| 1. |
- Bengtson, Per, et al.
(författare)
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Identification of a Missense Mutation (G329A; Arg110→ Gln) in the Human FUT7 Gene
- 2001
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 276:34, s. 31575-31582
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Tidskriftsartikel (refereegranskat)abstract
- The human FUT7 gene codes for the α1,3-fucosyltransferase VII (Fuc-TVII), which is involved in the biosynthesis of the sialyl Lewis x (SLex) epitope on human leukocytes. The FUT7 gene has so far been considered to be monomorphic. Neutrophils isolated from patients with ulcerative colitis were examined for apparent alterations in protein glycosylation patterns by Western blot analysis using monoclonal antibodies directed against SLex and SLex-related epitopes. One individual showed lower levels of SLex expression and an elevated expression of CD65s compared to controls. The coding regions of the FUT7 gene from this individual were cloned, and a G329A point mutation (Arg110 → Gln) was found in one allele, whereas the other FUT7 allele was wild type. No Fuc-TVII enzyme activity was detected in COS-7 cells transiently transfected with the mutated FUT7 construct. TheFUT7 Arg110 is conserved in all previously cloned vertebrate α1,3-fucosyltransferases. Polymerase chain reaction followed by restriction enzyme cleavage was used to screen 364 unselected Caucasians for the G329A mutation, and a frequency of ≤1% for this mutation was found (3 heterozygotes). Genetic characterization of the family members of one of the additional heterozygotes identified one individual carrying the G329A mutation in both FUT7alleles. Peripheral blood neutrophils of this homozygously mutated individual showed a lowered expression of SLex and an elevated expression of CD65s when analyzed by Western blot and flow cytometry. The homozygous individual was diagnosed with ulcer disease, non-insulin-dependent diabetes, osteoporosis, spondyloarthrosis, and Sjögren's syndrome but had no history of recurrent bacterial infections or leukocytosis.
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| 2. |
- Noborn, Fredrik, et al.
(författare)
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Subtyping of cardiac amyloidosis by mass spectrometry-based proteomics of endomyocardial biopsies.
- 2023
-
Ingår i: Amyloid : the international journal of experimental and clinical investigation : the official journal of the International Society of Amyloidosis. - : Informa UK Limited. - 1350-6129 .- 1744-2818. ; 30:1, s. 96-108
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Tidskriftsartikel (refereegranskat)abstract
- Cardiac amyloidosis is a severe condition leading to restrictive cardiomyopathy and heart failure. Mass spectrometry-based methods for cardiac amyloid subtyping have become important diagnostic tools but are currently used only in a few reference laboratories. Such methods include laser-capture microdissection to ensure the specific analysis of amyloid deposits. Here we introduce a direct proteomics-based method for subtyping of cardiac amyloidosis.Endomyocardial biopsies were retrospectively analysed from fresh frozen material of 78 patients with cardiac amyloidosis and from 12 biopsies of unused donor heart explants. Cryostat sections were digested with trypsin and analysed with liquid chromatography - mass spectrometry, and data were evaluated by proteomic software.With a diagnostic threshold set to 70% for each of the four most common amyloid proteins affecting the heart (LC κ, LC λ, TTR and SAA), 65 of the cases (87%) could be diagnosed, and of these, 61 cases (94%) were in concordance with the original diagnoses. The specimens were also analysed for the summed intensities of the amyloid signature proteins (ApoE, ApoA-IV and SAP). The intensities were significantly higher (p<0.001) for all assigned cases compared with controls.Cardiac amyloidosis can be successfully subtyped without the prior enrichment of amyloid deposits with laser microdissection.
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| 3. |
- Schael, S, et al.
(författare)
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Precision electroweak measurements on the Z resonance
- 2006
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Ingår i: Physics Reports. - : Elsevier BV. - 0370-1573 .- 1873-6270. ; 427:5-6, s. 257-454
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Forskningsöversikt (refereegranskat)abstract
- We report on the final electroweak measurements performed with data taken at the Z resonance by the experiments operating at the electron-positron colliders SLC and LEP. The data consist of 17 million Z decays accumulated by the ALEPH, DELPHI, L3 and OPAL experiments at LEP, and 600 thousand Z decays by the SLID experiment using a polarised beam at SLC. The measurements include cross-sections, forward-backward asymmetries and polarised asymmetries. The mass and width of the Z boson, m(Z) and Gamma(Z), and its couplings to fermions, for example the p parameter and the effective electroweak mixing angle for leptons, are precisely measured: m(Z) = 91.1875 +/- 0.0021 GeV, Gamma(Z) = 2.4952 +/- 0.0023 GeV, rho(l) = 1.0050 +/- 0.0010, sin(2)theta(eff)(lept) = 0.23153 +/- 0.00016. The number of light neutrino species is determined to be 2.9840 +/- 0.0082, in agreement with the three observed generations of fundamental fermions. The results are compared to the predictions of the Standard Model (SM). At the Z-pole, electroweak radiative corrections beyond the running of the QED and QCD coupling constants are observed with a significance of five standard deviations, and in agreement with the Standard Model. Of the many Z-pole measurements, the forward-backward asymmetry in b-quark production shows the largest difference with respect to its SM expectation, at the level of 2.8 standard deviations. Through radiative corrections evaluated in the framework of the Standard Model, the Z-pole data are also used to predict the mass of the top quark, m(t) = 173(+10)(+13) GeV, and the mass of the W boson, m(W) = 80.363 +/- 0.032 GeV. These indirect constraints are compared to the direct measurements, providing a stringent test of the SM. Using in addition the direct measurements of m(t) and m(W), the mass of the as yet unobserved SM Higgs boson is predicted with a relative uncertainty of about 50% and found to be less than 285 GeV at 95% confidence level. (c) 2006 Elsevier B.V. All rights reserved.
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| 4. |
- Svanberg, Sune, et al.
(författare)
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Applications of terawatt lasers
- 1994
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Ingår i: LASER SPECTROSCOPY - XITH INTERNATIONAL CONFERENCE. - : AIP. - 1551-7616 .- 0094-243X. - 1563962624 ; :290, s. 264-269
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Konferensbidrag (refereegranskat)
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| 5. |
- Arnaud, Julie, et al.
(författare)
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Reduction of lectin valency drastically changes glycolipid dynamics in membranes but not surface avidity.
- 2013
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Ingår i: ACS chemical biology. - : American Chemical Society (ACS). - 1554-8937 .- 1554-8929. ; 8:9, s. 1918-24
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Tidskriftsartikel (refereegranskat)abstract
- Multivalency is proposed to play a role in the strong avidity of lectins for glycosylated cell surfaces and also in their ability to affect membrane dynamics by clustering glycosphingolipids. Lectins with modified valency were designed from the β-propeller fold of Ralstonia solanacearum lectin (RSL) that presents six fucose binding sites. After identification of key amino acids by molecular dynamics calculations, two mutants with reduced valency were produced. Isothermal titration calorimetry confirmed the loss of three high affinity binding sites for both mutants. Crystal structures indicated that residual low affinity binding occurred in W76A but not in R17A. The trivalent R17A mutant presented unchanged avidity toward fucosylated surfaces, when compared to hexavalent RSL. However, R17A is not able anymore to induce formation of membrane invaginations on giant unilamellar vesicules, indicating the crucial role of number of binding sites for clustering of glycolipids. In the human lung epithelial cell line H1299, wt-RSL is internalized within seconds whereas the kinetics of R17A uptake is largely delayed. Neolectins with tailored valency are promising tools to study membrane dynamics.
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| 6. |
- Bally, Marta, 1981, et al.
(författare)
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A virus biosensor with single virus-particle sensitivity based on fluorescent vesicle labels and equilibrium fluctuation analysis
- 2013
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Ingår i: Biointerphases. - : American Vacuum Society. - 1559-4106 .- 1934-8630. ; 8
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Tidskriftsartikel (refereegranskat)abstract
- Biosensors allowing for the rapid and sensitive detection of viral pathogens in environmental or clinical samples are urgently needed to prevent disease outbreaks and spreading. We present a bioanalytical assay for the detection of whole viral particles with single virus sensitivity. Specifically, we focus on the detection of human norovirus, a highly infectious virus causing gastroenteritis. In our assay configuration, virus-like particles are captured onto a supported lipid bilayer containing a virus-specific glycolipid and detected after recognition by a glycolipid-containing fluorescent vesicle. Read-out is performed after illumination of the vesicle labels by total internal reflection fluorescence microscopy. This allows for visualization of individual vesicles and for recording of their binding kinetics under equilibrium conditions (equilibrium fluctuation analysis), as demonstrated previously. In this work we extend the concept and demonstrate that this simple assay setup can be used as a bioanalytical assay for the detection of virus particles at a limit of detection of 16 fM. Furthermore, we demonstrate how the analysis of the single vesicle-virus-like particle interaction dynamics can contribute to increase the accuracy and sensitivity of the assay by discriminating specific from non-specific binding events. This method is suggested to be generally applicable, provided that these events display different interaction kinetics.
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| 7. |
- Bally, Marta, 1981, et al.
