| 1. |
- Deadman, Mary E., et al.
(författare)
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Specific amino acids of the glycosyltransferase LpsA direct the addition of glucose or galactose to the terminal inner core heptose of Haemophilus influenzae lipopolysaccharide via alternative linkages
- 2006
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 281:40, s. 29455-29467
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Tidskriftsartikel (refereegranskat)abstract
- Lipopolysaccharide is the major glycolipid of the cell wall of the bacterium Haemophilus influenzae, a Gram-negative commensal and pathogen of humans. Lipopolysaccharide is both a virulence determinant and a target for host immune responses. Glycosyltransferases have high donor and acceptor substrate specificities that are generally limited to catalysis of one unique glycosidic linkage. The H. influenzae glycosyltransferase LpsA is responsible for the addition of a hexose to the distal heptose of the inner core of the lipopolysaccharide molecule and belongs to the glycosyltransferase family 25. The hexose added can be either glucose or galactose and linkage to the heptose can be either beta 1-2 or beta 1-3. Each H. influenzae strain uniquely produces only one of the four possible combinations of linked sugar in its lipopolysaccharide. We show that, in any given strain, a specific allelic variant of LpsA directs the anomeric linkage and the added hexose, glucose, or galactose. Site-directed mutagenesis of a single key amino acid at position 151 changed the hexose added in vivo from glucose to galactose or vice versa. By constructing chimeric lpsA gene sequences, it was shown that the 3' end of the gene directs the anomeric linkage (beta 1-2 or beta 1-3) of the added hexose. The lpsA gene is the first known example where interstrain variation in lipopolysaccharide core structure is directed by the specific sequence of a genetic locus encoding enzymes directing one of four alternative possible sugar additions from the inner core.
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| 2. |
- Lundström, Susanna L., et al.
(författare)
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Application of capillary electrophoresis mass spectrometry and liquid chromatography multiple-step tandem electrospray mass spectrometry to profile glycoform expression during Haemophilus influenzae pathogenesis in the chinchilla model of experimental otitis media
- 2008
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Ingår i: Infection and Immunity. - 0019-9567 .- 1098-5522. ; 76:7, s. 3255-3267
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Tidskriftsartikel (refereegranskat)abstract
- Otitis media caused by nontypeable Haemophilus influenzae (NTHi) is a common and recurrent bacterial infection of childhood. The structural variability and diversity of H. influenzae lipopolysaccharide (LPS) glycoforms are known to play a significant role in the commensal and disease-causing behavior of this pathogen. In this study, we determined LPS glycoform populations from NTHi strain 1003 during the course of experimental otitis media in the chinchilla model of infection by mass spectrometric techniques. Building on an established structural model of the major LPS glycoforms expressed by this NTHi strain in vitro (M. Mansson, W. Hood, J. Li, J. C. Richards, E. R. Moxon, and E. K. Schweda, Eur. J. Biochem. 269:808-818, 2002), minor isomeric glycoform populations were determined by liquid chromatography multiple-step tandem electrospray mass spectrometry (LC-ESI-MSn). Using capillary electrophoresis ESI-MS (CE-ESI-MS), we determined glycoform profiles for bacteria from direct middle ear fluid (MEF) samples. The LPS glycan profiles were essentially the same when the MEF samples of 7 of 10 animals were passaged on solid medium (chocolate agar). LC-ESI-MSn provided a sensitive method for determining the isomeric distribution of LPS glycoforms in MEF and passaged specimens. To investigate changes in LPS glycoform distribution during the course of infection, MEF samples were analyzed at 2, 5, and 9 days postinfection by CE-ESI-MS following minimal passage on chocolate agar. As previously observed, sialic acid-containing glycoforms were detected during the early stages of infection, but a trend toward more-truncated and less-complex LPS glycoforms that lacked sialic acid was found as disease progressed.
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| 3. |
- Lundström, Susanna L., et al.
(författare)
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Asthmatics exhibit altered oxylipin profiles compared to healthy individuals after subway air exposure
- 2011
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Ingår i: PLOS ONE. - San Francisco, CA : Public Library of Science. - 1932-6203. ; 6:8, s. e23864-
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Tidskriftsartikel (refereegranskat)abstract
- Background: Asthma is a chronic inflammatory lung disease that causes significant morbidity and mortality worldwide. Air pollutants such as particulate matter (PM) and oxidants are important factors in causing exacerbations in asthmatics, and the source and composition of pollutants greatly affects pathological implications.Objectives: This randomized crossover study investigated responses of the respiratory system to Stockholm subway air in asthmatics and healthy individuals. Eicosanoids and other oxylipins were quantified in the distal lung to provide a measure of shifts in lipid mediators in association with exposure to subway air relative to ambient air.Methods: Sixty-four oxylipins representing the cyclooxygenase (COX), lipoxygenase (LOX) and cytochrome P450 (CYP) metabolic pathways were screened using liquid chromatography-tandem mass spectrometry (LC-MS/MS) of bronchoalveolar lavage (BAL)-fluid. Validations through immunocytochemistry staining of BAL-cells were performed for 15-LOX-1, COX-1, COX-2 and peroxisome proliferator-activated receptor gamma (PPAR gamma). Multivariate statistics were employed to interrogate acquired oxylipin and immunocytochemistry data in combination with patient clinical information.Results: Asthmatics and healthy individuals exhibited divergent oxylipin profiles following exposure to ambient and subway air. Significant changes were observed in 8 metabolites of linoleic- and alpha-linolenic acid synthesized via the 15-LOX pathway, and of the COX product prostaglandin E(2) (PGE(2)). Oxylipin levels were increased in healthy individuals following exposure to subway air, whereas asthmatics evidenced decreases or no change.Conclusions: Several of the altered oxylipins have known or suspected bronchoprotective or anti-inflammatory effects, suggesting a possible reduced anti-inflammatory response in asthmatics following exposure to subway air. These observations may have ramifications for sensitive subpopulations in urban areas.
