Jonas Nilsson

University of Gothenburg | GU · Sambio Core Facilities Proteomics

Human noroviruses, globally the main cause of viral gastroenteritis, show strain specific affinity for histo-blood group antigens (HBGA) and can successfully be propagated ex vivo in human intestinal enteroids (HIEs). HIEs established from jejunal stem cells of individuals with different ABO, Lewis and secretor geno- and phenotypes, show varying su...

Influenza A virus (IAV) pandemics result from interspecies transmission events within the avian reservoir and further into mammals including humans. Receptor incompatibility due to differently expressed glycan structures between species has been suggested to limit zoonotic IAV transmission from the wild bird reservoir as well as between different b...

Fasciola hepatica is a global helminth parasite of humans and their livestock. The invasive stage of the parasite, the newly excysted juvenile (NEJs), relies on glycosylated excreted-secreted (ES) products and surface/somatic molecules to interact with host cells and tissues and to evade the host's immune responses, such as disarming complement and...

Introduction The glycosylation of immunoglobulin (Ig) G regulates IgG interaction capability with Fc gamma receptors found in all immune cells. In pathogenic conditions, estrogen can impact IgG levels and glycosylation. Following menopause, when estrogen levels decline affecting the immune system and potentially leading to a heightened susceptibili...

In recent years, several rational designed therapies have been developed for treatment of mucopolysaccharidoses (MPS), a group of inherited metabolic disorders in which glycosaminoglycans (GAGs) are accumulated in various tissues and organs. Thus, improved disease-specific biomarkers for diagnosis and monitoring treatment efficacy are of paramount...

Chondroitin sulfate proteoglycans (CSPGs) control key events in human health and disease and are composed of chondroitin sulfate (CS) polysaccharide(s) attached to different core proteins. Detailed information on the biological effects of site-specific CS structures is scarce as the polysaccharides are typically released from their core proteins pr...

In recent years, several rational designed therapies have been developed for treatment of mucopolysaccharidoses (MPS), a group of inherited metabolic disorders in which glycosaminoglycans (GAGs) are accumulated in various tissues and organs. Thus, improved disease-specific biomarkers for diagnosis and monitoring treatment efficacy are of paramount...

Background 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 anal...

Proteoglycans (PGs), a class of carbohydrate-modified proteins, are present in essentially all metazoan organisms investigated to date. PGs are composed of glycosaminoglycan (GAG) chains attached to various core proteins and are important for embryogenesis and normal homeostasis. PGs exert many of their functions via their GAG chains and understand...

The spondylodysplastic type of Ehlers–Danlos syndrome (spEDS) is caused by genetic defects in the B4GALT7 or B3GALT6 genes both deranging the biosynthesis of the glycosaminoglycan linkage region of chondroitin/dermatan sulfate and heparan sulfate proteoglycans. In this study, we have analyzed the linkage regions of urinary chondroitin sulfate prote...

The Sda histo-blood group antigen (GalNAcβ1-4(NeuAcα2-3)Galβ-R) is implicated in various infections and constitutes a potential biomarker for colon cancer. Sd(a−) individuals (2–4% of Europeans) may produce anti-Sda, which can lead to incompatible blood transfusions, especially if donors with the high-expressing Sd(a++)/Cad phenotype are involved....

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 g...

In this chapter, we describe a glycoproteomic approach for the identification of novel chondroitin sulfate proteoglycans (CSPGs) using a combination of biochemical enrichments, enzymatic digestions, and nanoscale liquid chromatography tandem mass spectrometry (nLC-MS/MS) analysis. The identification is achieved by trypsin digestion of CSPG-containi...

Chondroitin sulfate proteoglycans (CSPGs) are found at cell surfaces and in connective tissues, where they interact with a multitude of proteins involved in various pathophysiological processes. From a methodological perspective, the identification of CSPGs is challenging, as the identification requires the combined sequencing of specific core prot...

