Supplementary data for article: Grozdanović, M. M.; Burazer, L. M.; Gavrović-Jankulović, M. Kiwifruit (Actinidia Deliciosa) Extract Shows Potential as a Low-Cost and Efficient Milk-Clotting Agent. International Dairy Journal 2013, 32 (1), 46–52. https://doi.org/10.1016/j.idairyj.2013.03.001

Supplementary material for: [https://doi.org/10.1016/j.idairyj.2013.03.001]Related to published version: [http://cherry.chem.bg.ac.rs/handle/123456789/1589

Supplementary data for article: Grozdanović, M. M.; Burazer, L. M.; Gavrović-Jankulović, M. Kiwifruit (Actinidia Deliciosa) Extract Shows Potential as a Low-Cost and Efficient Milk-Clotting Agent. International Dairy Journal 2013, 32 (1), 46–52. https://doi.org/10.1016/j.idairyj.2013.03.001

Supplementary material for: [https://doi.org/10.1016/j.idairyj.2013.03.001]Related to published version: [http://cherry.chem.bg.ac.rs/handle/123456789/1589

Evaluacija kriterijuma za dijagnozu atopijskog dermatitisa i detekcija alergen specifičnih IgE antitela kod pasa alergičnih na polen biljke Ambrosia artemisiifolia

Common ragweed (Ambrosia atremisiifolia) is one of the most frequent causes of pollen-induced allergic reactions both in humans and dogs. It has not been defined yet, what is the major allergen(s) to which most dogs allergic to ragweed show a positive result on intradermal skin test (IDST). In the present study sensitization to Ambrosia artemisiifolia pollen allergens in dogs with atopic dermatitis was examined with both in vivo and in vitro tests, including IDST and serum allergen specific IgE test. Detection of specific-IgE antibodies against ragweed allergens by immunoblotting in the sera of allergic dogs was optimized, as well. Dogs that were positive, as judged by IDST reactions to ragweed pollen allergens, also had alergen specific IgE antibodies in their sera. Results indicate that major allergens of A. artemisifolia pollen in dogs are Amb a 1 and Amb a 2. Further characterization of ragweed allergens is needed before they could potentially be used in intradermal testing or allergen immunotherapy in affected dogs. Also, we evaluated new Favrots diagnostic criteria for canine atopic dermatitis in dogs allergic to Ambrosia atremisiifolia pollen. It might be concluded that proposed criteria are of great assistance for seting up suspected diagnosis of canine atopic dermatitis, after ruling out other pruritic dermatoses.Kratka ambrozija (Ambrosia artemisiifolia) je jedan od najčešćih uzročnika alergijskih reakcija izazvanih polenom kod ljudi i pasa. Još uvek nije definisano koji je glavni alergen (i), na koji, većina pasa alergičnih na polen ambrozije, ispoljava pozitivnu reakciju na intradermalnom testu. U ovoj studiji je ispitana senzibilizacija na polen ove biljke kod pasa sa simptomima atopijskog dermatitisa in vivo i in vitro testovima, uključujući intradermalni test i dokazivanje prisustva alergen specifičnih IgE antitela u serumu. Optimizovani su uslovi za detekciju IgE specifičnih antitela iz seruma pasa alergičnih na polen ambrozije imunoblot tehnikom. Psi koji su imali pozitivnu reakciju na polen ove biljke na intradermalnom testu, takođe su imali specifična IgE antitela u serumu. Dobijeni rezultati ukazuju da su glavni alergeni Ambrosia artemisiifolia kod pasa Amb a 1 i Amb a 2. Neophodna je dalja karakterizacija alergena ambrozije kako bi se oni mogli primeniti pri rutinskom intradermalnom testiranju ili u alergen specifičnoj imunoterapiji obolelih pasa. Takođe je razmatrana i validnost Favrotovih dijagnostičkih kriterijuma kod pasa alergičnih na polen ambrozije. Može se zaključiti da su predloženi kriterijumi od velike pomoći u postavljanju suspektne dijagnoze atopijskog dermatitisa pasa, nakon isključenja drugih pruritičnih dermatoza

Highly improved method for in-depth post-translational modification profiling: example of Timothy grass (Phleum pratense) pollen proteomes from polluted and preserved environments

Field-realistic exposure studies provide the most relevant assessment of the effects of the intensity and diversity of urban and industrial contamination on pollen structure and allergenicity. The significance of in-depth post-translational modification (PTM) studies of pollen proteomes, when compared with studies on other aspects of pollution and altered pollen allergenicity, has not yet been determined; hence, little progress has been made within this field