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| 4. |
- Lundström, Susanna L., et al.
(författare)
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Novel globoside-like oligosaccharide expression patterns in nontypeable Haemophilus influenzae lipopolysaccharide
- 2007
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Ingår i: The FEBS Journal. - : Wiley. - 1742-464X .- 1742-4658. ; 274:18, s. 4886-4903
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Tidskriftsartikel (refereegranskat)abstract
- We report the novel pattern of lipopolysaccharide (LPS) expressed by two disease-associated nontypeable Haemophilus influenzae strains, 1268 and 1200. The strains express the common structural motifs of H. influenzae; globotetraose [beta-D-GalpNAc-(1 -> 3)-alpha-D-Galp-(1 -> 4)-beta-D-Galp-(1 -> 4)-beta-d-Glcp] and its truncated versions globoside [alpha-D-Galp-(1 -> 4)-beta-D-Galp-(1 -> 4)-beta-D-Glcp] and lactose [beta-D-Galp-(1 -> 4)-beta-D-Glcp] linked to the terminal heptose (HepIII) and the corresponding structures with an alpha-D-Glcp as the reducing sugar linked to the middle heptose (HepII) in the same LPS molecule. Previously these motifs had been found linked only to either the proximal heptose (HepI) or HepIII of the triheptosyl inner-core moiety l-alpha-D-Hepp-(1 -> 2)-[PEtn -> 6]-l-alpha-D-Hepp-(1 -> 3)-l-alpha-D-Hepp-(1 -> 5)-[PPEtn -> 4]-alpha-Kdo-(2 -> 6)-lipid A. This novel finding was obtained by structural studies of LPS using NMR techniques and ESI-MS on O-deacylated LPS and core oligosaccharide material, as well as electrospray ionization-multiple-step tandem mass spectrometry on permethylated dephosphorylated oligosaccharide material. A lpsA mutant of strain 1268 expressed LPS of reduced complexity that facilitated unambiguous structural determination. Using capillary electrophoresis-ESI-MS/MS we identified sialylated glycoforms that included sialyllactose as an extension from HepII, this is a further novel finding for H. influenzae LPS. In addition, each LPS was found to carry phosphocholine and O-linked glycine. Nontypeable H. influenzae strain 1200 expressed identical LPS structures to 1268 with the difference that strain 1200 LPS had acetates substituting HepIII, whereas strain 1268 LPS has glycine at the same position.
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| 5. |
- Lundström, Susanna L., et al.
(författare)
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Structural analysis of the lipopolysaccharide from nontypeable Haemophilus influenzae strain R2846
- 2008
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Ingår i: Biochemistry. - : American Chemical Society (ACS). - 0006-2960 .- 1520-4995. ; 47:22, s. 6025-6038
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Tidskriftsartikel (refereegranskat)abstract
- We here report the lipopolysaccharide (LPS) structures expressed by nontypeable Haemophilus influenzae R2846, a strain whose complete genome sequence has recently been obtained. Results were obtained by using NMR techniques and ESI-MS on O-deacylated LPS and core oligosaccharide material (OS) as well as ESI-MSn on permethylated dephosphorylated OS. A beta-D-Glcp-(1 -> 4)-D-alpha-D-Hepp-(1 -> 6)-beta-D-Glcp-(1 -> 4) unit was found linked to the proximal heptose (HepI) of the conserved triheptosyl inner-core moiety, L-alpha-D-Hepp-(1 -> 2)-[PEtn -> 6]-L-alpha-D-Hepp-(1 -> 3)-L-alpha-D-Hepp-(1 -> 5)-[PPEtn -> 4]-alpha-Kdo-(2 -> 6)-lipid A. The beta-D-Glcp (GlcI) linked to HepI was also branched with oligosaccharide extensions from O-4 and O-6. O-4 of GlcI was substituted with sialyllacto-N-neotetraose [alpha-Neu5Ac-(2 -> 3)-beta-D-Galp-(1 -> 4)-beta-GlcpNAc-(1 -> 3)-beta-D-Galp-(1 -> 4)-beta-D-Glcp-(1 ->] and the related structure [(PEtn -> 6)-alpha-D-GalpNAc-(1 -> 6)-beta-D-Galp-(1 -> 4)-beta-D-GlcpNAc-(1 -> 3)-beta-D-Galp-(1 -> 4)-beta-Glcp-(1-]. The distal heptose (HepIII) was substituted at O-2 by beta-D-Gal. Phosphate, phosphoethanolamine, phosphocholine, acetate, and glycine were found to substitute the core oligosaccharide. Two heptosyltransferase genes, losB1 and losB2, have been identified from the R2846 genome sequence and are candidates to add the noncore heptose to the LPS. Mutant strain R2846losB1 did not show DD-heptose in the extension from HepI but still contained minor quantities of LD-heptose at the same position, indicating that the losB1 gene is required to add DD-heptose to Glcl. The LPS from strain R2846losB1/losB2 expressed no noncore heptose, consistent with losB2 directing the addition of LD-heptose.
