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Sve veći globalni problem predstavlja zagađenje vodenih sistema razičitim otpadnim materijama, pre svega fenolnim jedinjenjima i organskim bojama ispuštenim iz mnogobrojnih industrijskih postrojenja. S obzirom da su fenoli označeni kao najčešći zagađivači voda širom sveta, pronalazak metoda za njihovo uklanjanje predstavlja jedan od najvećih izazova savremene naučne zajednice. Različiti biokatalizatori su pronašli primenu u ove svrhe, a među njima je posebno mesto zauzela peroksidaza iz rena (HRP). Kao najveće ograničenje ove metode navodi se inaktivacija enzima, koja se može prevazići njegovom imobilizacijom na različitim nosačima. U ovom radu su razvijene metode za uklanjanje fenola i degradaciju tekstilnih boja imobilizovanom peroksidazom iz rena i ćelijskim zidovima sa lignin-peroksidazom (LiP). Kao nosači za imobilizaciju su korišćeni oksidovani alginati modifikovani tiraminom, makroporozni kopolimeri različite veličine pora, kao i neorganski glineni nosač – sepiolit. Tiramin-alginatne mikro-kuglice su iskorišćene za enkapsulaciju peroksidaze iz rena primenom kuplovane emulziono-polimerizacione reakcije. Ispitan je uticaj stepena modifikacije alginata tiraminom na specifičnu aktivnost enzima. Ustanovljeno je da sa povećanjem koncentracije perjodata korišćenog za oksidaciju alginata od 2,5 do 20 mol% pre modifikacije tiraminom dolazi do postepenog povećanja specifične aktivnosti. Imobilizovana HRP je potom upotrebljena za uklanjanje fenola u šaržnom reaktoru. Optimizovani su reakcioni uslovi sa ciljem postizanja visokog stepena efikasnosti i poboljšane kontinualne primene. Za interno generisanje vodonik-peroksida (H2O2) tokom oksidacije fenola primenjen je sistem sastavljen od glukoze i glukoza-oksidaze. Znatno veći stepen uklanjanja fenola (96%) iz reakcionog sistema za 6 h je postignut primenom interne dostave H2O2, dok se njegovim direktnim dodavanjem značajno smanjuje efikasnost uklanjanja fenola na 42%. Primenom glukoza-oksidaze u koncentraciji od 0,187 U/m, A growing global concern is the pollution of water courses with various waste materials, primarly phenolic compounds and organic dyes released from numerous industrial plants. Since phenols are designated as the most common water pollutants worldwide, finding methods for their removal is nowadays one of the biggest challenges for the broad scientific community. To this end, various biocatalysts have been used, among them Horseradish peroxidase (HRP) has been in prime position. The main limitation of this method is an enzyme inactivation, which can be overcome by its immobilization on different carriers. In this study, various methods for the removal of phenol and degradation of textile dyes by immobilized Horseradish peroxidase and cell walls with lignin peroxidase (LiP) were developed. Oxidized alginates modified with tyramine, macroporous copolymers with different pore characteristics, as well as inorganic clay mineral – sepiolite, were used as carriers in immobilization. Tyramine-alginate micro-beads were used for the encapsulation of peroxidase by coupled emulsion polymerization reaction. An effect of a degree of the alginate modification by tyramine on the specific activity of enzyme was examined. A gradual increase in the specific activity, with an increase in the concentration of periodate used for oxidation of alginate from 2.5 to 20 mol% prior to modification by tyramine, was observed. Immobilized HRP was then used for the removal of phenol in a batch reactor. Reaction conditions were optimized with an aim of achieving a high degree of efficiency and improved reusability. A system composed of glucose and glucose oxidase was used for internal generation of hydrogen peroxide (H2O2). When internal delivery of H2O2 was employed, 96% of phenol was removed from the reaction system in 6 h, whereas its straightforward addition significantly reduced the efficiency of phenol removal (42%). The highest efficiency was achieved by using glucose oxidase at the concentrat

Banana allergy is often associated with the pollen and latex allergies, which led us to the hypothesis that some yet unidentified banana allergen could provide a basis of the latex-pollen-fruit syndrome. S-adenosyl-L-homocysteine hydrolase (SAHH) was recently identified in the literature as a novel plant allergen. This study aimed to assess the allergenic potential of the naturally occurring banana SAHH (nSAHH) and its recombinant homolog produced in E. coli (rSAHH). nSAHH showed IgE reactivity with a serum pool of twelve banana-allergic persons, while rSAHH displayed IgE reactivity in ten out of the twelve tested patients. Five linear B-cell epitopes were identified on the rSAHH surface, exhibiting ≥ 90 % sequence homology with relevant plant SAHH allergens. Our findings have elucidated SAHH as a novel plant panallergen, underlying the cross-reactivity between plant derived food and respiratory allergens, confirming our initial hypothesis.