Izolovanje i karakterizacija 68 kD alergena iz ekstrakta kućnih grinja

House dust mites (HDM) represent a major source of allergens, contributing to the increasing incidence of type I hypersensitivity disease worldwide. Over 30 different IgE-binding proteins from the HDM extract were detected. Although group 1 and 2 have been identified as major allergens, due to the safety and efficacy of allergy diagnosis and immunotherapy, there is a need to carefully evaluate the clinical relevance of other allergens present in the HDM extract. In regard to this, a high molecular mass allergen of about 68 kD was purified from the HDM extract using a combination of gel permeation chromatography and reversed-phase chromatography. The IgG and IgE reactivity of the purified protein were preserved during the purification process, as confirmed by Western blot analysis with polyclonal rabbit antibodies and dot blot analysis with a pool of sera from subjects with house dust mite allergy, respectively. In addition, the IgE reactivity was confirmed using ELISA testing with nine patient sera. The biological potency of the 68 kD allergen was confirmed by skin prick testing in five allergic subjects, suggesting that the high molecular mass allergen is a good candidate for component-resolved diagnosis of house dust mite allergy and eventual therapeutic treatment.Grinje iz kućne prašine predstavljaju jedan od glavnih izvora alergena koji su u značajnoj meri doprineli porastu prvog tipa preosetljivosti. Preko 30 IgE-vezujućih proteina iz kućne prašine je detektovano do danas. Alergeni grupe 1 i 2 označeni su kao glavni alergeni kućne prašine. Međutim, da bi se poboljšala sigurnost i efikasnost dijagnoze i terapije alergijskih oboljenja izazvanih grinjama iz kućne prašine, neophodno je odrediti klinički značaj svih alergena iz ovog alergenskog izvora. U ovom radu izolovan je alergen visoke molekulske mase od 68 kD iz ekstrakta kućne prašine kombinovanjem gel-permeacione hromatografije i reversno-fazne hromatografije. IgG i IgE reaktivnost prečišćenog proteina je proverena u 'Western blot'-u i 'dot blot'-u sa poliklonskim zečijim antitelima na ekstrakt kućne prašine i 'pool'-om seruma osoba alergičnih na kućnu prašinu, redom. 64 % pacijenata je pokazalo IgE reaktivnost na prečišćeni protein u ELISA testu. Biološka reaktivnost prečišćenog alergena je potvrđena u kožnim probama na pet pacijenata, ukazujući da je prečišćen alergen dobar kandidat za dijagnozu alergije na kućnu prašinu pojedinačnim komponentama i eventualni terapeutski tretman

Supplementary data for article: Smiljanic, K.; Apostolovic, D.; Trifunovic, S.; Ognjenovic, J.; Perusko, M.; Mihajlovic, L.; Burazer, L.; van Hage, M.; Cirkovic Velickovic, T. Subpollen Particles Are Rich Carriers of Major Short Ragweed Allergens and NADH Dehydrogenases: Quantitative Proteomic and Allergomic Study. Clinical and Experimental Allergy 2017, 47 (6), 815–828. https://doi.org/10.1111/cea.12874

Supporting information for: [https://doi.org/10.1111/cea.12874]Related to published version: [http://cherry.chem.bg.ac.rs/handle/123456789/2468]Related to accepted version: [http://cherry.chem.bg.ac.rs/handle/123456789/3126

Deep and quantitative profiling of PTMs in ecologically preserved and polluted pollen proteomes of timothy grass reveals predominant source of contamination

Objective: to create method for unrestrictive deep, relative quantification of post translational modifications (PTMs) within different proteomes. Pollution field studies of bio indicators such as pollen are valuable because of realistic situation of target contamination, however they carry the great extent of uncertainty in attributing and delineating the polluting effect from multiple sources. Holistic research platform focusing on comprehensively characterized and quantified PTMs of comparable bio-indicator proteomes could help and overcome these obstacles of field pollution studies. Material and Methods: Scanning electron and light microscopy assessed surface and sub pollen particle (SPP) releasing features of timothy grass (TG) pollen. Inductively coupled atomic emission spectrometry revealed metal elemental content in pollen while in solution trypsin digested pollen proteomes analysed with high resolution Orbitrap mass tandem spectrometry and PEAKS Suite 8.5 brought quantitative information on protein expression level and its PTM profiling. Results: TG polluted pollen samples (P2) collected along regional road and chemical plant, exposed to air contaminants from road traffics and chemical plants showed 4.5 times higher SPP releasing capacity, with notable surface changes, as well as significantly higher contents of Mn, Hg and Cd. Antioxidative enzymes (oxidoreductases, superoxide dismutases and peroxidases), including actin, were upregulated several times in polluted sample compared to ecologically preserved pollen (P1). While the level of spontaneous and physiological PTMs including methylation, acetylation, deamidation and formylation, was similar without significant changes in P1 and P2 pollens, oxidative PTMs including oxidation of Met, Lys, His, Pro and HNE and hexose adducts showed several times higher and significant increase in abundancy of P2 compared to P1. PTMs connected to road traffic such as tyrosine nitration were very rare and low abundant. Conclusion: Results suggest prominent role of chemical pollution compared to effect of road traffic pollution, with primary consequences from oxidative properties of mercury (Hg) and cadmium (Cd).Book of Abstract