Proteoglycans (PGs) are proteins with glycosaminoglycan (GAG) chains, such as chondroitin sulfate (CS) or heparan sulfate (HS), attached to serine residues. We have earlier shown that prohormones can carry CS, constituting a novel class of PGs. The mapping of GAG modifications of proteins in endocrine cells may thus assist us in delineating possibl...

Glycoproteome profiling (glycoproteomics) remains a considerable analytical challenge that hinders rapid progress in glycobiology. The complex tandem mass spectra generated from glycopeptide mixtures require sophisticated analysis pipelines for structural determination. Diverse informatics solutions aiding the process have appeared, but their relat...

Of all post-translational modifications known, glycosaminoglycans (GAGs) remain one of the most challenging to study, and despite the recent years of advancement in MS technologies and bioinformatics, detailed knowledge about the complete structures of GAGs as part of proteoglycans (PGs) is limited. To address this issue, we have developed a protoc...

Influenza A virus (IAV) pandemics result from interspecies transmission events within the avian reservoir and further to mammals including humans. Investigating molecular virus–host interactions dictating this process and the adaptations to the new hosts that follow is vital to understand zoonotic IAV spread. Receptor incompatibility has been sugge...

The molecular mechanisms behind infection and propagation of human restricted pathogens such as human norovirus (HuNoV) have defied interrogation because they were previously unculturable. However, human intestinal enteroids (HIEs) have emerged to offer unique ex vivo models for targeted studies of intestinal biology, including inflammatory and inf...

Chondroitin sulfate (CS) is the placental receptor for the VAR2CSA malaria protein, expressed at the surface of infected erythrocytes during Plasmodium falciparum infection. Infected cells adhere to syncytiotrophoblasts or get trapped within the intervillous space by binding to a determinant in a 4-O-sulfated CS chains. However, the exact structure...

Chondroitin sulfate (CS) is the placental receptor for the VAR2CSA malaria protein, expressed at the surface of infected erythrocytes during Plasmodium falciparum infection. Infected cells adhere to syncytiotrophoblasts or get trapped within the intervillous space by binding to a determinant in a 4-O-sulfated CS chains. However, the exact structure...

Glycosaminoglycans (GAGs) are polysaccharides produced by most mammalian cells and involved in a variety of biological processes. However, due to the size and complexity of GAGs, detailed knowledge about the structure and expression of GAGs by cells, the glycosaminoglycome, is lacking. Here we report a straightforward and versatile approach for str...

Proteoglycans, a class of carbohydrate-modified proteins, often modulate growth factor signaling on the cell surface. However, the molecular mechanism by which proteoglycans regulate signal transduction is largely unknown. In this study, using a recently-developed glycoproteomic method, we found that Windpipe (Wdp) is a novel chondroitin sulfate pr...

Glycosylation is a topic of intense current interest in the development of biopharmaceuticals since 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, universit...

An early pathological hallmark of Alzheimer’s disease (AD) is amyloid-β (Aβ) deposits in the brain, which largely consist of up to 43 amino acids long Aβ peptides derived from the amyloid precursor protein (APP). We previously identified a series of sialylated Tyr-10 O-glycosylated Aβ peptides, 15–20 residues long, from human cerebrospinal fluid (C...

It is generally accepted that the biosynthesis of chondroitin sulfate and heparan sulfate is proceeding from a common linkage region tetrasaccharide comprising GlcA-Gal-Gal-Xyl-O-. The linkage region can undergo various modifications such as sulfation, phosphorylation, and sialylation, and as the methods for studying glycosaminoglycan structure hav...

A recombinant subunit vaccine (Shingrix®) was recently licensed for use against herpes zoster. This vaccine is based on glycoprotein E (gE) of varicella zoster virus (VZV), the most abundantly expressed protein of VZV, harboring sites for N- and O-linked glycosylation. The subunit vaccine elicits stronger virus-specific CD4+ T cell response as well...

Proteoglycans, a class of carbohydrate-modified proteins, often modulate growth factor signaling on the cell surface. However, the molecular mechanism by which proteoglycans regulate signal transduction is largely unknown. In this study, using a recently-developed glycoproteomic method, we found that Windpipe (Wdp) is a novel chondroitin sulfate pr...

