Terrestrial carnivorous plants of genera Drosera, Dionaea and Nepenthes within the order Caryophyllales employ jasmonates for the induction of digestive processes in their traps. Here, we focused on two aquatic carnivorous plant genera with different trapping mechanism from distinct families and orders: Aldrovanda (Droseraceae, Caryophyllales) with snap-traps and Utricularia (Lentibulariaceae, Lamiales) with suction traps. Using phytohormone analyses and simple biotest, we asked whether the jasmonates are involved in the activation of carnivorous response similar to that known in traps of terrestrial genera of Droseraceae (Drosera, Dionaea). The results showed that Utricularia, in contrast with Aldrovanda, does not use jasmonates for activation of carnivorous response and is the second genus in Lamiales, which has not co-opted jasmonate signalling for botanical carnivory. On the other hand, the nLC-MS/MS analyses revealed that both genera secreted digestive fluid containing cysteine protease homologous to dionain although the mode of its regulation may differ. Whereas in Utricularia the cysteine protease is present constitutively in digestive fluid, it is induced by prey and exogenous application of jasmonic acid in Aldrovanda.

• MeSH
• Cyclopentanes MeSH
• Droseraceae * MeSH
• Lamiales * MeSH
• Carnivorous Plant MeSH
• Oxylipins MeSH
• Tandem Mass Spectrometry MeSH
• Publication type
• Journal Article MeSH

Background: The rumen microbiota is one of the most complex consortia of anaerobes, involving archaea, bacteria, protozoa, fungi and phages. They are very effective at utilizing plant polysaccharides, especially cellulose and hemicelluloses. The most important hemicellulose decomposers are clustered with the genus Butyrivibrio. As the related species differ in their range of hydrolytic activities and substrate preferences, Butyrivibrio fibrisolvens was selected as one of the most effective isolates and thus suitable for proteomic studies on substrate comparisons in the extracellular fraction. The B. fibrisolvens genome is the biggest in the butyrivibria cluster and is focused on "environmental information processing" and "carbohydrate metabolism". Methods: The study of the effect of carbon source on B. fibrisolvens 3071 was based on cultures grown on four substrates: xylose, glucose, xylan, xylan with 25% glucose. The enzymatic activities were studied by spectrophotometric and zymogram methods. Proteomic study was based on genomics, 2D electrophoresis and nLC/MS (Bruker Daltonics) analysis. Results: Extracellular β-endoxylanase as well as xylan β-xylosidase activities were induced with xylan. The presence of the xylan polymer induced hemicellulolytic enzymes and increased the protein fraction in the interval from 40 to 80 kDa. 2D electrophoresis with nLC/MS analysis of extracellular B. fibrisolvens 3071 proteins found 14 diverse proteins with significantly different expression on the tested substrates. Conclusion: The comparison of four carbon sources resulted in the main significant changes in B. fibrisolvens proteome occurring outside the fibrolytic cluster of proteins. The affected proteins mainly belonged to the glycolysis and protein synthesis cluster.

• Publication type
• Journal Article MeSH

Honey is a unique natural product produced by European honeybees. Due to its high economic value, honey is considered to be well characterized chemically, and it is often discovered to be an adulterated commodity. However, this study shows that our knowledge of honey protein composition, which is of high medical and pharmaceutical importance, is incomplete. In this in-depth proteomic study of 13 honeys, we identified a number of proteins that are important for an understanding of honey properties and merit additional pharmaceutical research. Our major result is an expanded understanding of the proteins underlying honey's antimicrobial properties, such as hymenoptaecin and defensin-1, glucose dehydrogenase isoforms, venom allergens and other venom-like proteins, serine proteases and serine protease inhibitors, and a series of royal jelly proteins. In addition, we performed quantitative comparisons of all of the proteins previously known or newly identified. The honey proteins, determined using label-free nLC-MS/MS in which the same protein quantity was analyzed in one series, were found in relatively similar proportions, although eucalyptus honey differed most widely from the remaining honeys. Overall, the proteome analysis indicated that honeybees supply proteins to honey in a relatively stable ratio within each proteome, but total protein quantity can differ by approximately an order of magnitude in different honeys.

• MeSH
• Allergens analysis   MeSH
• Anti-Bacterial Agents pharmacology   MeSH
• Serine Proteinase Inhibitors analysis   MeSH
• Fatty Acids chemistry   MeSH
• Honey analysis   MeSH
• Proteomics methods   MeSH
• Serine Proteases analysis   MeSH
• Venoms analysis   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Zubní kaz je ve většině vyspělých zemí nejrozšířenější orální nemoc, kterou trpí převážná většina dospělých. Malá část světové populace je vůči tomuto onemocnění zatím z nejasných příčin rezistentní. Jednou z možností tohoto jevu je specifické složení proteinů slin, pelikuly, či zubů u těchto jedinců. Cílem naší studie je porovnat proteinové složení slin, pelikuly a zubů dvou skupin osob (skupina se zubními kazy a skupina bez zubních kazů - z obou skupin okolo deseti vzorků) metodami jednodimenzionální (SDS-PAGE) a dvojdimenzionální gelové elektroforézy (2-DE), nano-kapalinové chromatografie ve spojení s hmotnostním spektrometrem (nLC-MS/MS) a diferenční gelovou elektroforézou (DIGE). Doposud nebyla provedena žádná takováto srovnávací studie zaměřená na proteinové složení lidských zubů. Dále bychom chtěli u těchto dvou skupin vůbec poprvé využít techniku DIGE pro porovnání proteinového složení slin. Dosažené výsledky o proteomu budou porovnány s údaji teoretické studie.; The majority of the worldwide population experiences dental caries. Only a minor part of the population is resistant to this oral disease for unclear reasons. One possibility of this occurrence is specific protein composition of saliva, pellicle, or teeth of these persons. The aim of this study is to compare protein content of saliva, pellicle and teeth from two groups of people: “caries susceptible” (with tooth caries) and “caries resistant” (without any tooth caries) (about ten samples from each group). This comparison of these samples will be done by various methods: gel electrophoresis (one-dimensional (SDS-PAGE) and two-dimensional (2-DE)), difference gel electrophoresis (DIGE), and liquid chromatography coupled with mass spectrometer (nLC-MS/MS). It was not carried out any similar study aimed to protein content of human tooth to this date. In this project we would like to apply the DIGE method to compare the proteom of the saliva (in this case for the very first time).