Unrestricted and quantitative method of post translational modifications profiling: Timothy grass pollen proteome in relation to increased oxidative stress caused by enviromental pollution

The significance of in-depth post-translational modification (PTM) studies of pollen proteomes, when compared with studies on other aspects of pollution and altered pollen allergenicity, has not yet been determined; hence, little progress has been made within this field. Therefore, we created a comprehensive approach for the comparison of pollen from polluted and environmentally preserved areas. To examine the effects of long-term, in vivo pollen exposure to multiple source pollutants, Phleum pratense (Timothy grass) pollen samples were collected along a regional road in Kruševac, central Serbia and were compared with pollen samples from rural, environmentally preserved area over two consecutive pollination seasons. We combined the quantitative comparison of proteome expression profiles from in-solution and 2D-gels with unrestrictive in-depth quantitative PTM profiling using high resolution tandem mass spectrometry and the PEAKS 8.5 Suite platform. An increased phenolic content and release of sub-pollen particles was found in pollen samples from the polluted area, and significantly higher content of mercury, cadmium, and manganese. Antioxidative defense-related enzymes were significantly upregulated. Seven oxidative PTMs were significantly increased (methionine, histidine, lysine, and proline oxidation; tyrosine glycosylation, lysine 4-hydroxy-2-nonenal adduct, and lysine carbamylation) in pollen exposed to the chemical plant and road traffic pollution sources. Oxidative modifications affected pollen allergens, especially Phl p 6, with several different oxidative modifications. Quantitative, unrestricted, and detailed PTM searches using an enrichment-free approach was used for the first time to map extensive modifications in the pollen proteome, reflecting increased environmental oxidative stress, primarily caused by increased content of heavy metals in pollen.Acknowledgement to European Commission, under the Horizon2020, FoodEnTwin project

Unrestricted and quantitative method of post translational modifications profiling: Timothy grass pollen proteome in relation to increased oxidative stress caused by enviromental pollution

The significance of in-depth post-translational modification (PTM) studies of pollen proteomes, when compared with studies on other aspects of pollution and altered pollen allergenicity, has not yet been determined; hence, little progress has been made within this field. Therefore, we created a comprehensive approach for the comparison of pollen from polluted and environmentally preserved areas. To examine the effects of long-term, in vivo pollen exposure to multiple source pollutants, Phleum pratense (Timothy grass) pollen samples were collected along a regional road in Kruševac, central Serbia and were compared with pollen samples from rural, environmentally preserved area over two consecutive pollination seasons. We combined the quantitative comparison of proteome expression profiles from in-solution and 2D-gels with unrestrictive in-depth quantitative PTM profiling using high resolution tandem mass spectrometry and the PEAKS 8.5 Suite platform. An increased phenolic content and release of sub-pollen particles was found in pollen samples from the polluted area, and significantly higher content of mercury, cadmium, and manganese. Antioxidative defense-related enzymes were significantly upregulated. Seven oxidative PTMs were significantly increased (methionine, histidine, lysine, and proline oxidation; tyrosine glycosylation, lysine 4-hydroxy-2-nonenal adduct, and lysine carbamylation) in pollen exposed to the chemical plant and road traffic pollution sources. Oxidative modifications affected pollen allergens, especially Phl p 6, with several different oxidative modifications. Quantitative, unrestricted, and detailed PTM searches using an enrichment-free approach was used for the first time to map extensive modifications in the pollen proteome, reflecting increased environmental oxidative stress, primarily caused by increased content of heavy metals in pollen.Acknowledgement to European Commission, under the Horizon2020, FoodEnTwin project

Digestomics of cow's milk: casein-derived digestion-resistant peptides aggregate into functional complexes

This paper discusses the improvements to flight simulators and their use in flight training