(författare)
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Interaction of Single Viruslike Particles with Vesicles Containing Glycosphingolipids
- 2011
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Ingår i: Physical Review Letters. - : American Physical Society. - 0031-9007 .- 1079-7114. ; 107:18
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Tidskriftsartikel (refereegranskat)abstract
- Glycosphingolipids are involved in the first steps of virus-cell interaction, where they mediate specific recognition of the host cell membrane. We have employed total-internal-reflection fluorescence microscopy to explore the interaction kinetics between individual unlabeled noroviruslike particles, which are attached to a glycosphingolipid-containing lipid bilayer, and fluorescent vesicles containing different types and concentrations of glycosphingolipids. Under association equilibrium, the vesicle-binding rate is found to be kinetically limited, yielding information on the corresponding activation energy. The dissociation kinetics are logarithmic over a wide range of time. The latter is explained by the vesicle-size-related distribution of the dissociation activation energy. The biological, pharmaceutical, and diagnostic relevance of the study is briefly discussed.
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| 8. |
- Bally, Marta, 1981, et al.
(författare)
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Interaction of virions with membrane glycolipids
- 2012
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Ingår i: Physical Biology. - : IOP Publishing. - 1478-3967 .- 1478-3975. ; 9:2
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Tidskriftsartikel (refereegranskat)abstract
- Cellular membranes contain various lipids including glycolipids (GLs). The hydrophilic head groups of GLs extend from the membrane into the aqueous environment outside the cell where they act as recognition sites for specific interactions. The first steps of interaction of virions with cells often include contacts with GLs. To clarify the details of such contacts, we have used the total internal reflection fluorescence microscopy to explore the interaction of individual unlabelled virus-like particles (or, more specifically, norovirus protein capsids), which are firmly bound to a lipid bilayer, and fluorescent vesicles containing glycosphingolipids (these lipids form a subclass of GLs). The corresponding binding kinetics were earlier found to be kinetically limited, while the detachment kinetics were logarithmic over a wide range of time. Here, the detachment rate is observed to dramatically decrease with increasing concentration of glycosphingolipids from 1% to 8%. This effect has been analytically explained by using a generic model describing the statistics of bonds in the contact area between a virion and a lipid membrane. Among other factors, the model takes the formation of GL domains into account. Our analysis indicates that in the system under consideration, such domains, if present, have a characteristic size smaller than the contact area between the vesicle and the virus-like particle.
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| 9. |
- Bally, Marta, 1981, et al.
(författare)
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Norovirus GII.4 Virus-like Particles Recognize Galactosylceramides in Domains of Planar Supported Lipid Bilayers.
- 2012
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Ingår i: Angewandte Chemie International Edition. - : Wiley. - 1433-7851 .- 1521-3773. ; 51:48, s. 12020-4
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Tidskriftsartikel (refereegranskat)abstract
- A sticky situation: Domain-dependent recognition of the glycosphingolipid galactosylceramide by norovirus-like particles (see picture; red/yellow) is shown using supported lipid bilayers (purple) as model membranes. Optimal ligand presentation is found to promote strong binding to GalCer. This presentation can be found at the edges of the glycosphingolipid-enriched domains (green) and binding is repressed in the absence of these domains.
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| 10. |
- Bally, Marta, 1981, et al.
(författare)
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Physicochemical tools for studying virus interactions with targeted cell membranes in a molecular and spatiotemporally resolved context
- 2021
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Ingår i: Analytical and Bioanalytical Chemistry. - : Springer Science and Business Media LLC. - 1618-2642 .- 1618-2650. ; 413, s. 7157-7178
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Tidskriftsartikel (refereegranskat)abstract
- The objective of this critical review is to provide an overview of how emerging bioanalytical techniques are expanding our understanding of the complex physicochemical nature of virus interactions with host cell surfaces. Herein, selected model viruses representing both non-enveloped (simian virus 40 and human norovirus) and enveloped (influenza A virus, human herpes simplex virus, and human immunodeficiency virus type 1) viruses are highlighted. The technologies covered utilize a wide range of cell membrane mimics, from supported lipid bilayers (SLBs) containing a single purified host membrane component to SLBs derived from the plasma membrane of a target cell, which can be compared with live-cell experiments to better understand the role of individual interaction pairs in virus attachment and entry. These platforms are used to quantify binding strengths, residence times, diffusion characteristics, and binding kinetics down to the single virus particle and single receptor, and even to provide assessments of multivalent interactions. The technologies covered herein are surface plasmon resonance (SPR), quartz crystal microbalance with dissipation (QCM-D), dynamic force spectroscopy (DFS), total internal reflection fluorescence (TIRF) microscopy combined with equilibrium fluctuation analysis (EFA) and single particle tracking (SPT), and finally confocal microscopy using multi-labeling techniques to visualize entry of individual virus particles in live cells. Considering the growing scientific and societal needs for untangling, and interfering with, the complex mechanisms of virus binding and entry, we hope that this review will stimulate the community to implement these emerging tools and strategies in conjunction with more traditional methods. The gained knowledge will not only contribute to a better understanding of the virus biology, but may also facilitate the design of effective inhibitors to block virus entry.
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| 11. |
- Bengtson, Per, 1971-, et al.
(författare)
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Characterization of EBV-transformed B-cells established from an individual homozygously mutated (G329A) in the FUT7 alpha1,3-fucosyltransferase gene
- 2005
-
Ingår i: Scand J Immunol. - : Wiley. ; 62:3, s. 251-8
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Tidskriftsartikel (refereegranskat)abstract
- The alpha1,3-fucosyltransferase VII (Fuc-TVII) is involved in the biosynthesis of E- and P-selectin ligands such as sialyl Lewis x (SLe(x)) on human leukocytes. Recently, individuals were characterized carrying a missense mutation (G329A; Arg110-Gln) in the FUT7 gene encoding this enzyme. The mutated FUT7 construct produced a Fuc-TVII enzyme with impaired activity compared with the wildtype enzyme. Polymorphonuclear granulocytes from an individual carrying this mutation homozygously also showed a reduced expression of SLe(x). In the present study, we have established Epstein-Barr virus-transformed B-cell lines from this individual (SIGN) and from an individual not carrying the mutation (IWO). The cell lines were confirmed to be of B-cell origin by flow cytometry analysis. IWO cells interacted with E-selectin in an in vitro flow chamber analysis whereas SIGN cell did not. However, when SIGN cell was transiently transfected with wildtype FUT7 cDNA, interaction with E-selectin could be restored. Cell surface expression of the SLe(x)-related epitopes recognized by antibodies CSLEX-1, KM-93 and HECA-452 was elevated on IWO cells compared with that on SIGN cells, consistent with a role of these antigens in E-selectin recognition. These cell lines will be useful in further characterization of E-selectin ligands and encourage further studies on the consequences of the FUT7-G329A mutation in vivo.
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| 12. |
- Bengtson, Per, et al.
(författare)
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EBV-transformed B-cells from an individual homozygously mutated (G329A) in FUT7 do not roll on E-selectin
-
Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- The α1,3 fucosyltransferase VII (Fuc-TVII) is involved in the biosynthesis of E- and P-selectin ligands such as sialyl Lewis x on human leukocytes. Recently, individuals were characterized carrying a missense mutation (G329A; Arg110-Gln) in the FUT7 gene encoding this enzyme. The mutated FUT7 construct was not able to produce an active Fuc-TVII enzyme in transfection studies and polymorphonuclear granulocytes from an individual carrying the mutation homozygously showed severely reduced expression of sialyl Lewis x. In the present study we have established Epstein-Barr virus transformed B-celllines from this individual (SIGN) and from an individual not carrying the mutation (IWO). The cell lines were confirmed to be of B-cell origin by flow cytometry analysis. IWO cells interacted with E-selectin in an in vitro flow chamber analysis whereas SIGN cell did not. However, when SIGN cell were transfected with wild type FUT7 cDNA interaction with E-selectin could be restored. Cell surface expression of the sLex related epitopes recognized by antibodies CSLEX-1, KM-93 and HECA-452 was elevated on IWO cells compared to SIGN cells, consistent with a role of these antigens in E-selectin recognition. Despite high expression of PSGL-1 neither of the cell lines interacted with P-selectin under flow. These cell lines may be useful in the further characterization ofEselectin ligands and the obtained results encourage further studies on the consequences of the FUT7-G329A mutation in vivo.
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| 13. |
- Bengtson, Per, et al.
