Project description:Food proteins differ in their allergenic potential. Currently, there is no predictive and validated bio-assay to evaluate the allergenicity of novel food proteins. The objective of this study was to investigate the potential of a human peripheral blood mononuclear cell (PBMC) gene expression assay to identify biomarkers to predict the allergenicity of legume proteins. PBMCs from healthy donors were exposed to weakly and strongly allergenic legume proteins. Possible biomarkers for allergenicity were investigated by exposing PBMCs to a protein pair of weakly (white bean) and strongly allergenic (soybean) 7S globulins in a pilot experiment. Gene expression was measured by RNA-sequencing and differentially expressed genes were selected as biomarkers. 153 genes were identified as having significantly different expression levels to the 7S globulin of white bean compared to soybean.
Project description:Ulva sp., a promising yet underexplored food source, offers rich protein content and nutritional benefits. However, assessing its allergenicity remains challenging due to limited research and understanding. In this translational research, we focus on a laboratory-produced Ulva sp. product (UlvaLe) and gather comprehensive data to assess its potential allergenicity. Therefore, we employed a weight-of-evidence approach inspired by regulatory standards, conducting a literature review, proteomic analysis, and clinical trial. This article addresses challenges in assessing the allergenicity of novel foods, using Ulva sp. as an example, while enhancing the understanding of Ulva sp.'s allergenic potential and offers insights for future research and regulatory processes. Our findings reveal no reported cases of Ulva allergenicity in the literature but highlight the need to address polysaccharide safety. Proteins with homology exceeding 70% to known allergens were identified as putative allergens. Nonetheless, interim clinical trial results demonstrate no allergic reactions to UlvaLe ingestion, suggesting a favorable safety profile. The presented data suggests a promising allergenicity safety profile for UlvaLe as a novel food source. Further research into macro-algae allergenicity, including DNA and protein composition decoding, is warranted to ensure safe utilization in the food industry. Nevertheless, the most effective means of assessing allergenicity for any new food is through widespread consumption among large populations, which statistically reveals the true public risk. As a modern society seeking innovative solutions and sustainable growth, it is imperative to be mindful of this and devise strategies for its safe integration.
Project description:Food proteins differ in their allergenic potential. Currently, there is no predictive and validated bio-assay to evaluate the allergenicity of novel food proteins. The objective of this study was to investigate the potential of a human peripheral blood mononuclear cell (PBMC) gene expression assay to identify biomarkers to predict the allergenicity of legume proteins. PBMCs from healthy donors were exposed to weakly and strongly allergenic legume proteins. Inclusion of multiple protein pairs from 2S albumins (lupine and peanut) and 7S globulins (white bean and soybean) in a larger study, led to the selection of CCL2, CCL7, and RASD2 as biomarkers to distinguish weakly from strongly allergenic proteins.
Project description:Na+/H+ exchanger 1 (NHE1; SLC9A1) is a transport protein responsible for pH regulation in various types of cells. In cardiac myocytes, NHE1 generates the largest transmembrane acid-extrusion flux and is therefore the most significant controller of the intracellular acid/base milieu. The protein itself is regulated by various enzymes, including kinases. A number of critically important sites have been identified, coupling NHE1 to signalling cascades such as cAMP. NHE1 activity is very sensitive to oxygen tension, thus linking metabolism with pH. However, the sites implicated in this effect are not known. This study identified novel phosphorylation sites on NHE1. The state of these residues regulates NHE1 activity, hence pH and a myriad of pH sensitive downstream processes. Dysregulated NHE1 activity has been shown in diseases of the heart. Identification of novel regulatory sites can help demarcate the mechanisms of disease and suggest possible interventions to correct NHE1 activity.
