Glycoproteomic Analysis of Seven Major Allergenic Proteins Reveals Novel Post-translational Modifications

Halim, Adnan; Carlsson, Michael C.; Madsen, Caroline B.; Brand, Stéphanie; Møller, Svenning Rune; Olsen, Carl Erik; Vakhrushev, Sergey Y.; Brimnes, Jens; Würtzen, Peter Adler; Ipsen, Henrik; Petersen, Bent Larsen; Wandall, Hans H.

doi:10.1074/mcp.m114.042614

Cited by 36 publications

(31 citation statements)

References 69 publications

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“…Purification of groups 1 and 2 from Der p was performed by ion exchange and size exclusion chromatography as previously described 15, 16, 17, 18. Purified proteins were separated by SDS‐PAGE to evaluate purity and stability (Figure 1).…”

Section: Methodsmentioning

confidence: 99%

Subcutaneous immunotherapy with purified Der p1 and 2 suppresses type 2 immunity in a murine asthma model

Hesse

Ieperen

Habraken

et al. 2018

Allergy

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BackgroundAllergen‐specific immunotherapy can induce long‐term suppression of allergic symptoms, reduce medication use, and prevent exacerbations of allergic rhinitis and asthma. Current treatment is based on crude allergen extracts, which contain immunostimulatory components such as β‐glucans, chitins, and endotoxin. Use of purified or recombinant allergens might therefore increase efficacy of treatment.AimsHere, we test application of purified natural group 1 and 2 allergens from Dermatophagoides pteronyssinus (Der p) for subcutaneous immunotherapy (SCIT) treatment in a house dust mite (HDM)‐driven mouse model of allergic asthma.Materials and methods HDM‐sensitized mice received SCIT with crude HDM extract, a mixture of purified Der p1 and 2 (DerP1/2), or placebo. Upon challenges, we measured specific immunoglobulin responses, allergen‐induced ear swelling response (ESR), airway hyperresponsiveness (AHR), and inflammation in bronchoalveolar lavage fluid (BAL) and lung tissue.Results ESR measurement shows suppression of early allergic response in HDM‐SCIT– and DerP1/2‐SCIT–treated mice. Both HDM‐SCIT and DerP1/2‐SCIT are able to suppress AHR and eosinophilic inflammation. In contrast, only DerP1/2‐SCIT is able to significantly suppress type 2 cytokines in lung tissue and BAL fluid. Moreover, DerP1/2‐SCIT treatment is uniquely able suppress CCL20 and showed a trend toward suppression of IL‐33, CCL17 and eotaxin levels in lung tissue.DiscussionTaken together, these data show that purified DerP1/2‐SCIT is able to not only suppress AHR and inflammation, but also has superior activity toward suppression of Th2 cells and HDM‐induced activation of lung structural cells including airway epithelium.ConclusionsWe postulate that treatment with purified natural major allergens derived from HDM will likely increase clinical efficacy of SCIT.

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Section: Methodsmentioning

confidence: 99%

Subcutaneous immunotherapy with purified Der p1 and 2 suppresses type 2 immunity in a murine asthma model

Hesse

Ieperen

Habraken

et al. 2018

Allergy

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“…(180)(181)(182). In food safety, these analyses are important for the determination of allergenicity of proteins and peptides (183,184). Recently, Ghaste et al (185) gave a comprehensive overview about the use of FT-ICR MS in metabolomics and lipidomics.…”

Section: Fourier Transform Ion Cyclotron Resonance Mass Spectrometrymentioning

confidence: 99%

“…The advantage of Orbitrap over FT-ICR is in its speed of up to 20 Hz, which is much more suitable for coupling with LC when analyzing samples of high complexity. Hybrid Orbitrap mass spectrometers can perform combined fragmentations, and this property of the instrument is currently the most important fragmentation scheme for the analysis of postt ranslational modifi cations of proteins (134,183,184,192). In food safety, these investigations have been shown to be crucial for the studying of allergenicity of proteins (183,184).…”

Section: Hybrid Mass Spectrometers -Orbitrap and Other Instrumentsmentioning

confidence: 99%

“…Hybrid Orbitrap mass spectrometers can perform combined fragmentations, and this property of the instrument is currently the most important fragmentation scheme for the analysis of postt ranslational modifi cations of proteins (134,183,184,192). In food safety, these investigations have been shown to be crucial for the studying of allergenicity of proteins (183,184). It has to be stressed that the ongoing optimization of speed, performance, practicability, collaboration with leading scientist in this fi eld from both academia and industry (33,34,134,173,183,184), its user friendliness and excellent marketing, make the Orbitrap a leading instrument for mass spectrometry analyses in foodomics (33,133,134,135,183,186,193), as well as one of the most expensive ones.…”

Section: Hybrid Mass Spectrometers -Orbitrap and Other Instrumentsmentioning

confidence: 99%

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Foodomics and Food Safety: Where We Are

Andjelković

Gajdošik

Gašo-Sokač

et al. 2017

Food Technol. Biotechnol.