(författare)
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Polymorphonuclear Leukocytes from Individuals Carrying the G329A Mutation in the α1,3-Fucosyltransferase VII Gene (FUT7) Roll on E- and P-Selectins
- 2002
-
Ingår i: Journal of Immunology. - 0022-1767 .- 1550-6606. ; 169:7, s. 3940-3946
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Tidskriftsartikel (refereegranskat)abstract
- We recently identified several individuals carrying a missense mutation (G329A; Arg110-Gln) in the FUT7 gene encoding fucosyltransferase VII. This enzyme is involved in the biosynthesis of the sialyl Lewis x (Lex) epitope on human leukocytes, which has been identified as an important component of leukocyte ligands for E- and P-selectin. No enzyme activity was measurable in expression studies in COS-7 cells using the mutated FUT7 construct. One of the identified individuals carried this mutation homozygously. Flow cytometry analysis of polymorphonuclear leukocytes (PMN) from this individual showed a nearly complete absence of staining with mAbs directed against sialyl Lex and a diminished staining with an E-selectin IgG chimera. However, staining with P-selectin IgG chimera and Abs directed against P-selectin glycoprotein ligand-1 was not affected by the mutation. PMN from the homozygously mutated individual was further analyzed in an in vitro flow chamber assay. The number of rolling PMN and the rolling velocities on both E- and P-selectin were in the range of PMN from nonmutated individuals. FUT4 and FUT7 mRNA was quantified in PMN isolated from individuals carrying the FUT7 mutation. It was found that PMN from both FUT7 homozygously and heterozygously mutated individuals exhibited an elevated expression of FUT4 mRNA compared with PMN from FUT7 nonmutated individuals. The elevated expression of fucosyltransferase IV was reflected as an increased expression of the Lex and CD65s Ags on PMN from these individuals. The significance of the mutation was supported by transfection of BJAB cells.
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| 14. |
- Bergström, Anders, et al.
(författare)
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Origins and genetic legacy of prehistoric dogs
- 2020
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Ingår i: Science. - : American Association for the Advancement of Science (AAAS). - 0036-8075 .- 1095-9203. ; 370:6516, s. 557-563
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Tidskriftsartikel (refereegranskat)abstract
- Dogs were the first domestic animal, but little is known about their population history and to what extent it was linked to humans. We sequenced 27 ancient dog genomes and found that all dogs share a common ancestry distinct from present-day wolves, with limited gene flow from wolves since domestication but substantial dog-to-wolf gene flow. By 11,000 years ago, at least five major ancestry lineages had diversified, demonstrating a deep genetic history of dogs during the Paleolithic. Coanalysis with human genomes reveals aspects of dog population history that mirror humans, including Levant-related ancestry in Africa and early agricultural Europe. Other aspects differ, including the impacts of steppe pastoralist expansions in West and East Eurasia and a near-complete turnover of Neolithic European dog ancestry.
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| 15. |
- Bernasconi, Valentina, 1989, et al.
(författare)
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A vaccine combination of lipid nanoparticles and a cholera toxin adjuvant derivative greatly improves lung protection against influenza virus infection
- 2021
-
Ingår i: Mucosal Immunology. - : Elsevier BV. - 1933-0219 .- 1935-3456. ; 14:2, s. 523-536
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Tidskriftsartikel (refereegranskat)abstract
- This is a proof-of-principle study demonstrating that the combination of a cholera toxin derived adjuvant, CTA1-DD, and lipid nanoparticles (LNP) can significantly improve the immunogenicity and protective capacity of an intranasal vaccine. We explored the self-adjuvanted universal influenza vaccine candidate, CTA1-3M2e-DD (FPM2e), linked to LNPs. We found that the combined vector greatly enhanced survival against a highly virulent PR8 strain of influenza virus as compared to when mice were immunized with FPM2e alone. The combined vaccine vector enhanced early endosomal processing and peptide presentation in dendritic cells and upregulated co-stimulation. The augmenting effect was CTA1-enzyme dependent. Whereas systemic anti-M2e antibody and CD4(+)T-cell responses were comparable to those of the soluble protein, the local respiratory tract IgA and the specific Th1 and Th17 responses were strongly enhanced. Surprisingly, the lung tissue did not exhibit gross pathology upon recovery from infection and M2e-specific lung resident CD4(+)T cells were threefold higher than in FPM2e-immunized mice. This study conveys optimism as to the protective ability of a combination vaccine based on LNPs and various forms of the CTA1-DD adjuvant platform, in general, and, more specifically, an important way forward to develop a universal vaccine against influenza.
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| 16. |
- Bobbio, Emanuele, et al.
(författare)
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Clinical Diagnosis and Subtyping of Cardiac Amyloidosis by Mass Spectrometry.
- 2020
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Ingår i: The Journal of heart and lung transplantation : the official publication of the International Society for Heart Transplantation. - : Elsevier BV. - 1557-3117. ; 39:4S
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Tidskriftsartikel (refereegranskat)abstract
- Medical treatment for cardiac amyloidosis (CA) is evolving rapidly. Heart transplantation can be a valid option when followed by transplantation of bone marrow or liver, dependent on the type and origin of the amyloid protein. Thus, accurate typing of amyloidosis has implications for treatment, prognosis, and genetic counseling. Although non-invasive diagnostic techniques can type CA, endomyocardial biopsy (EMB) may be needed in the case of equivocal imaging findings or discordant data. We aimed to define the role of mass spectrometry (MS) for diagnosis and subtyping of CA.Nineteen previously diagnosed CA cases, who underwent EMB at Sahlgrenska University Hospital (SU), Gothenburg, between the beginning 1990s and 2016, were selected. MS analysis, modified from was conducted on duplicate samples from myocardial tissue for each case included.1 Clinical features and diagnoses were used as gold standard and compared to the MS findings.Clinical diagnosis and the MS analysis agreed in 14 cases (73.7 %); in 3/19 (15.8 %) diagnosis was unclear or discordant (Fig.1). MS analysis revealed that transthyretin (TTR) amyloidosis was the most abundant amyloid protein in the samples examined (9/19; 47.3 %), whereas the AA subtype only occurred in 1 case (5.2 %). The AL κ type amyloidosis occurred in 3 cases (15.8 %), and AL λ type in six cases (31.6 %). These results strongly correlated with the clinical features in all patients. Clinical diagnosis could not be retrieved from the medical records in 2 cases (10.4 %). Additional 20 patients with clinical CA are presently under study.MS analysis of a small amount of endomyocardial tissue can be used to subtype CA with a high diagnostic validity. The method differentiated between TTR, SAA and Ig light chain amyloidosis. AL κ and AL λ identities correlated to those found in serum and urine electrophoreses. MS can therefore be of use to subtype CA for cases in which clinical findings are inconclusive. 1) Brambilla F et al. Blood. 2012 Feb 23;119(8):1844-7.
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| 17. |
- Breimer, Michael, 1951, et al.
(författare)
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Glycosphingolipid composition of epithelial cells isolated along the villus axis of small intestine of a single human individual
- 2012
-
Ingår i: Glycobiology. - : Oxford University Press (OUP). - 1460-2423 .- 0959-6658. ; 22:12, s. 1721-1730
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Tidskriftsartikel (refereegranskat)abstract
- A 6-cm fresh proximal ileum surgical specimen from a blood group A(1)Le(a-b+) secretor individual was used for stepwise isolation of epithelial cells from villus tip to crypt bottom by gentle washing with ethylenediaminetetraacetic acid-containing buffer. Acid and non-acid sphingolipids were prepared from the epithelial cell fractions and the non-epithelial intestinal residue. Molecular information on the sphingolipid composition was obtained without further isolation of individual species by applying thin-layer chromatography using chemical and biological (monoclonal antibodies, cholera toxin, Escherichia coli) detection reagents, mass spectrometry and proton NMR spectroscopy of derivatized glycolipids. In this way, the structure of major and minor saccharides, ceramide components and their relative amounts were obtained. Epithelial cells and non-epithelial residue were distinctly different in their sphingolipid composition. Sphingomyelin was the major single component in both compartments. Characteristic for epithelial cells was the dominance of monoglycosylceramides, sulphatides and blood group fucolipids (mainly Le(b) hexaglycosylceramides and ALe(b) heptaglycosylceramides). The non-epithelial residue had about five times less glycolipids mainly mono-, di-, tri- and tetra-glycosylceramides and gangliosides, including the GM1 ganglioside. The ceramides were more hydroxylated (1-2 additional hydroxyls) in epithelial cell glycolipids compared with the non-epithelial residue. Combined with a separate detailed study on the glycoproteins of the same epithelial cell preparation, this human intestinal sample is the only epithelial cell preparation where both protein- and lipid-linked saccharides are characterized in detail.
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| 18. |
- Brinkmalm, Gunnar, et al.
(författare)
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An online nano-LC-ESI-FTICR-MS method for comprehensive characterization of endogenous fragments from amyloid β and amyloid precursor protein in human and cat cerebrospinal fluid.
- 2012
-
Ingår i: Journal of mass spectrometry : JMS. - : Wiley. - 1096-9888 .- 1076-5174. ; 47:5, s. 591-603
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Tidskriftsartikel (refereegranskat)abstract
- Amyloid precursor protein (APP) is the precursor protein to amyloid β (Aβ), the main constituent of senile plaques in Alzheimer's disease (AD). Endogenous Aβ peptides reflect the APP processing, and greater knowledge of different APP degradation pathways is important to understand the mechanism underlying AD pathology. When one analyzes longer Aβ peptides by low-energy collision-induced dissociation tandem mass spectrometry (MS/MS), mainly long b-fragments are observed, limiting the possibility to determine variations such as amino acid variants or post-translational modifications (PTMs) within the N-terminal half of the peptide. However, by using electron capture dissociation (ECD), we obtained a more comprehensive sequence coverage for several APP/Aβ peptide species, thus enabling a deeper characterization of possible variants and PTMs. Abnormal APP/Aβ processing has also been described in the lysosomal storage disease Niemann-Pick type C and the major large animal used for studying this disease is cat. By ECD MS/MS, a substitution of Asp7 → Glu in cat Aβ was identified. Further, sialylated core 1 like O-glycans at Tyr10, recently discovered in human Aβ (a previously unknown glycosylation type), were identified also in cat cerebrospinal fluid (CSF). It is therefore likely that this unusual type of glycosylation is common for (at least) species belonging to the magnorder Boreoeutheria. We here describe a detailed characterization of endogenous APP/Aβ peptide species in CSF by using an online top-down MS-based method.