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| 6. |
- Nandakumar, Kutty Selva, 1965-, et al.
(författare)
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Streptococcal Endo-β-N-Acetylglucosaminidase Suppresses Antibody-Mediated Inflammation In Vivo
- 2018
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Ingår i: Frontiers in Immunology. - : Frontiers Media SA. - 1664-3224. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Endo-β-N-acetylglucosaminidase (EndoS) is a family 18 glycosyl hydrolase secreted by Streptococcus pyogenes. Recombinant EndoS hydrolyzes the β-1,4-di-N-acetylchitobiose core of the N-linked complex type glycan on the asparagine 297 of the γ-chains of IgG. Here, we report that EndoS and IgG hydrolyzed by EndoS induced suppression of local immune complex (IC)-mediated arthritis. A small amount (1 µg given i.v. to a mouse) of EndoS was sufficient to inhibit IgG-mediated arthritis in mice. The presence of EndoS disturbed larger IC lattice formation both in vitro and in vivo, as visualized with anti-C3b staining. Neither complement binding in vitro nor antigen-antibody binding per se were affected. Thus, EndoS could potentially be used for treating patients with IC-mediated pathology.
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| 7. |
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| 8. |
- Saei, Amir Ata, et al.
(författare)
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System-wide identification and prioritization of enzyme substrates by thermal analysis
- 2021
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Ingår i: Nature Communications. - : Springer Nature. - 2041-1723. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- Despite the immense importance of enzyme-substrate reactions, there is a lack of general and unbiased tools for identifying and prioritizing substrate proteins that are modified by the enzyme on the structural level. Here we describe a high-throughput unbiased proteomics method called System-wide Identification and prioritization of Enzyme Substrates by Thermal Analysis (SIESTA). The approach assumes that the enzymatic post-translational modification of substrate proteins is likely to change their thermal stability. In our proof-of-concept studies, SIESTA successfully identifies several known and novel substrate candidates for selenoprotein thioredoxin reductase 1, protein kinase B (AKT1) and poly-(ADP-ribose) polymerase-10 systems. Wider application of SIESTA can enhance our understanding of the role of enzymes in homeostasis and disease, opening opportunities to investigate the effect of post-translational modifications on signal transduction and facilitate drug discovery.
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| 9. |
- Wang, Jijing, et al.
(författare)
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First Immunoassay for Measuring Isoaspartate in Human Serum Albumin
- 2021
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Ingår i: Molecules. - : MDPI. - 1431-5157 .- 1420-3049. ; 26:21
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Tidskriftsartikel (refereegranskat)abstract
- Isoaspartate (isoAsp) is a damaging amino acid residue formed in proteins mostly as a result of spontaneous deamidation of asparaginyl residues. An association has been found between isoAsp in human serum albumin (HSA) and Alzheimer’s disease (AD). Here we report on a novel monoclonal antibody (mAb) 1A3 with excellent specificity to isoAsp in the functionally important domain of HSA. Based on 1A3 mAb, an indirect enzyme-linked immunosorbent assay (ELISA) was developed, and the isoAsp occupancy in 100 healthy plasma samples was quantified for the first time, providing the average value of (0.74 ± 0.13)%. These results suggest potential of isoAsp measurements for supplementary AD diagnostics as well as for assessing the freshness of stored donor blood and its suitability for transfusion.
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| 10. |
- Wang, Jijing, et al.
(författare)
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SpotLight Proteomics Identifies Variable Sequences of Blood Antibodies Specific Against Deamidated Human Serum Albumin
- 2023
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Ingår i: Molecular & Cellular Proteomics. - : Elsevier. - 1535-9476 .- 1535-9484. ; 22:7
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Tidskriftsartikel (refereegranskat)abstract
- Spontaneous deamidation of asparaginyl residues in proteins, if not repaired or cleared, can set in motion a cascade that leads to deteriorated health. Previously, we have discovered that deamidated human serum albumin (HSA) is elevated in the blood of patients with Alzheimer's disease and other neurodegenerative diseases, while the level of endogenous antibodies against deamidated HSA is significantly diminished, creating an imbalance between the risk factor and the defense against it. Endogenous antibodies against deamidated proteins are still unexplored. In the current study, we employed the SpotLight proteomics approach to identify novel amino acid sequences in antibodies specific to deamidated HSA. The results provide new insights into the clearance mechanism of deamidated proteins, a possible avenue for prevention of neurodegeneration.
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