Structural characterization of glycosaminoglycans remains a challenge but is essential for determining structure-function relationships between glycosaminoglycans and the biomolecules with which they interact and for gaining insight into the biosynthesis of glycosaminoglycans. We have recently reported that xyloside-primed chondroitin/dermatan sulf...

Distinct structural changes of the α2,3/α2,6 sialic acid glycosidic linkages on glycoproteins are of importance in cancer biology, inflammatory diseases and virus tropism. Current glycoproteomic methodologies are, however, not amenable toward high‐throughput characterization of sialic acid isomers. To enable such assignments, we developed a mass sp...

Distinct structural changes of the α2,3/α2,6 sialic acid glycosidic linkages on glycoproteins are of importance in cancer biology, inflammatory diseases and virus tropism. Current glycoproteomic methodologies are, however, not amenable toward high‐throughput characterization of sialic acid isomers. To enable such assignments, we developed a mass sp...

Chondroitin sulfate proteoglycans (CSPGs) are important structural components of connective tissues in essentially all metazoan organisms. In vertebrates, CSPGs are involved also in more specialized processes such as neurogenesis and growth factor signaling. In invertebrates, however, knowledge of CSPGs core proteins and proteoglycan-related functi...

Purification and liquid chromatography-tandem mass spectrometry (LC-MS/MS) characterization of glycopeptides, originating from protease digests of glycoproteins, enables site-specific analysis of protein N- and O-glycosylations. We have described a protocol to enrich, hydrolyze by chondroitinase ABC, and characterize chondroitin sulfate-containing...

Heparan sulfate (HS) and chondroitin sulfate (CS) are complex polysaccharides that regulate important biological pathways in virtually all metazoan organisms. The polysaccharides often display opposite effects on cell functions with HS and CS structural motifs presenting unique binding sites for specific ligands. Still, the mechanisms by which glyc...

Glycoproteomics has rapidly become an independent analytical platform bridging the fields of glycomics and proteomics to address site specific protein glycosylation and its impact in biology. Current glycopeptide characterization relies on time-consuming manual interpretations and demands high levels of personal expertise. Efficient data interpreta...

The use of liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS(n)) for the glycoproteomic characterization of glycopeptides is a growing field of research. The N- and O-glycosylated peptides (N- and O-glycopeptides) analyzed typically originate from protease-digested glycoproteins where many of them are expected to be...

Post-translational glycosylation of proteins play key roles in cellular processes and the site-specific characterisation of glycan structures is critical to understanding these events. Given the challenges regarding identification of glycan isomers, glycoproteomic studies generally rely on the assumption of conserved biosynthetic pathways. However,...

The inter-alpha-trypsin inhibitor (IαI) complex is a macromolecular arrangement of structurally related heavy chain proteins covalently cross-linked to the chondroitin sulfate (CS) chain of the proteoglycan bikunin. The IαI complex is abundant in plasma and associated with inflammation, kidney diseases, cancer and diabetes. Bikunin is modified at S...

The herpes simplex virus type 1 (HSV-1) glycoprotein gC-1, participating in viral receptor interactions and immunity interference, harbors a mucin-like domain with multiple clustered O-linked glycans. Using HSV-1 infected diploid human fibroblasts, an authentic target for HSV-1 infection, and a protein immunoaffinity procedure, we enriched fully gl...

Methods are provided for the prevention, treatment and diagnosis of Alzheimer's disease, based on the glycosylation pattern of amyloid-beta peptides in body fluids and tissues.

SorLA is a neuronal sorting receptor that is genetically associated with Alzheimer's disease. SorLA interacts directly with the amyloid precursor protein (APP) and affects the processing of the precursor, leading to a decreased generation of the amyloid-β (Aβ) peptide. The sorLA complement-type repeat (CR)-domains associate in vitro with APP, but t...

Protein glycosylation plays critical roles for 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 conser...