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| 19. |
- Bucardo, Filemon, et al.
(författare)
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Genetic susceptibility to symptomatic norovirus infection in Nicaragua. : norovirus susceptibility in Nicaragua
- 2009
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Ingår i: Journal of medical virology. - : Wiley. - 1096-9071 .- 0146-6615. ; 81:4, s. 728-35
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Tidskriftsartikel (refereegranskat)abstract
- Host genetic resistance to Norovirus (NoV) has been observed in challenge and outbreak studies in populations from Europe, Asia, and USA. In this study, we have investigated if histo-blood group antigens can predict susceptibility to diarrhea caused by NoV in Nicaragua, Central America, and if this can be reflected in antibody-prevalence and titer to NoV among individuals with different histo-blood group antigen phenotypes. Investigation of 28 individuals infected with NoV and 131 population controls revealed 6% of non-secretors in the population and nil non-secretors among patients infected with NoV, suggesting that non-secretors may be protected against NoV disease in Nicaragua. Surprisingly, 25% of the population was Lewis negative (Le(a-b-)). NoV infections with genogroup I (GI) and GII occurred irrespective of Lewis genotype, but none of the Lewis a positive (Le(a + b-)) were infected. The globally dominating GII.4 virus infected individuals of all blood groups except AB (n = 5), while the GI viruses (n = 4) infected only blood type O individuals. Furthermore, O blood types were susceptible to infections with GI.4, GII.4, GII.7, GII.17, and GII.18-Nica viruses, suggesting that secretors with blood type O are susceptible (OR = 1.52) and non-secretors resistant. The overall antibody-prevalence to NoV GII.3 VLP was 62% with the highest prevalence among blood type B carriers (70%) followed by A (68%) and O (62%). All four investigated individuals carrying blood type AB were antibody-negative. Among secretors, 63% were antibody-positive compared to 33% among non-secretors (P = 0.151). This study extends previous knowledge about the histo-blood group antigens role in NoV disease in a population with different genetic background than North American and European.
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| 20. |
- Bucardo, Filemon, et al.
(författare)
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Susceptibility of Children to Sapovirus Infections, Nicaragua, 2005–2006
- 2012
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Ingår i: Emerging Infectious Diseases. - Atlanta, GA, USA : U.S. Department of Health and Human Services * Centers for Disease Control and Prevention. - 1080-6040 .- 1080-6059. ; 18:11, s. 1875-1878
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Tidskriftsartikel (refereegranskat)abstract
- We describe the genetic diversity of sapovirus (SaV) in children in Nicaragua and investigate the role of host genetic factors and susceptibility to SaV infections. Our results indicate that neither ABO blood group, Lewis phenotype, nor secretor status affects susceptibility to SaV infection in Nicaragua.
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| 21. |
- Carlsson, Beatrice, et al.
(författare)
-
Susceptibility to symptomatic sapovirus infection in Denmark is not associated with secretor or Lewis status
-
Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Background. Sapovirus (SaV) infections are increasing globally but there is no information available regarding factors determining susceptibility to SaV infections in the Caucasian population. Methods. Saliva samples were collected from 64 individuals with sapovirus gastroenteritis in Denmark between October 2008 and November 2010. These were genotyped for the FUT2 G428A nonsense mutation (secretor status) and phenotyped for ABO and Lewis histo-blood groups. Results. We found that neither secretor status nor Lewis phenotype, were associated with susceptibility to symptomatic infection with SaV. However, individuals of histo-blood groups B and AB had significantly lower risk to be infected (OR 0.18, p≤0.01 and OR 0.10, p<0.05, respectively). For 39 of the 64 SaV positive samples viral strains were genotyped and 41%, (16/39) belonged to genotype GI.2, 10% was GI.1 (4/39), 2.5% was GI.5 (1/39), 8% was GII.1 (3/39), 5% was GII.4 (2/39), 18% was GIV (7/39) and 15.5% was GV (6/39). Conclusion. This is the first report investigating the role of host genetic factors in SaV susceptibility in the Caucasian population. We found a reduced risk of infection in individuals with blood group B (and AB), but no association to the FUT2 G428A nonsense mutation determining secretor status nor to the Lewis status.
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| 22. |
- Carlsson, Beatrice, et al.
(författare)
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The G428A nonsense mutation in FUT2 provides strong but not absolute protection against symptomatic GII.4 Norovirus infection. : Novel GII.4 disease pattern
- 2009
-
Ingår i: PloS one. - : Public Library of Science (PLoS). - 1932-6203. ; 4:5
-
Tidskriftsartikel (refereegranskat)abstract
- In November 2004, 116 individuals in an elderly nursing home in El Grao de Castellón, Spain were symptomatically infected with genogroup II.4 (GII.4) norovirus. The global attack rate was 54.2%. Genotyping of 34 symptomatic individuals regarding the FUT2 gene revealed that one patient was, surprisingly, a non-secretor, hence indicating secretor-independent infection. Lewis genotyping revealed that Lewis-positive and negative individuals were susceptible to symptomatic norovirus infection indicating that Lewis status did not predict susceptibility. Saliva based ELISA assays were used to determine binding of the outbreak virus to saliva samples. Saliva from a secretor-negative individual bound the authentic outbreak GII.4 Valencia/2004/Es virus, but did not in contrast to secretor-positive saliva bind VLP of other strains including the GII.4 Dijon strain. Amino acid comparison of antigenic A and B sites located on the external loops of the P2 domain revealed distinct differences between the Valencia/2004/Es and Dijon strains. All three aa in each antigenic site as well as 10/11 recently identified evolutionary hot spots, were unique in the Valencia/2004/Es strain compared to the Dijon strain. To the best of our knowledge, this is the first example of symptomatic GII.4 norovirus infection of a Le(a+b-) individual homozygous for the G428A nonsense mutation in FUT2. Taken together, our study provides new insights into the host genetic susceptibility to norovirus infections and evolution of the globally dominating GII.4 viruses.
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| 23. |
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| 24. |
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| 25. |
- De Leoz, M. L. A., et al.
(författare)
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NIST Interlaboratory Study on Glycosylation Analysis of Monoclonal Antibodies: Comparison of Results from Diverse Analytical Methods
- 2020
-
Ingår i: Molecular & Cellular Proteomics. - 1535-9476. ; 19:1, s. 11-30
-
Tidskriftsartikel (refereegranskat)abstract
- A broad-based interlaboratory study of glycosylation profiles of a reference and modified IgG antibody involving 103 reports from 76 laboratories. Glycosylation is a topic of intense current interest in the development of biopharmaceuticals because it is related to drug safety and efficacy. This work describes results of an interlaboratory study on the glycosylation of the Primary Sample (PS) of NISTmAb, a monoclonal antibody reference material. Seventy-six laboratories from industry, university, research, government, and hospital sectors in Europe, North America, Asia, and Australia submitted a total of 103 reports on glycan distributions. The principal objective of this study was to report and compare results for the full range of analytical methods presently used in the glycosylation analysis of mAbs. Therefore, participation was unrestricted, with laboratories choosing their own measurement techniques. Protein glycosylation was determined in various ways, including at the level of intact mAb, protein fragments, glycopeptides, or released glycans, using a wide variety of methods for derivatization, separation, identification, and quantification. Consequently, the diversity of results was enormous, with the number of glycan compositions identified by each laboratory ranging from 4 to 48. In total, one hundred sixteen glycan compositions were reported, of which 57 compositions could be assigned consensus abundance values. These consensus medians provide community-derived values for NISTmAb PS. Agreement with the consensus medians did not depend on the specific method or laboratory type. The study provides a view of the current state-of-the-art for biologic glycosylation measurement and suggests a clear need for harmonization of glycosylation analysis methods.
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| 26. |
- Delbaere, S., et al.