Supplementary Table S2: A)List of T-cell epitopes in kiwifruit SAHH. B)List of T cell epitopes in latex SAHH.C)List of T-cell epitopes in ryegrass SAHH

Food allergies mediated by specific IgE (sIgE) have a significant socioeconomic impact on society. Evaluating the IgE cross-reactivity between allergens from different allergen sources can enable the better management of these potentially life-threatening adverse reactions to food proteins and enhance food safety. A novel banana fruit allergen, S-adenosyl-L-homocysteine hydrolase (SAHH), has been recently identified and its recombinant homolog was heterologously overproduced in E. coli. In this study, we performed a search in the NCBI (National Center for Biotechnology Information) for SAHH homologs in ryegrass, latex, and kiwifruit, all of which are commonly associated with pollen-latex-fruit syndrome. In addition, Western immunoblot analysis was utilized to identify the cross-reactive IgE to banana SAHH in the sera of patients with a latex allergy, kiwifruit allergy, and ryegrass allergy. ClustalOmega analysis showed more than 92% amino acid sequence identity among the banana SAHH homologs in ryegrass, latex, and kiwifruit. In addition to five B-cell epitopes, in silico analysis predicted eleven T-cell epitopes in banana SAHH, seventeen in kiwifruit SAHH, twelve in ryegrass SAHH, and eight in latex SAHH, which were related to the seven-allele HLA reference set (HLA-DRB1*03:01, HLA-DRB1*07:01, HLA-DRB1*15:01, HLA-DRB3*01:01, HLA-DRB3*02:02, HLA-DRB4*01:01, HLA-DRB5*01:01). Four T-cell epitopes were identical in banana and kiwifruit SAHH (positions 328, 278, 142, 341), as well as banana and ryegrass SAHH (positions 278, 142, 96, and 341). All four SAHHs shared two T-cell epitopes (positions 278 and 341). In line with the high amino acid sequence identity and B-cell epitope homology among the analyzed proteins, the cross-reactive IgE to banana SAHH was detected in three of three latex-allergic patients, five of six ryegrass-allergic patients, and two of three kiwifruit-allergic patients. Although banana SAHH has only been studied in a small group of allergic individu

Allergen immunotherapy (AIT) is currently the only disease-modifying treatment forallergies. Pre-clinical models for the evaluation of novel therapeutics are crucial forensuring their efficacy and safety. While cell culture models are cost-effective andefficient, they cannot fully replicate the cellular interactions in vivo. Therefore, it isessential to use more sophisticated model systems, such as co-cultures, to assess thepotential of new therapeutics more accurately. Immunomodulatory protein banana lectin(BLwt) is an attractive candidate for adjuvant in AIT. Its mutant BLH84T was developed toreduce its potential mitogenicity. The aim of this study was the development of the coculture model system for testing the immunomodulatory effect of chimeras composed ofthe major birch pollen allergen (Bv1a) and BLwt (Bv1a-BLwt, Cwt), the hypoallergenicisoform of Bv1a (Bv1l) and BLH84T (Bv1l-BLH84T, C1 and BLH84T-Bv1l, C2). Chimericstructures were designed in silico, fully minimized, and relaxed without van der Waalsatomic clashes. Afterward, proteins were successfully expressed in Escherichia coli andpurified by IMAC yielding around 0.4 mg per 1L of expression medium. The IgE bindingcapacity was assessed using ELISA inhibition with birch pollen allergic patients’ sera.Caco-2 intestinal epithelial cells and THP-1 differentiated macrophages were used for theco-culture model system development. After protein application on the apical side of theco-culture, the integrity of the epithelial monolayer was not disturbed. Theimmunomodulatory potential of antigens was tested by measuring the gene expressionlevels for pro- and anti-inflammatory cytokines in both cell lines from co-culture. Theobtained results indicate that the best anti-inflammatory response was favored aftertreatment with Cwt. Additionally, to further confirm the immunomodulatory effect of therecombinant chimeras, PBMCs obtained from individuals allergic to birch pollen wereemployed and treated with recombinant prote

Ovalbumin (OVA) is the most abundant egg white protein. It is a globular, acidic phosphorylated glycoprotein of the serpin family with a molecular weight of 45 kDa. OVA is rich in essential amino acids and upon proteolytic digestion yields bioactive peptides (BAPs), recognized nutraceuticals with hypotensive, antimicrobial, antioxidant, and anticancer properties that contribute to the overall nutritional and health benefits of eggs. OVA is a common choice in the food, biomedical, and pharmaceutical industries due to its useful properties during food processing, capacity to form biocompatible gels, and special properties as an effective transporter for a variety of nutraceuticals and pharmaceuticals. Cellular agriculture is an innovative interdisciplinary approach that bypasses conventional animal husbandry in the production of animal proteins. OVA expressed in Trichoderma reesei (T. reesei) most closely mimics the structural and functional properties of its natural homolog and is therefore considered a sustainable alternative to chicken egg white protein powder. Egg allergy poses serious concerns for food safety and an important socioeconomic burden to the food sector and public health. OVA has been extensively studied as an important egg allergen in mice and in vitro experimental models, providing fundamental insights into the molecular mechanisms of allergy and identifying new therapeutic targets. This chapter focuses on providing a comprehensive overview of the state-of-the-art of OVA in human nutrition and the food industry. After presenting the structure underlying the functional properties of OVA, we provide a critical perspective on cellular agriculture as a non-poultry production of OVA. Additionally, the detailed nutritional and biotechnological significance of OVA is elaborated. The final part of this chapter provides a comprehensive insight into OVA as a model antigen and food allergen from a food safety perspective.