Identification of Cross-Species Marker Peptides for the Detection of Mammalian and Poultry Meat in Vegan and Vegetarian Foods.

Authentication of vegan and vegetarian foods is important since these increasingly popular food items could be adulterated with cheap meat to increase profit margins. In this study, nine marker peptides for the detection of meat (several species) were identified applying liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). These marker peptides enable the crucial differentiation of beef versus milk and chicken meat versus egg, demonstrated by the investigation of 19 commercial vegetarian meat substitutes containing milk and egg.

Genome-wide deposition of 6-methyladenine in human DNA reduces the viability of HEK293 cells and directly influences gene expression.

While cytosine-C5 methylation of DNA is an essential regulatory system in higher eukaryotes, the presence and relevance of 6-methyladenine (m6dA) in human cells is controversial. To study the role of m6dA in human DNA, we introduced it in human cells at a genome-wide scale at GANTC and GATC sites by expression of bacterial DNA methyltransferases and observed concomitant reductions in cell viability, in particular after global GANTC methylation. We identified several genes that are directly regulated by m6dA in a GANTC context.

Characterization and Quantification of Mal d 1 Isoallergen Profiles and Contents in Traditional and Commercial Apple Varieties by Mass Spectrometry.

The apple allergy in Northern Europe is a cross-reaction to the birch pollen allergy. No correlation between the allergenicity of an apple variety and the content of the major apple allergen Mal d 1, a homologue to the Bet v 1 allergen in birch, could be found using ELISA, so far. Therefore, an impact of polyphenols and/or differences in the isoallergen profile are discussed.

The R736H cancer mutation in DNMT3A modulates the properties of the FF-subunit interface.

The DNMT3A DNA methyltransferase is an important epigenetic enzyme that is frequently mutated in cancers, particularly in AML. The heterozygous R736H mutation in the FF-interface of the tetrameric enzyme is the second most frequently observed DNMT3A cancer mutation, but its pathogenic mechanism is unclear. We show here that R736H leads to a moderate reduction in catalytic activity of 20-40% depending on the substrate, but no changes in CpG specificity, flanking sequence preferences and subnuclear localization.

The lipidome of an omnivorous insect responds to diet composition and social environment.

Lipids are biomolecules with essential roles in metabolic processes, signaling, and cellular architecture. In this study, we investigated changes in the lipidome of the house cricket subjected to diets of different nutritional composition (i.e.

New Mass Spectrometric Approach to Quantify the Major Isoallergens of the Apple Allergen Mal d 1.

Patients who suffer from birch pollinosis often develop adverse reactions to the consumption of fresh apples due to the structural similarity of the allergens Bet v 1 and Mal d 1 from birch and apples, respectively. A different allergenic potential for Mal d 1 isoallergens is postulated, but approaches to quantify the Mal d 1 isoallergen-specific are missing. Therefore, a bottom-up proteomics approach was developed to quantify Mal d 1 by stable isotope dilution and microHPLC-QTOF analyses.

Relative quantification of pork and beef in meat products using global and species-specific peptide markers for the authentication of meat composition.

We used global and species-specific peptide markers for a relative quantitative determination of pork and beef in raw and processed meat products made of the two meat species. Four groups of products were prepared (i.e.

Flanking sequences influence the activity of TET1 and TET2 methylcytosine dioxygenases and affect genomic 5hmC patterns.

TET dioxygenases convert 5-methylcytosine (5mC) preferentially in a CpG context into 5-hydroxymethylcytosine (5hmC) and higher oxidized forms, thereby initiating DNA demethylation, but details regarding the effects of the DNA sequences flanking the target 5mC site on TET activity are unknown. We investigated oxidation of libraries of DNA substrates containing one 5mC or 5hmC residue in randomized sequence context using single molecule readout of oxidation activity and sequence and show pronounced 20 and 70-fold flanking sequence effects on the catalytic activities of TET1 and TET2, respectively. Flanking sequence preferences were similar for TET1 and TET2 and also for 5mC and 5hmC substrates.

Identification and Characterization of IgE-Reactive Proteins and a New Allergen (Cic a 1.01) from Chickpea (Cicer arietinum).

Scope: Chickpea (Cicer arietinum) allergy has frequently been reported particularly in Spain and India. Nevertheless, chickpea allergens are poorly characterized. The authors aim to identify and characterize potential allergens from chickpea.