(författare)
-
b3galt6 Knock-Out Zebrafish Recapitulate beta 3GalT6-Deficiency Disorders in Human and Reveal a Trisaccharide Proteoglycan Linkage Region
- 2020
-
Ingår i: Frontiers in Cell and Developmental Biology. - : Frontiers Media SA. - 2296-634X. ; 8
-
Tidskriftsartikel (refereegranskat)abstract
- Proteoglycans are structurally and functionally diverse biomacromolecules found abundantly on cell membranes and in the extracellular matrix. They consist of a core protein linked to glycosaminoglycan chains via a tetrasaccharide linkage region. Here, we show that CRISPR/Cas9-mediated b3galt6 knock-out zebrafish, lacking galactosyltransferase II, which adds the third sugar in the linkage region, largely recapitulate the phenotypic abnormalities seen in human beta 3GalT6-deficiency disorders. These comprise craniofacial dysmorphism, generalized skeletal dysplasia, skin involvement and indications for muscle hypotonia. In-depth TEM analysis revealed disturbed collagen fibril organization as the most consistent ultrastructural characteristic throughout different affected tissues. Strikingly, despite a strong reduction in glycosaminoglycan content, as demonstrated by anion-exchange HPLC, subsequent LC-MS/MS analysis revealed a small amount of proteoglycans containing a unique linkage region consisting of only three sugars. This implies that formation of glycosaminoglycans with an immature linkage region is possible in a pathogenic context. Our study, therefore unveils a novel rescue mechanism for proteoglycan production in the absence of galactosyltransferase II, hereby opening new avenues for therapeutic intervention.
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| 27. |
- El Masri, R., et al.
(författare)
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Extracellular endosulfatase Sulf-2 harbors a chondroitin/dermatan sulfate chain that modulates its enzyme activity
- 2022
-
Ingår i: Cell Reports. - : Elsevier BV. - 2211-1247. ; 38:11
-
Tidskriftsartikel (refereegranskat)abstract
- Sulfs represent a class of unconventional sulfatases which provide an original post-synthetic regulatory mechanism for heparan sulfate polysaccharides and are involved in multiple physiopathological processes, including cancer. However, Sulfs remain poorly characterized enzymes, with major discrepancies regarding their in vivo functions. Here we show that human SuIf-2 (HSulf-2) harbors a chondroitin/dermatan sulfate glycosaminoglycan (GAG) chain, attached to the enzyme substrate-binding domain. We demonstrate that this GAG chain affects enzyme/substrate recognition and tunes HSulf-2 activity in vitro and in vivo. In addition, we show that mammalian hyaluronidase acts as a promoter of HSulf-2 activity by digesting its GAG chain. In conclusion, our results highlight HSulf-2 as a proteoglycan-related enzyme and its GAG chain as a critical non-catalytic modulator of the enzyme activity. These findings contribute to clarifying the conflicting data on the activities of the Sulfs.
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| 28. |
- Elmgren, A, et al.
(författare)
-
DNA sequencing and screening for point mutations in the human Lewis (FUT3) gene enables molecular genotyping of the human Lewis blood group system.
- 1996
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Ingår i: Vox sanguinis. - 0042-9007. ; 70:2, s. 97-103
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Tidskriftsartikel (refereegranskat)abstract
- The human Lewis gene encodes an alpha(1,3/1,4)-fucosyltransferase responsible for synthesis of the Le(a) and a Le(b) antigens. To define the molecular background for non-functional Lewis genes we have sequenced PCR-amplified DNA fragments from two Le(a-b-) individuals. One was homozygously mutated at nucleotides 202(T --> C) and 314 (C --> T), altering Trp68 to Arg and Thr105 to Met, and the other was homozygously mutated at nucleotides 59 (T --> G) and 1067 (T --> A), altering Leu20 to Arg and Ile356 to Lys. Using PCR we screened for these and additionally one other mutation at nucleotide 508 (G --> A) among 40 Caucasians. Of 15 Le(a-b-) individuals, 7 typed as le59/1067le202/314, 4 as le202/314le202/314 and 1 as le59/1067le59/1067. Of 21 Le(a-b+) and 4 Le(a+b-), 17 typed as LeLe and 7 as Lele202/314. A pedigree study of 8 Lewis-positive individuals showed that the mutations at nucleotides 202 and 314 were located on the same allele.
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| 29. |
- Eriksson, Jonas
(författare)
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Genetic and Genomic Studies in Chicken : Assigning Function to Vertebrate Genes
- 2012
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- A major challenge in the post-genomic era is to understand how genome sequence variants (genotype) give rise to the enormous diversity observed in terms of morphology, physiology and behavior (phenotype) among living organisms. Domestic animals—with their tremendous phenotypic variation—are excellent model organisms for determining the relationships between genotype and phenotype. In this thesis, I describe the utilization of the chicken, in combination with modern genetic and genomic approaches, in developing our understanding of the genetic mechanisms underlying phenotypic variation. These studies provide novel information on the genetics behind variation in carotenoid- and melanin-based pigmentation—observed in many organisms—and also cast light on the genetic basis of chicken domestication. In paper I, we report that the yellow skin phenotype—observed in most commercial chickens—is caused by one or several tissue-specific mutations altering the expression of beta-carotene oxygenase 2 (BCO2 or BCDO2) in skin. In addition, we present the first conclusive evidence of a hybrid origin of the domestic chicken, since the allele causing yellow skin most likely originates from the grey jungle fowl (Gallus sonneratii) and not from the previously described sole ancestor, the red jungle fowl (Gallus gallus). In paper II, we detect a number of loci that were likely important during the domestication process of chicken and the later specialization into meat (broiler) and egg (layer) producing lines. One of the major findings was that worldwide, almost all domestic chickens carry a missense mutation in TSHR (thyroid stimulating hormone receptor) in a position that is completely conserved amongst vertebrates. We speculate that this “domestication-mutation” has played an important role in the transformation of the wild red jungle fowl ancestor into the modern domestic chicken. In paper III, we demonstrate that the dilution of red (pheomelanin) pigmentation—observed in the plumage of the Inhibitor of Gold chicken—is caused by a frame-shift mutation in the catechol-O-methyltransferase domain containing 1 (COMTD1) gene. The production and regulation of pheomelanin is poorly understood and this discovery advances our current knowledge of this pathway.
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| 30. |
- Fernandez-Mateos, P, et al.
(författare)
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Point mutations and deletion responsible for the Bombay H null and the Reunion H weak blood groups.
- 1998
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Ingår i: Vox sanguinis. - 0042-9007. ; 75:1, s. 37-46
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE: Definition of the molecular basis of the Reunion and the Bombay red cell and salivary H-deficient phenotypes. METHODS: Sequence and expression of FUT1 and FUT2 genes from H-deficient individuals. Family segregation analysis of the mutations responsible for the fucosyltransferase defects of H, secretor and Lewis systems. RESULTS: The Indian red cell H null Bombay phenotype depends on a new mutation of the FUT1 gene. T725-->G changing Leu242-->Arg. Their salivary nonsecretor phenotype is secondary to a complete deletion of the FUT2 gene. The red cell H weak Reunion phenotype depends on another new mutation of FUT1, C349-->T which induces a change of His117-->Tyr. Their salivary nonsecretor phenotype is due to the known Caucasian inactivating mutation G428-->A. CONCLUSION: Single prevalent FUT1 and FUT2 point mutations and a deletion are responsible for the Indian Bombay H null and the Reunion H weak phenotypes found on Reunion island. This is in contrast with other H-deficient phenotypes where sporadic nonprevalent inactivating mutations are the rule.
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| 31. |
- Grahn, Ammi, 1961, et al.
(författare)
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Determination of Lewis FUT3 gene mutations by PCR using sequence-specific primers enables efficient genotyping of clinical samples.
- 2001
-
Ingår i: Human mutation. - : Hindawi Limited. - 1098-1004 .- 1059-7794. ; 18:4, s. 358-9
-
Tidskriftsartikel (refereegranskat)abstract
- We have developed a polymerase chain reaction method using sequence-specific primers (PCR-SSP) for rapid and correct genotyping of the common Lewis (FUT3) gene mutations 59T>G, 202T>C, 314C>T, 508G>A, and 1067T>A. The PCR-SSP method was validated on 20 healthy blood donors and 16 non-insulin-dependent diabetic patients. All individuals were in parallel genotyped by our established polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. The FUT3 genotypes, determined with the PCR-SSP method, were in complete accordance with the results of the PCR-RFLP reference method. The PCR-SSP method could also be adapted to assign the presence of a specific mutation to the respective FUT3 alleles. We found the method to be reliable, rapid and cheap with no requirements for restriction enzyme processing.
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| 32. |
- Grahn, Ammi, 1961, et al.
(författare)
-
Glycobiology and cancer
- 2009
-
Ingår i: Encyclopedia of Cancer. - Berlin : Springer Verlag. - 9783540476481
-
Bokkapitel (övrigt vetenskapligt/konstnärligt)
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| 33. |
- Grahn, Ammi, 1961, et al.