Project description:Purpose: Delonix regia or Gulmohor commonly grows here and there in Indian villages as well as it is used for megacity beautification and environmental management due to its evergreen nature and vibrant flower colour. However, an increasing incidence of seasonal pollinosis was observed among the inhabitants living in close vicinity to this tree suggesting a possible link between the airborne pollen load and the concomitant respiratory hazards. This prompted us to investigate the allergens in the pollen of this dominant avenue tree. Methodology: Allergenicity of D. regia pollen grains was first checked by Skin Prick Test (SPT) and further confirmed by in vitro tests, such as, ELISA, Dot Blot and histamine release assay. The total proteome profiling was done by 2D PAGE and it was confronted with the pooled sera of 10 patients. The IgE reactive proteins were identified by MALDI TOF/TOF in Autoflex speed (Bruker,Germany). The raw spectra were searched against NCBInr database using MASCOT search engine. Result: Delonix regia, pollen grains have been found in considerable amount in the air during its flowering season (May to July). Approximately 31% of atopic individuals were found allergic to D. regia, pollen with elevated level of specific IgE and histamine in the serum. Total 8 IgE reactive proteins have been identified by homology driven proteomics. These proteins are ATP synthase beta subunit (spot no 5), Actin (spot no 6), Hypothetical actin like protein (spot no 7), Recombinant S-adenosylmethionine synthase 2 (spot no 9), UDP-arabinopyranose mutase (spot no 13), Luminal-binding 5 (spot no 2), ELF3-like protein (spot no 8) and hypothetical protein OsJ_04810 (spot no 11). Among these eight identified allergenic proteins, five were previously reported as allergen from different sources whereas the rest three have been reported for the first time as novel allergens. Conclusion: Novelty of this study was to identify 8 allergens from D.regia for the first time using immune-biochemical and proteomic techniques. Further studies will open up new avenues in component resolved diagnosis of pollen allergy.
Project description:Shotgun proteomics focusing on the membrane proteomes, of four Synechococcus spp. strains namely CC9311 (clade I), CC9605 (Clade II), WH8102 (clade III) and CC9902 (clade IV) were conducted to gain insight into the amount of resources these unicellular organisms invest into adaptation strategies.
Project description:To characterize the nature of the cytochrome c1 (CYC1) processivity defect in native CIII2 assemblies we identified CYC1 peptides using mass spectrometry analysis of blue native (BN)-PAGE. Gel slices ranging from ~600-900kDa and containing CIII2 assemblies were excised from U2OS control cells, U2OS OCIAD1 knockdown cells, and U2OS OCIAD1 knockdown cells rescued with wildtype OCIAD1. Gel slices were then digested and analyzed by mass spectrometry.
Project description:Glaucoma is an optic neuropathy, the second leading cause of irreversible blindness worldwide. In our earlier studies, we demonstrated the isolation and functional efficacy (wound healing and anti-oxidant potential) of small extracellular vesicles (sEV) from the adult tissue-resident stem cells of the trabecular meshwork (TMSC). This study aimed to elucidate the protein profile of sEV derived from TMSCs to correlate these functional differences for a better understanding of their potential as a cell-free therapeutic agents for primary open angle glaucoma (POAG). TM and TMSC sEV were isolated by ultracentrifugation and characterized. The protein cargo of these sEV were analyzed by Mass spectrometry. Mass spectrometry analysis identified 2802 proteins in TMSC sEV and 2848 in TM sEV. Further Maxquant analysis revealed that 511 proteins were reproducibly quantified across the samples. and 18% of the proteins were significantly altered between TMSC and TM sEV. Differential expression analysis identified distinct protein profiles between TMSC and TM sEV. Notably, sEV from TMSCs were enriched with proteins associated with wound healing, cell proliferation, migration, anti-oxidant and anti-apoptotic activities, consistent with the findings in other mesenchymal stem cells. Pathway analysis highlighted the enrichment of proteins associated with PI3K-AKT and MAPK signaling pathways. Further validation by western blotting confirmed that TMSC sEV effectively modulated these pathways in TM cells, which are essential for cell proliferation and survival under oxidative stress. To our knowledge, this is the first comprehensive protein profiling of TMSC-derived sEV, demonstrating the presence of functional proteins and their capacity to regulate the MAPK and PI3K-AKT signaling pathways in the recipient TM cells. This highlights the potential of TMSC sEV in advancing cell-free therapeutic strategies for POAG in the future.