostic relevance (P > 0.05). A >= 1-log reduction in WT1 expression in bone marrow samples taken < 1 month after diagnosis significantly correlated with an improved overall survival (P = 0.004) and freedom from relapse (P = 0.010) when beta-actin was used as control gene. Furthermore, a reduction in WT1 expression by >= 2 logs in peripheral blood samples taken at a later time point significantly correlated with a better outcome for overall survival (P = 0.004) and freedom from relapse (P = 0.012). This result was achieved when normalizing against both b-actin and ABL1. These results therefore suggest that WT1 gene expression can provide useful information for minimal residual disease detection in adult AML patients and that combined use of control genes can give more informative results.

  • 148.
    Andersson, Charlotta S.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Berthold, Catrine L.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Högbom, Martin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    A dynamic c terminal segment in the mycobacterium tuberculosis mn/fe r2lox protein can adopt a helical structure with possible functional consequences2012Inngår i: Chemistry and Biodiversity, ISSN 1612-1872, E-ISSN 1612-1880, Vol. 9, nr 9, s. 1981-1988Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Mycobacterium tuberculosis R2-like ligand-binding oxidase (MtR2lox) belongs to a recently discovered group of proteins that are homologous to the ribonucleotide reductase R2 proteins. MtR2lox carries a heterodinuclear Mn/Fe cofactor and, unlike R2 proteins, a large ligand-binding cavity. A unique tyrosine-valine cross link is also found in the vicinity of the active site. To date, all known structures of R2 and R2lox proteins show a disordered C-terminal segment. Here, we present two new crystal forms of MtR2lox, revealing an ordered helical C-terminal. The ability of alternating between an ordered and disordered state agrees well with bioinformatic analysis of the protein sequence. Interestingly, ordering of the C-terminal helix shields a large positively charged patch on the protein surface, potentially used for interaction with other cellular components. We hypothesize that the dynamic C-terminal segment may be involved in control of protein function in vivo.

    Fulltekst (pdf)
    fulltext
  • 149.
    Andersson, Charlotta S.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Lundgren, Camilla A. K.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Magnúsdóttir, Auður
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Ge, Changrong
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Wieslander, Åke
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Martinez Molina, Daniel
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Högbom, Martin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    The Mycobacterium tuberculosis Very-Long-Chain Fatty Acyl-CoA Synthetase: Structural Basis for Housing Lipid Substrates Longer than the Enzyme2012Inngår i: Structure, ISSN 0969-2126, E-ISSN 1878-4186, Vol. 20, nr 6, s. 1062-1070Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The Mycobacterium tuberculosis acid-induced operon MymA encodes the fatty acyl-CoA synthetase FadD13 and is essential for virulence and intracellular growth of the pathogen. Fatty acyl-CoA synthetases activate lipids before entering into the metabolic pathways and are also involved in transmembrane lipid transport. Unlike soluble fatty acyl-CoA synthetases, but like the mammalian integral-membrane very-long-chain acyl-CoA synthetases, FadD13 accepts lipid substrates up to the maximum length tested (C-26). Here, we show that FadD13 is a peripheral membrane protein. The structure and mutational studies reveal an arginine- and aromatic-rich surface patch as the site for membrane interaction. The protein accommodates a hydrophobic tunnel that extends from the active site toward the positive patch and is sealed by an arginine-rich lid-loop at the protein surface. Based on this and previous data, we propose a structural basis for accommodation of lipid substrates longer than the enzyme and transmembrane lipid transport by vectorial CoA-esterification.

    Fulltekst (pdf)
    fulltext
  • 150.
    Andersson, Charlotta Selina
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Structural studies of R2 and R2–like proteins with a heterodinuclear Mn/Fe cofactor and enzymes involved in Mycobacterium tuberculosis lipid metabolism2012Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

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

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

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

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

    Fulltekst (pdf)
    Fulltext
1234567 101 - 150 of 6026
RefereraExporteraLink til resultatlisten
Permanent link
Referera
Referensformat
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Annet format
Fler format
Språk
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Annet språk
Fler språk
Utmatningsformat
  • html
  • text
  • asciidoc
  • rtf