(författare)
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Identification of seven new alpha2,3-sialyltransferase III, ST3Gal III, transcripts from human foetal brain
- 2004
-
Ingår i: Glycoconj J. ; 20:7-8, s. 493-500
-
Tidskriftsartikel (refereegranskat)abstract
- We have recently cloned and sequenced 19 human ST3Gal III gene isotranscripts from peripheral blood leukocytes and identified very complex patterns of isotranscripts of this gene in neuronal tissues. We have now cloned and sequenced additionally seven new isotranscripts from foetal brain. These novel isotranscripts showed losses of complete exons along the whole length of the coding sequence. None of the new isotranscripts coded for proteins with the two (L- and S-) sialylmotifs intact. One of the isotranscripts belonged to the isoform ST3Gal III -B, five to the ST3Gal III -C isoform and one to ST3Gal III -D isoform of isotranscripts, which lacks exon 3, exons 3 and 4 and exon 4 respectively. Two of the C series isotranscripts, ST3Gal III C4 and C11 had both lost exons 12 and 13 containing the S-motif but had otherwise the L- and the VS-motifs intact. Three isotranscripts, ST3Gal III C5, C12 and D5, were similar in the 3'-end coding for an identical amino acid sequence unrelated to the original enzyme. Isotranscripts ST3Gal III C9 and B10 were distinctly different from all other forms identified so far. The splice variants reported here are unlikely to express enzymatic activities but may other biological functions.
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| 34. |
- Groenen, M. A., et al.
(författare)
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Analyses of pig genomes provide insight into porcine demography and evolution
- 2012
-
Ingår i: Nature. - : Springer Science and Business Media LLC. - 0028-0836 .- 1476-4687. ; 491:7424, s. 393-398
-
Tidskriftsartikel (refereegranskat)abstract
- For 10,000 years pigs and humans have shared a close and complex relationship. From domestication to modern breeding practices, humans have shaped the genomes of domestic pigs. Here we present the assembly and analysis of the genome sequence of a female domestic Duroc pig (Sus scrofa) and a comparison with the genomes of wild and domestic pigs from Europe and Asia. Wild pigs emerged in South East Asia and subsequently spread across Eurasia. Our results reveal a deep phylogenetic split between European and Asian wild boars approximately 1 million years ago, and a selective sweep analysis indicates selection on genes involved in RNA processing and regulation. Genes associated with immune response and olfaction exhibit fast evolution. Pigs have the largest repertoire of functional olfactory receptor genes, reflecting the importance of smell in this scavenging animal. The pig genome sequence provides an important resource for further improvements of this important livestock species, and our identification of many putative disease-causing variants extends the potential of the pig as a biomedical model.
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| 35. |
- Haga, K., et al.
(författare)
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Genetic Manipulation of Human Intestinal Enteroids Demonstrates the Necessity of a Functional Fucosyltransferase 2 Gene for Secretor-Dependent Human Norovirus Infection
- 2020
-
Ingår i: mBio. - 2150-7511. ; 11:2
-
Tidskriftsartikel (refereegranskat)abstract
- Human noroviruses (HuNoVs) are the leading cause of nonbacterial gastroenteritis worldwide. Histo-blood group antigen (HBGA) expression is an important susceptibility factor for HuNoV infection based on controlled human infection models and epidemiologic studies that show an association of secretor status with infection caused by several genotypes. The fucosyltransferase 2 gene (FUT2) affects HBGA expression in intestinal epithelial cells; secretors express a functional FUT2 enzyme, while nonsecretors lack this enzyme and are highly resistant to infection and gastroenteritis caused by many HuNoV strains. These epidemiologic associations are confirmed by infections in stem cell-derived human intestinal enteroid (HIE) cultures. GII.4 HuNoV does not replicate in HIE cultures derived from nonsecretor individuals, while HIEs from secretors are permissive to infection. However, whether FUT2 expression alone is critical for infection remains unproven, since routinely used secretor-positive transformed cell lines are resistant to HuNoV replication. To evaluate the role of FUT2 in HuNoV replication, we used CRISPR or overexpression to genetically manipulate FUT2 gene function to produce isogenic HIE lines with or without FUT2 expression. We show that FUT2 expression alone affects both HuNoV binding to the HIE cell surface and susceptibility to HuNoV infection. These findings indicate that initial binding to a molecule(s) glycosylated by FUT2 is critical for HuNoV infection and that the HuNoV receptor is present in nonsecretor HIEs. In addition to HuNoV studies, these isogenic HIE lines will be useful tools to study other enteric microbes where infection and/or disease outcome is associated with secretor status. IMPORTANCE Several studies have demonstrated that secretor status is associated with susceptibility to human norovirus (HuNoV) infection; however, previous reports found that FUT2 expression is not sufficient to allow infection with HuNoV in a variety of continuous laboratory cell lines. Which cellular factor(s) regulates susceptibility to HuNoV infection remains unknown. We used genetic manipulation of HIE cultures to show that secretor status determined by FUT2 gene expression is necessary and sufficient to support HuNoV replication based on analyses of isogenic lines that lack or express FUT2. Fucosylation of HBGAs is critical for initial binding and for modification of another putative receptor(s) in HIEs needed for virus uptake or uncoating and necessary for successful infection by GII and several GII HuNoV strains.
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| 36. |
- Halim, Adnan, et al.
(författare)
-
Assignment of Saccharide Identities through Analysis of Oxonium Ion Fragmentation Profiles in LC-MS/MS of Glycopeptides.
- 2014
-
Ingår i: Journal of proteome research. - : American Chemical Society (ACS). - 1535-3907 .- 1535-3893. ; 13:12, s. 6024-32
-
Tidskriftsartikel (refereegranskat)abstract
- Protein glycosylation plays critical roles in the regulation of diverse biological processes, and determination of glycan structure-function relationships is important to better understand these events. However, characterization of glycan and glycopeptide structural isomers remains challenging and often relies on biosynthetic pathways being conserved. In glycoproteomic analysis with liquid chromatography-tandem mass spectrometry (LC-MS/MS) using collision-induced dissociation (CID), saccharide oxonium ions containing N-acetylhexosamine (HexNAc) residues are prominent. Through analysis of beam-type CID spectra and ion trap CID spectra of synthetic and natively derived N- and O-glycopeptides, we found that the fragmentation patterns of oxonium ions characteristically differ between glycopeptides terminally substituted with GalNAcα1-O-, GlcNAcβ1-O-, Galβ3GalNAcα1-O-, Galβ4GlcNAcβ-O-, and Galβ3GlcNAcβ-O- structures. The difference in the oxonium ion fragmentation profiles of such glycopeptides may thus be used to distinguish among these glycan structures and could be of importance in LC-MS/MS-based glycoproteomic studies.
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| 37. |
- Halim, Adnan, et al.
(författare)
-
Human urinary glycoproteomics; attachment site specific analysis of N- and O-linked glycosylations by CID and ECD.
- 2012
-
Ingår i: Molecular & cellular proteomics : MCP. - 1535-9484. ; 11:4
-
Tidskriftsartikel (refereegranskat)abstract
- Urine is a complex mixture of proteins and waste products and a challenging biological fluid for biomarker discovery. Previous proteomic studies have identified more than 2800 urinary proteins but analyses aimed at unraveling glycan structures and glycosylation sites of urinary glycoproteins are lacking. Glycoproteomic characterization remains difficult because of the complexity of glycan structures found mainly on asparagine (N-linked) or serine/threonine (O-linked) residues. We have developed a glycoproteomic approach that combines efficient purification of urinary glycoproteins/glycopeptides with complementary MS-fragmentation techniques for glycopeptide analysis. Starting from clinical sample size, we eliminated interfering urinary compounds by dialysis and concentrated the purified urinary proteins by lyophilization. Sialylated urinary glycoproteins were conjugated to a solid support by hydrazide chemistry and trypsin digested. Desialylated glycopeptides, released through mild acid hydrolysis, were characterized by tandem MS experiments utilizing collision induced dissociation (CID) and electron capture dissociation fragmentation techniques. In CID-MS(2), Hex(5)HexNAc(4)-N-Asn and HexHexNAc-O-Ser/Thr were typically observed, in agreement with known N-linked biantennary complex-type and O-linked core 1-like structures, respectively. Additional glycoforms for specific N- and O-linked glycopeptides were also identified, e.g. tetra-antennary N-glycans and fucosylated core 2-like O-glycans. Subsequent CID-MS(3), of selected fragment-ions from the CID-MS(2) analysis, generated peptide specific b- and y-ions that were used for peptide identification. In total, 58 N- and 63 O-linked glycopeptides from 53 glycoproteins were characterized with respect to glycan- and peptide sequences. The combination of CID and electron capture dissociation techniques allowed for the exact identification of Ser/Thr attachment site(s) for 40 of 57 putative O-glycosylation sites. We defined 29 O-glycosylation sites which have, to our knowledge, not been previously reported. This is the first study of human urinary glycoproteins where "intact" glycopeptides were studied, i.e. the presence of glycans and their attachment sites were proven without doubt.
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| 38. |
- Halim, Adnan, et al.
(författare)
-
LC-MS/MS characterization of O-glycosylation sites and glycan structures of human cerebrospinal fluid glycoproteins.