Unique tyrosine glycosylated amyloid-β(1-15) glycopeptides were synthesized with well-defined stereochemistry at the glycosidic linkages. Aided by these glycopeptides and tandem mass spectrometry analysis, the naturally existing amyloid-β glycopeptides, isolated from Alzheimer's disease patients, were determined to contain an α-linked N-acetyl gala...

Vertebrates produce various chondroitin sulfate proteoglycans (CSPGs) that are important structural components of cartilage and other connective tissues. CSPGs also contribute to the regulation of more specialized processes such as neurogenesis and angiogenesis. Although many aspects of CSPGs have been studied extensively, little is known of where...

Studies have suggested that the glycosphingolipid globoside (Gb4Cer) is a receptor for human parvovirus B19. Virus-like particles bind to Gb4Cer on thin-layer chromatograms, but a direct interaction between the virus and lipid membrane-associated Gb4Cer has been debated. Here, we characterized the binding of parvovirus B19 VP1/VP2 virus-like partic...

Glycogen synthesis is initiated by self-glucosylation of the glycosyltransferases glycogenin-1 and -2 that, in the presence of UDP-glucose, form both the first glucose-O-tyrosine linkage, and then stepwise add a series of α1,4-linked glucoses to a growing chain of variable length. Glycogen-1 and -2 coexist in liver glycogen preparations where the p...

One of the principal goals of glycoprotein research is to correlate glycan structure and function. Such correlation is necessary to understand the mechanisms whereby glycoprotein structure elaborates the functions of myriad proteins. Accurate comparison of glycoforms and quantification of glycosites is an essential step in this direction. Mass spec...

Extracellular glycoproteins frequently carry terminal sialic acids on their N-linked and/or O-linked glycan structures. In this chapter a sialic acid specific capture-and-release protocol for the enrichment of N- and O-glycopeptides originating from glycoproteins in complex biological samples is described. The enriched glycopeptides are subjected t...

Several herpesviruses induce expression of the selectin receptor sialyl-Lewis X (sLe(x) ) by activating transcription of one or more of silent host FUT genes, each one encoding a fucosyltransferase that catalyses the rate-limiting step of sLe(x) synthesis. The aim here was to identify the identity of the glycoconjugate associated with sLe(x) glycoe...

Despite numerous original publications describing the structural complexity of N- and O-linked glycans on glycoproteins, only very few answer the basic question of which particular glycans are linked to which amino acid residues along the polypeptide chain. Such structural information is of fundamental importance for understanding the biological ro...

Defects in the O-linked glycosylation of the peripheral membrane protein α-dystroglycan (α-DG) are the main cause of several forms of congenital muscular dystrophies and thus the characterization of the glycosylation of α-DG is of great medical importance. A detailed investigation of the glycosylation pattern of the native α-DG protein is essential...

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 p...

Norde´nNorde´n R, Nystro¨mNystro¨m K, Adamiak B, Halim A, Nilsson J, Larson G, Trybala E, Olofsson S. Involvement of viral glycoprotein gC-1 in expression of the selectin ligand sialyl-Lewis X induced after infection with herpes simplex virus type 1. APMIS 2013; 121: 280–89. Several herpesviruses induce expression of the selectin receptor sialyl-Le...

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 diff...

Glycogenin-1 initiates the glycogen synthesis in skeletal muscle by the autocatalytic formation of a short oligosaccharide at tyrosine 195. Glycogenin-1 catalyzes both the glucose-O-tyrosine linkage and the α1,4 glucosidic bonds linking the glucose molecules in the oligosaccharide. We recently described a patient with glycogen depletion in skeletal...

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...

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The glycosylation of the extracellular protein α-dystroglycan is important for its ligand-binding activity, and altered or blocked glycosylation is associated with several forms of congenital muscular dystrophies. By immunoprecipitation and sialic acid capture-and-release enrichment strategies, we isolated tryptic glycopeptides of α-dystroglycan fr...