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SummaryThe power of foodomics as a discipline that is now broadly used for quality assurance of food products and adulteration identifi cation, as well as for determining the safety of food, is presented. Concerning sample preparation and application, maintenance of highly sophisticated instruments for both high-performance and high-throughput techniques, and analysis and data interpretation, special att ention has to be paid to the development of skilled analysts. The obtained data shall be integrated under a strong bioinformatics environment. Modern mass spectrometry is an extremely powerful analytical tool since it can provide direct qualitative and quantitative information about a molecule of interest from only a minute amount of sample. Quality of this information is infl uenced by the sample preparation procedure, the type of mass spectrometer used and the analyst's skills. Technical advances are bringing new instruments of increased sensitivity, resolution and speed to the market. Other methods presented here give additional information and can be used as complementary tools to mass spectrometry or for validation of obtained results. Genomics and transcriptomics, as well as affi nity-based methods, still have a broad use in food analysis. Serious drawbacks of some of them, especially the affi nity-based methods, are the cross-reactivity between similar molecules and the infl uence of complex food matrices. However, these techniques can be used for pre-screening in order to reduce the large number of samples. Great progress has been made in the application of bioinformatics in foodomics. These developments enabled processing of large amounts of generated data for both identifi cation and quantifi cation, and for corresponding modeling.

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“…A particular advantage of intact MS is that it can quantitatively elucidate microheterogeneity and macroheterogeneity of the glycoprotein species (193)(194)(195)(196). One drawback of this technique is that it typically requires a high level of protein homogeneity, and a small number of putative sites of glycosylation.…”

Section: Mass Spectrometry For Protein Glycosylationmentioning

confidence: 99%

Structural characterisation of glycosylation of paramyxovirus attachment and fusion proteins by mass spectrometry

Pegg¹

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The family Paramyxoviridae (paramyxovirus) contains several significant human and animal pathogens.Represented within this family are human respiratory syncytial virus (hRSV) and Newcastle disease virus (NDV). The former contributes significantly to severe respiratory tract disease in infants, children and immunocompromised individuals. At present, highly efficacious therapeutics or safe and effective vaccines are not available for hRSV. NDV is the causative agent of Newcastle disease, afflicting a wide range of avian species. The desire to study NDV is due not only to the significant economic impact it has on the poultry industry worldwide, but also its potential use as an oncolytic agent and vaccine vector in humans and animals. Additionally, findings on NDV may be translated to closely related viruses that cause disease in humans, such as parainfluenza viruses.As outlined in Chapter 1 of this thesis, the infectious processes of all members of this family are driven by two major membrane glycoproteins whose ectodomains project from the viral envelope: the fusion (F) and attachment glycoproteins. The F glycoprotein is responsible for viral entry by means of fusion with host cell membranes while the variable attachment glycoproteins, haemagglutinin, haemagglutininneuraminidase (HN) and major surface glycoprotein (G), are involved in viral attachment to host cells.Previous studies have shown that altering the glycosylation profile of these proteins can modulate the ability of the virus to infect host cells and stimulate the host immune system. As yet, site-specific glycan heterogeneity of hRSV and NDV surface glycoproteins has not been defined at a chemical level. As described in Chapter 2, reverse-phase liquid chromatography tandem mass spectrometry (MS) strategies were implemented to characterise intact glycopeptides from the attachment and F glycoproteins of hRSV and NDV. Site-occupancy and monosaccharide compositions at a given site were determined using collision-induced dissociation, higher-energy collision dissociation, electron-transfer dissociation and electron-transfer dissociation combined with collision dissociation. As described in Chapter 3, a spectral processing program was developed called OxoExtract to aid identification of intact glycopeptides that were fragmented with higher-energy collision dissociation.The F and HN proteins of NDV were derived from virions propagated in embryonic eggs. Analyses of HN in Chapter 4 revealed high mannose N-linked glycans and complex or hybrid N-linked glycans that were variably fucosylated, sialylated and sulfated or phosphorylated. In total 63, 58, and 37 glycans were identified at sites N341, N433 and N481, respectively. In addition, a previously undocumented Olinked glycosylation site was identified in the stalk domain of the protein. Observed glycans from NDV F described in Chapter 5 were mainly high mannose, containing variations of five to nine mannose ii residues across four sites, N85, N191, N366 and N471. There was also evidence of fucosylated c...

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Glycoproteomic Analysis of Seven Major Allergenic Proteins Reveals Novel Post-translational Modifications

Cited by 36 publications

References 69 publications

Subcutaneous immunotherapy with purified Der p1 and 2 suppresses type 2 immunity in a murine asthma model

Subcutaneous immunotherapy with purified Der p1 and 2 suppresses type 2 immunity in a murine asthma model

Foodomics and Food Safety: Where We Are

Structural characterisation of glycosylation of paramyxovirus attachment and fusion proteins by mass spectrometry

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