- 2013
-
Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 12:2, s. 573-584
-
Tidskriftsartikel (refereegranskat)abstract
- The GalNAc O-glycosylation on Ser/Thr residues of extracellular proteins has not been well characterized from a proteomics perspective. We previously reported a sialic acid capture-and-release protocol to enrich tryptic N- and O-glycopeptides from human cerebrospinal fluid glycoproteins using nanoLC-ESI-MS/MS with collision-induced dissociation (CID) for glycopeptide characterization. Here, we have introduced peptide N-glycosidase F (PNGase F) pre-treatment of CSF samples to remove the N-glycans facilitating the selective characterization of O-glycopeptides and enabling the use of an automated CID-MS2/MS3 search protocol for glycopeptide identification. We used electron capture and transfer dissociation (ECD/ETD) to pinpoint the glycosylation site(s) of the glycopeptides, identified as predominantly core 1 like HexHexNAc-O- glycans attached to 1-4 Ser/Thr residues. We characterized 108 O-glycosylations and found Pro residues preferentially in the n-1, n+1 and/or n+3 positions in relation to the Ser/Thr attachment site (n). The characterization of glycans and glycosylation sites in glycoproteins from human clinical samples provides a basis for future studies addressing the biological and diagnostic importance of specific protein glycosylations in relation to human disease.
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| 39. |
- Halim, Adnan, et al.
(författare)
-
Site-specific characterization of threonine, serine, and tyrosine glycosylations of amyloid precursor protein/amyloid {beta}-peptides in human cerebrospinal fluid.
- 2011
-
Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 1091-6490. ; 108:29, s. 11848-53
-
Tidskriftsartikel (refereegranskat)abstract
- The proteolytic processing of human amyloid precursor protein (APP) into shorter aggregating amyloid β (Aβ)-peptides, e.g., Aβ1-42, is considered a critical step in the pathogenesis of Alzheimer's disease (AD). Although APP is a well-known membrane glycoprotein carrying both N- and O-glycans, nothing is known about the occurrence of released APP/Aβ glycopeptides in cerebrospinal fluid (CSF). We used the 6E10 antibody and immunopurified Aβ peptides and glycopeptides from CSF samples and then liquid chromatography-tandem mass spectrometry for structural analysis using collision-induced dissociation and electron capture dissociation. In addition to 33 unglycosylated APP/Aβ peptides, we identified 37 APP/Aβ glycopeptides with sialylated core 1 like O-glycans attached to Thr(-39, -21, -20, and -13), in a series of APP/AβX-15 glycopeptides, where X was -63, -57, -52, and -45, in relation to Asp1 of the Aβ sequence. Unexpectedly, we also identified a series of 27 glycopeptides, the Aβ1-X series, where X was 20 (DAEFRHDSGYEVHHQKLVFF), 19, 18, 17, 16, and 15, which were all uniquely glycosylated on Tyr10. The Tyr10 linked O-glycans were (Neu5Ac)(1-2)Hex(Neu5Ac)HexNAc-O- structures with the disialylated terminals occasionally O-acetylated or lactonized, indicating a terminal Neu5Acα2,8Neu5Ac linkage. We could not detect any glycosylation of the Aβ1-38/40/42 isoforms. We observed an increase of up to 2.5 times of Tyr10 glycosylated Aβ peptides in CSF in six AD patients compared to seven non-AD patients. APP/Aβ sialylated O-glycans, including that of a Tyr residue, the first in a mammalian protein, may modulate APP processing, inhibiting the amyloidogenic pathway associated with AD.
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| 40. |
- Hedberg, Carola, 1969, et al.
(författare)
-
Hereditary myopathy with early respiratory failure is associated with misfolding of the titin fibronectin III 119 subdomain
- 2014
-
Ingår i: Neuromuscular Disorders. - : Elsevier BV. - 0960-8966. ; 24:5, s. 373-379
-
Tidskriftsartikel (refereegranskat)abstract
- Hereditary myopathy with early respiratory failure is a rare disease with muscle weakness and respiratory failure as early symptoms. Muscle pathology is characterized by the presence of multiple cytoplasmic bodies and other protein aggregates in muscle fibers. The disease is associated with mutations in the titin gene (TTN). All patients harbor mutations located in exon 343 in the TTN gene that codes for the fibronectin III domain 119 (FN3 119) in the 10th motif of the 11-element motif A-band super-repeat. We investigated how such disease-causing mutations affect the biochemical behavior of this titin domain. All five disease-causing amino acid changes analyzed by us (p.P30068R, p.C30071R, p.W30088R, p.W30088C and p.P30091L) resulted in impaired FN3 119 domain solubility. In contrast, amino acid changes associated with common SNPs (p.V30076I, p.R30107C and p.S30125F) did not have this effect. In silica analyses further support the notion that disease-causing mutations impair proper folding of the FN3 119 domain. The results suggest that hereditary myopathy with early respiratory failure is caused by defective protein folding. (C) 2014 Elsevier B.V. All rights reserved.
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| 41. |
- Hellström, Göran, et al.
(författare)
-
Seasonal thermal energy storage - the HYDROCK concept
- 2001
-
Ingår i: Bulletine of Engineering Geology and the Environment. ; 60, s. 145-156
-
Tidskriftsartikel (refereegranskat)abstract
- A method for seasonal storage of heat or cold in the bedrock (the HYDROCK concept) is presented and its thermal performance discussed.
|
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| 42. |
- Hellström, Göran, et al.
(författare)
-
Seasonal thermal energy storage - The HYDROCK concept
- 2001
-
Ingår i: Bulletin of Engineering Geology and the Environment. - : Springer Science and Business Media LLC. - 1435-9529 .- 1435-9537. ; 60:2, s. 145-156
-
Tidskriftsartikel (refereegranskat)abstract
- A method for seasonal storage of heat or cold in the bedrock (the HYDROCK concept) is presented and its thermal performance discussed. It involves the use of a fractured bedrock at shallow depths (ca. 50-250 m), where existing fractures are stimulated or new fractures artificially created and used as flow-paths for a heat/cold carrier (usually water). The fracture surfaces are used as heat exchangers and the bedrock is loaded and unloaded to suit the energy needs. Propants are injected into the fractures to keep them open and reduce the energy needed for pumping water through the system. Field tests confirm that stacked parallel fractures can be produced by hydraulic fracturing. The thermal performance of the store is modelled and compared with a ducted ground heat store. It is shown that the HYDROCK store may yield 10-20% more energy during extraction than a ducted ground heat store for similar amounts of injected energy. This indicates that the HYDROCK concept is competitive as a seasonal energy store and may be of particular importance as an alternative energy source where existing methods cannot be economically justified.
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| 43. |
- Henriksson, Sara, 1981-
(författare)
-
Helicobacter pylori : multitalented adaptation of binding properties
- 2012
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Helicobacter pylori infects and persistently colonizes the stomach, which results in gastritis and in some individuals peptic ulcer disease or gastric cancer. Adherence of H. pylori to the epithelium is an important factor for development of disease. Attachment is mediated by the adhesins BabA and SabA that binds the ABO/Leb blood group antigens and sialylated glycoconjugates respectively. High-affinity attachment could be anticipated to be of disadvantage for H. pylori because epithelial cells have a fast turnover rate and the dislocated and shed epithelial cells would carry attached bacteria to the acidic gastric juice in the lumen. However, here we describe that H. pylori manage to adapt to this innate clearance mechanism by unique acid regulatory binding properties of its adhesins. We propose that pH regulated binding properties enable bacteria to detachment from host cells for chemotactic guided motility and successful return to the more neutral epithelium for a fresh restart of the infectious cycle. By comparison of BabA from different stomach loci we identified amino acid key position for acid regulated binding activity.Previous studies found lower prevalence of Leb-binding among H. pylori isolates from southern Europe compared to Sweden. Here we tested if the reduced prevalence of Leb-binding could be explained by a novel binding mode; in among Spanish strains, we identified S812 that demonstrates preference for multivalent binding to ABO antigens in glycolipids; we found that 812 BabA had drifted in its preferred binding epitope away from the consensus a1,2fucosylation and towards the blood group A and B derivatives. Such epitope drift might in particular optimize binding to ABO antigens in densely packed lipid rafts.In parallel, we studied the influence of BabA for disease progression by an inventory of gastric biopsies. BabA correlated both with the oncoprotein CagA, the VacAs1 toxin and, in addition, to severe disease progression. We further correlate BabA expression with positive secretor phenotype and stronger adhesion of H. pylori in vitro.For functional adherence studies in vitro, we constructed a recombinant Leb-expressing cell lineage that supports BabA mediated H. pylori attachment.
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| 44. |
- Hesse, Camilla, et al.
(författare)
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The N-terminal domain of α-dystroglycan is released as a 38kDa protein and is increased in cerebrospinal fluid in patients with Lyme neuroborreliosis.
- 2011
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Ingår i: Biochemical and biophysical research communications. - : Elsevier BV. - 1090-2104 .- 0006-291X. ; 412:3, s. 494-499
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Tidskriftsartikel (refereegranskat)abstract
- α-Dystroglycan is an extracellular adhesion protein that is known to interact with different ligands. The interaction is thought to stabilize the integrity of the plasma membrane. The N-terminal part of α-dystroglycan may be proteolytically processed to generate a small 38kDa protein (α-DG-N). The physiological significance of α-DG-N is unclear but has been suggested to be involved in nerve regeneration and myelination and to function as a potential biomarker for neurodegenerative and neuromuscular diseases. In this report we show that α-DG-N is released into different body fluids, such as lachrimal fluid, cerebrospinal fluid (CSF), urine and plasma. To investigate the significance of α-DG-N in CSF we examined the levels of α-DG-N and known neurodegenerative markers in CSF from patients diagnosed with Lyme neuroborreliosis (LNB) and healthy controls. In untreated acute phase LNB patients, 67% showed a significant increase of CSF α-DG-N compared to healthy controls. After treatment with antibiotics the CSF α-DG-N levels were normalized in the LNB patients.