Diet composition and social environment determine food consumption, phenotype and fecundity in an omnivorous insect.

Nutrition is the single most important factor for individual's growth and reproduction. Consequently, the inability to reach the nutritional optimum imposes severe consequences for animal fitness. Yet, under natural conditions, organisms may face a mixture of stressors that can modulate the effects of nutritional asymmetry.

German Government Official Methods Board Points the Way Forward: Launch of a New Working Group for Mass Spectrometry for Protein Analysis to Detect Food Fraud and Food Allergens.

The detection of food fraud and undeclared food allergens is one of the major challenges for competent authorities. Because adulterations are continuously adapted to the methods used to uncover them, the accomplishment of this task has become increasingly difficult over time. In recent years, various new promising methods for the detection of multiple food adulterants and multiple food allergens have been developed.

Mass Spectrometry-Based Untargeted Proteomics for the Assessment of Food Authenticity: The Case of Farmed Versus Wild-Type Salmon.

Omics technologies have been widely applied in different fields, among which, proteomics is gaining increasing interest for its application to the authenticity of food products. MS, typically coupled with LC, represents a key technique for proteomics-related studies dedicated to fish and other seafood products by using a bottom-up approach. In this paper, the optimization of an untargeted proteomics-based method using LC separation and MS detection relying on a quadrupole time-of-flight mass spectrometer is described and applied to the analysis of Canadian farmed and wild-type salmon, followed by statistical analysis based on principal component (PC) analysis.

Determination of food allergens by LC-MS: Impacts of sample preparation, food matrix, and thermal processing on peptide detectability and quantification.

Food allergies are a growing worldwide concern and the contamination of products with food allergens represents a significant health risk to allergic consumers. With the introduction of reference doses, quantitative methods are needed for the monitoring of allergen levels, and the potential of LC-MS/MS is of hugely growing interest. In this study, we demonstrate that relevant food matrices (bakery products and chocolates) and thermal food processing substantially influence the quantification of 18 marker peptides from various nut and peanut allergens via targeted proteomics.

Characterization and Detection of Food Allergens Using High-Resolution Mass Spectrometry: Current Status and Future Perspective.

Allergic reactions to food are among the major food safety concerns in industrialized countries, and it is estimated that approximately 5% of the population suffers from immunoglobulin-E-mediated food allergy. High-resolution mass spectrometry has become one of the most important techniques for the molecular characterization of allergens, including structural modification, degradation in the gastrointestinal environment, or identification of suitable marker peptides for the development of novel analytical approaches, in the past decade. This perspective aims to briefly summarize the current situation and discuss future developments.

Novel Approaches for the MS-Based Detection of Food Allergens: High Resolution, MS³, and Beyond.

The prevalence of allergic reactions to food is believed to be increasing in industrialized countries worldwide. One of the major tasks in risk management is, therefore, the analytical surveillance of allergen contamination in food and targeted proteomics using MS, which is of hugely growing interest due to its specificity and sensitivity and the possibility to analyze multiple allergens in parallel. Though approximately 200 different foods have been described as having the potential to elicit allergic reactions, current regional labeling requirements are focused on the 5-14 priority allergens that elicit the vast majority of severe reactions or that pose a risk as hidden allergens in food production.

Gastrointestinal digestion of hazelnut allergens on molecular level: Elucidation of degradation kinetics and resistant immunoactive peptides using mass spectrometry.

Scope: Allergy to hazelnut seeds ranks among the most prevalent food allergies in Europe. The aim of this study was to elucidate the gastrointestinal digestion of hazelnut allergens on molecular level.

Methods And Results: Hazelnut flour was digested in vitro following the Infogest consensus model.

Structural Characterization of the Allergenic 2S Albumin Cor a 14: Comparing Proteoform Patterns across Hazelnut Cultivars.

The hazelnut allergen Cor a 14 belongs to the 2S albumins, a family of heterodimeric seed storage proteins exhibiting a high degree of structural diversity. Given its relevance as an allergen and the potential to elicit severe reactions, elucidation of the sequence heterogeneity of naturally occurring Cor a 14 is essential for the development of reliable diagnostics and risk evaluation. We therefore performed a comprehensive survey on the proteoforms of Cor a 14 and determined their quantitative distribution in three different hazelnut cultivars by a combinatory HPLC-HRMS approach including bottom-up and intact mass analysis.