We present a method to enrich for glycoproteins from proteomic samples. Sialylated glycoproteins were selectively periodate-oxidized, captured on hydrazide beads, trypsinized and released by acid hydrolysis of sialic acid glycosidic bonds. Mass spectrometric fragment analysis allowed identification of glycan structures, and additional fragmentation...

Noroviruses and norovirus virus-like particles (VLPs) exhibit strain specific patterns in their binding to ABH and Lewis histo-blood group antigens. In this study we demonstrate for the first time specific binding of Norwalk virus VLPs to type 1 and type 2 chain glycosphingolipids (GSLs) carrying ABH and Lewis antigens. N-succinimidyl-3-tributylsta...

The carbohydrate binding characteristics of a norovirus GII.3 (Chron1) and a GII.4 (Dijon) strain were investigated using virus-like particles (VLPs) and saliva samples from 81 individuals genotyped for FUT2 (secretor) and FUT3 (Lewis) and phenotyped for ABO and Lewis blood groups. The two VLPs showed a typical secretor-gene-dependent binding and b...

Here we report the first survey of galectins binding to glycoproteins of human serum. Serum was subjected to affinity chromatography using immobilized galectins, and the bound glycoproteins were analyzed by electrophoresis, Western blotting, and mass spectrometry. Galectins-3, -8, and -9 bound a much broader range of ligands in serum than previousl...

The different techniques used for analyzing protein folding were discussed. The design of polypeptides that fold into structures that are preorganized to form specific protein-protein interactions was also discussed. The construction of cavities was found to be a necessary step in the design of efficient catalysts and selective receptors.

Considerable progress has been made in the understanding of how to exploit hydrophobic and charge-charge interactions in forming binding sites for peptides and small molecules in folded polypeptide catalysts. This knowledge has enabled the introduction of feedback and control functions into catalytic cycles and the construction of folded polypeptid...

The second-order rate constants for the hydrolysis of nitrophenyl esters catalysed by a number of folded designed polypeptides have been determined, and 1900-fold rate enhancements over those of the 4-methylimidazole-catalysed reactions have been observed. The rate enhancements are much larger than those expected from the pKa depression of the nucl...

Peptides with 42 amino acid residues have been designed to fold into helix-loop-helix motifs that dimerize to form four-helix bundles and catalyze the hydrolysis of p-nitrophenyl esters. Their reactivities depend on nucleophilic and general-acid catalysis by cooperative HisH ⁺ -His pairs. The peptide catalyst MNV with the HisH ⁺ -His pair separated...

Peptides with 42 amino acid residues have been designed to fold into helix-loop-helix motifs that dimerize to form four-helix bundles and catalyze the hydrolysis of p-nitrophenyl esters. Their reactivities depend on nucleophilic and general-acid catalysis by cooperative HisH(+)-His pairs. The peptide catalyst MNV with the HisH(+)-His pair separated...

Four-helix bundle proteins have been designed that catalyze the hydrolysis and transesterification reactions of p-nitrophenyl esters by a cooperative nucleophilic and general acid mechanism. The catalysts consist of two 42-residue peptides that fold into helix-loop-helix motifs and dimerise. They have previously been shown to recognize anionic and...

Designed four-helix bundle proteins with reactive sites based on the cooperativity of HisH+−His pairs in helical sequences catalyze acyl-transfer reactions of p-nitrophenyl esters with large rate enhancements. The function of the HisH+−His site has been expanded by the introduction of flanking residues to provide recognition of substrate carboxylat...

A histidine-based two-residue reactive site for the catalysis of hydrolysis and transesterification reactions of p-nitrophenyl esters has been engineered into a helix in a designed helix−loop−helix motif, and it has been shown to function through cooperative nucleophilic and general-acid catalysis. The two-residue site has been expanded by the inco...

Receptor-active gangliosides with affinity for Helicobacter pylori and influenza virus were chemically modified and analyzed by negative ion fast atom bombardment mass spectrometry (FAB MS) or electron ionization mass spectrometry (EI MS) after permethylation. Derivatizations included mild periodate oxidation of the sialic acid glycerol tail or con...

Thesis (doctoral)--Linköpings Universitet, 2002.