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| 45. |
- Kawahara, R., et al.
(författare)
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Community evaluation of glycoproteomics informatics solutions reveals high-performance search strategies for serum glycopeptide analysis
- 2021
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Ingår i: Nature Methods. - : Springer Science and Business Media LLC. - 1548-7091 .- 1548-7105. ; 18, s. 1304-1316
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Tidskriftsartikel (refereegranskat)abstract
- Glycoproteomics is a powerful yet analytically challenging research tool. Software packages aiding the interpretation of complex glycopeptide tandem mass spectra have appeared, but their relative performance remains untested. Conducted through the HUPO Human Glycoproteomics Initiative, this community study, comprising both developers and users of glycoproteomics software, evaluates solutions for system-wide glycopeptide analysis. The same mass spectrometrybased glycoproteomics datasets from human serum were shared with participants and the relative team performance for N- and O-glycopeptide data analysis was comprehensively established by orthogonal performance tests. Although the results were variable, several high-performance glycoproteomics informatics strategies were identified. Deep analysis of the data revealed key performance-associated search parameters and led to recommendations for improved 'high-coverage' and 'high-accuracy' glycoproteomics search solutions. This study concludes that diverse software packages for comprehensive glycopeptide data analysis exist, points to several high-performance search strategies and specifies key variables that will guide future software developments and assist informatics decision-making in glycoproteomics. This analysis presents the results of a community-based evaluation of existing software for large-scale glycopeptide data analysis.
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| 46. |
- Kindberg, Elin, 1981-, et al.
(författare)
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Host genetic resistance to symptomatic norovirus (GGII.4) infections in Denmark
- 2007
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Ingår i: Journal of Clinical Microbiology. - 0095-1137 .- 1098-660X. ; 45:8, s. 2720-2
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Tidskriftsartikel (refereegranskat)abstract
- A total of 61 individuals involved in five norovirus outbreaks in Denmark were genotyped at nucleotides 428 and 571 of the FUT2 gene, determining secretor status, i.e., the presence of ABH antigens in secretions and on mucosa. A strong correlation (P = 0.003) was found between the secretor phenotype and symptomatic disease, extending previous knowledge and confirming that nonsense mutations in the FUT2 gene provide protection against symptomatic norovirus (GGII.4) infections. Copyright © 2007, American Society for Microbiology. All Rights Reserved.
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| 47. |
- Koppisetty, Ashok Krishna Chaitanya, 1982, et al.
(författare)
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Computational studies on the interaction of ABO-active saccharides with the norovirus VA387 capsid protein can explain experimental binding data
- 2010
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Ingår i: Journal of Computer-Aided Molecular Design. - : Springer Science and Business Media LLC. - 0920-654X .- 1573-4951. ; 24:5, s. 423-431
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Tidskriftsartikel (refereegranskat)abstract
- Norovirus strains are known to cause recurring epidemics of winter vomiting disease. The crystal structure of the capsid protein of VA387, a representative of the clinically important GII.4 genocluster, was recently solved in complex with histo-blood group A- and B-trisaccharides. However, the VA387 strain is known to bind also to other natural carbohydrates for which detailed structural information of the complexes is not available. In this study we have computationally explored the fit of the VA387 with a set of naturally occurring carbohydrate ligands containing a terminal alpha 1,2-linked fucose. MD simulations both with explicit and implicit solvent models indicate that type 1 and 3 extensions of the ABO-determinant including ALe(b) and BLe(b) pentasaccharides can be well accommodated in the site. Scoring with Glide XP indicates that the downstream extensions of the ABO-determinants give an increase in binding strength, although the alpha 1,2-linked fucose is the single strongest interacting residue. An error was discovered in the geometry of the GalNAc-Gal moiety of the published crystal structure of the A-trisaccharide/VA387 complex. The present modeling of the complexes with histo-blood group A-active structures shows some contacts which provide insight into mutational data, explaining the involvement of I389 and Q331. Our results can be applicable in structure-based design of adhesion inhibitors of noroviruses.
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| 48. |
- Kronstrand, Robert, et al.
(författare)
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Incorporation of Selegiline Metabolites into Hair after Oral Selegiline Intake
- 2001
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Ingår i: Journal of Analytical Toxicology. - : Oxford University Press (OUP). - 0146-4760 .- 1945-2403. ; 25:7, s. 594-601
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Tidskriftsartikel (refereegranskat)abstract
- We have previously shown that melanin in human hair has a great impact on the incorporation of codeine into hair. The present study on 10 subjects was performed to investigate whether or not these findings could also be extrapolated to other therapeutic drugs. We chose selegiline because it metabolizes to two commonly abused central stimulants, methamphetamine and amphetamine. The results would therefore also be of interest when studying the intake of such drugs and their incorporation into human hair. Selegiline and metabolites were determined by gas chromatography-mass spectrometry, total melanin by spectrophotometry, and pyrroletricarboxylic acid by high-performance liquid chromatography with ultraviolet detection. Our results show strong positive exponential relationships (y = ex) between melanin and the metabolites, which for methamphetamine improved by normalizing for plasma area under the curve. We conclude that the major metabolites of selegiline can be detected in hair up to four weeks after a single oral dose and that the incorporation closely relates to the melanin contents.
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| 49. |
- Kunze, Angelika, 1978, et al.
(författare)
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Equilibrium-fluctuation-analysis of single liposome binding events reveals how cholesterol and Ca2+ modulate glycosphingolipid trans-interactions
- 2013
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 3
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Tidskriftsartikel (refereegranskat)abstract
- Carbohydrate-carbohydrate interactions (CCIs) are of central importance for several biological processes. However, the ultra-weak nature of CCIs generates difficulties in studying this interaction, thus only little is known about CCIs. Here we present a highly sensitive equilibrium-fluctuation-analysis of single liposome binding events to supported lipid bilayers (SLBs) based on total internal reflection fluorescence (TIRF) microscopy that allows us to determine apparent kinetic rate constants of CCIs. The liposomes and SLBs both contained natural Le(x) glycosphingolipids (Gal beta 4( Fuc alpha 3) GlcNAc beta 3Gal beta 4Glc beta 1Cer), which were employed to mimic cell-cell contacts. The kinetic parameters of the self-interaction between Le(x)-containing liposomes and SLBs were measured and found to be modulated by bivalent cations. Even more interestingly, upon addition of cholesterol, the strength of the CCIs increases, suggesting that this interaction is strongly influenced by a cholesterol-dependent presentation and/or spatial organization of glycosphingolipids in cell membranes.
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| 50. |
- Larson, Göran, 1953, et al.
(författare)
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Typing for the human lewis blood group system by quantitative fluorescence-activated flow cytometry: large differences in antigen presentation on erythrocytes between A(1), A(2), B, O phenotypes.
- 1999
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Ingår i: Vox sanguinis. - 0042-9007. ; 77:4, s. 227-36
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Lewis phenotyping by hemagglutination is an unreliable routine method for Lewis antigen designation. Now genomic typing of the Lewis gene is available. Additionally, flow cytometry has been used for typing. We wanted to compare the results of Lewis typing in healthy individuals using the three methods. MATERIALS AND METHODS: Ninety-three randomly selected plasma donors were genotyped for inactivating Secretor (FUT2) G428A and Lewis (FUT3) T59G, T202C, C314T, G508A and T1067A point mutations. All Le(a+b-) individuals (nonsecretors) were homozygous for the FUT2 G428A mutation and all Le(a-b-) individuals had inactivating mutations on both FUT3 alleles. Fixed erythrocytes were analyzed by fluorescence-activated flow cytometry and the results were compared with hem- agglutination and genotypic data. Antigen availability was expressed as median fluorescence intensity and as percentage positive cells with fluorescence intensities > or =10(2). RESULTS: Using an anti-Le(a) reagent a mean of 99% of erythrocytes from Le(a+b-) individuals and 1% of erythrocytes from Le(a-b-) or Le(a-b+) individuals were stained positive. Using an anti-Le(b) reagent, a mean of 71% of erythrocytes from A(1), 95% from B and 99% from O and A(2) Le(a-b+) individuals and less than 10% of erythrocytes from Le(a-b-) or Le(a+b-) individuals were stained positive. After papain treatment 100% of the erythrocytes from A(1) and A(1)B Le(a-b+) individuals stained positive without increase in background staining. The flow cytometric technique revealed large differences in staining intensities, within each ABO Le(a-b+) subgroup which was not directly correlated to plasma donation frequencies nor to Secretor or Lewis genotypes. CONCLUSION: Flow cytometry may prove valuable as a Lewis blood group typing technique but also as a research tool when investigating Lewis phenotypes of human erythrocytes.
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