MRM-based LC-MS multi-method for the detection and quantification of nut allergens.

Food allergies have become a global challenge to food safety in industrialized countries in recent years. With governmental monitoring and legislation moving towards the establishment of threshold allergen doses, there is a need for sensitive and quantitative analytical methods for the determination of allergenic food contaminants. Targeted proteomics employing liquid chromatography-mass spectrometry (LC-MS) has emerged as a promising technique that offers increased specificity and reproducibility compared to antibody and DNA-based technologies.

New High-Performance Liquid Chromatography Coupled Mass Spectrometry Method for the Detection of Lobster and Shrimp Allergens in Food Samples via Multiple Reaction Monitoring and Multiple Reaction Monitoring Cubed.

Crustacean shellfish allergy ranks among the most frequent and severe food allergies for adults, demanding rugged and sensitive analytical routine methods. The objective of this study was therefore to develop a mass spectrometric approach for the detection of contamination with shrimp and lobster, two economically important types of crustaceans, in complex food matrices. Following a biomarker approach, we identified proteotypic peptides and developed a multiple reaction monitoring (MRM) method allowing for the identification and differentiation of shrimp and lobster in the food matrix at concentrations down to 0.

Comprehensive peptide marker identification for the detection of multiple nut allergens using a non-targeted LC-HRMS multi-method.

Food allergies have emerged as a global problem over the last few decades; therefore, reliable and sensitive analytical methods to ensure food safety for allergic consumers are required. The application of mass spectrometry is of growing interest in this field and several procedures based on low resolution tandem mass spectrometry using single tryptic peptides as analytical targets have recently been described. However, a comprehensive survey of marker peptides for the development of multi-methods is still missing, as is a consensus guide to marker identification.

Simple, Rapid, and Selective Isolation of 2S Albumins from Allergenic Seeds and Nuts.

The 2S albumins belong to the group of seed storage proteins present in different seeds and nuts. Due to their pronounced allergenic potential, which is often associated with severe allergic reactions, this protein family is of special interest in the field of allergen research. Here we present a simple, rapid, and selective method for the purification of 2S albumins directly from allergenic seeds and nuts.

Characterization of mustard 2S albumin allergens by bottom-up, middle-down, and top-down proteomics: a consensus set of isoforms of Sin a 1.

The mustard allergen Sin a 1 belongs to the 2S-albumin family of seed-storage proteins. Because of its high abundance in mustard seeds and the potential to elicit severe allergenic reactions, Sin a 1 is considered to be a major allergen in mustard. Eight Sin a 1 isoforms have been identified using DNA cloning and sequencing, and we aim in this study to thoroughly investigate sequence heterogeneity using a novel combination of bottom-up, middle-down, and top-down proteomics.

Structural and functional characterization of cleavage and inactivation of human serine protease inhibitors by the bacterial SPATE protease EspPα from enterohemorrhagic E. coli.

EspPα and EspI are serine protease autotransporters found in enterohemorrhagic Escherichia coli. They both belong to the SPATE autotransporter family and are believed to contribute to pathogenicity via proteolytic cleavage and inactivation of different key host proteins during infection. Here, we describe the specific cleavage and functional inactivation of serine protease inhibitors (serpins) by EspPα and compare this activity with the related SPATE EspI.

Biochemical characterization of the SPATE members EspPα and EspI.

The activity of serine proteases is influenced by their substrate specificity as well as by the physicochemical conditions. Here, we present the characterization of key biochemical features of the two SPATE members EspPα and EspI from Shiga-toxin producing Escherichia coli (STEC) and enterohemorrhagic E. coli (EHEC).

Meat authentication: a new HPLC-MS/MS based method for the fast and sensitive detection of horse and pork in highly processed food.

Fraudulent blending of food products with meat from undeclared species is a problem on a global scale, as exemplified by the European horse meat scandal in 2013. Routinely used methods such as ELISA and PCR can suffer from limited sensitivity or specificity when processed food samples are analyzed. In this study, we have developed an optimized method for the detection of horse and pork in different processed food matrices using MRM and MRM(3) detection of species-specific tryptic marker peptides.