• MeSH
• Electrophoresis, Gel, Two-Dimensional MeSH
• Chromatography, Liquid MeSH
• Dental Physiological Phenomena MeSH
• Dentin MeSH
• Mass Spectrometry MeSH
• Dental Caries Susceptibility MeSH
• Dental Pellicle MeSH
• Proteome MeSH
• Saliva MeSH
• Dental Caries MeSH

• Conspectus
• Stomatologie
• NML Fields
• zubní lékařství
• biochemie
• NML Publication type
• závěrečné zprávy o řešení grantu IGA MZ ČR

UNLABELLED: Most people in the world suffer from dental caries, >90% of adults experience caries on enamel and root surfaces during their life. However, the overall roles of all factors in the development of dental caries still remain unclear and are worthy of recent investigation. In this study we used a proteomic 2D-DIGE approach in connection with MS/MS to investigate the different protein abundances in the tooth pulp of human third molars obtained from caries-resistant and caries-susceptible people. Statistical analysis of the two protein maps obtained on large gel (17cm length) and mini gel (7cm length) followed by nLC-MS/MS analysis enabled the identification of 16 significantly changed spots with unique protein identifications corresponding to 12 non-redundant proteins. Seven proteins exhibited higher and four proteins exhibited lower expression in the caries-resistant samples compared to the caries-susceptible samples. Additionally, one protein (alpha-1-antitrypsin) exhibited both expressions (up and down). Most of the differentially expressed proteins were associated with protein metabolism, energy production, cytoskeletal organization and transport. These differentially expressed proteins are likely involved in the natural resistance or susceptibility of humans to the development of dental caries and suggest that the resistance mechanism is multifactorial. BIOLOGICAL SIGNIFICANCE: Dental caries are not a serious and life-threatening disease, but their healing requires many remedies and takes up a lot of time. Moreover, neglecting the problem may lead to tooth loss, which can strongly reduce the quality of life. Therefore the identifying effective and safe oral medicine and determining the causes of caries-resistance were viewed as the main aims of this study. Our work aims to elucidate the mechanism of natural human resistance to the development of dental caries by studying the proteomes of tooth pulp isolated from patients who displayed different prevalences of tooth caries. This study is the first protein tooth pulp comparison of sound teeth obtained from caries-resistant versus caries-susceptible people.

• MeSH
• Two-Dimensional Difference Gel Electrophoresis MeSH
• Humans MeSH
• Molar, Third MeSH
• Disease Susceptibility etiology   MeSH
• Disease Resistance MeSH
• Proteome analysis   MeSH
• Proteomics methods   MeSH
• Tandem Mass Spectrometry MeSH
• Dental Pulp chemistry   MeSH
• Dental Caries etiology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Comparative Study MeSH

Tobacco mature pollen has extremely desiccated cytoplasm, and is metabolically quiescent. Upon re-hydration it becomes metabolically active and that results in later emergence of rapidly growing pollen tube. These changes in cytoplasm hydration and metabolic activity are accompanied by protein phosphorylation. In this study, we subjected mature pollen, 5-min-activated pollen, and 30-min-activated pollen to TCA/acetone protein extraction, trypsin digestion and phosphopeptide enrichment by titanium dioxide. The enriched fraction was subjected to nLC-MS/MS. We identified 471 phosphopeptides that carried 432 phosphorylation sites, position of which was exactly matched by mass spectrometry. These 471 phosphopeptides were assigned to 301 phosphoproteins, because some proteins carried more phosphorylation sites. Of the 13 functional groups, the majority of proteins were put into these categories: transcription, protein synthesis, protein destination and storage, and signal transduction. Many proteins were of unknown function, reflecting the fact that male gametophyte contains many specific proteins that have not been fully functionally annotated. The quantitative data highlighted the dynamics of protein phosphorylation during pollen activation; the identified phosphopeptides were divided into seven groups based on the regulatory trends. The major group comprised mature pollen-specific phosphopeptides that were dephosphorylated during pollen activation. Several phosphopeptides representing the same phosphoprotein had different regulation, which pinpointed the complexity of protein phosphorylation and its clear functional context. Collectively, we showed the first phosphoproteomics data on activated pollen where the position of phosphorylation sites was clearly demonstrated and regulatory kinetics was resolved.

• MeSH
• Phosphoproteins chemistry   metabolism   MeSH
• Kinetics MeSH
• Proteomics methods   MeSH
• Pollen metabolism   MeSH
• Gene Expression Regulation, Plant MeSH
• Plant Proteins chemistry   metabolism   MeSH
• Nicotiana genetics   metabolism   MeSH
• Tandem Mass Spectrometry methods   MeSH
• Binding Sites MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

The eggshell is a barrier that plays an important role in the defense of the egg against microbial and other infections; it protects the developing bird against unfavorable impacts of the environment and is essential for the reproduction of birds. The avian eggshell is a complex structure that is formed during movement along the oviduct by producing a multilayered mineral-organic composite. The extractable proteins of avian eggshells have been studied extensively and many of them identified, however, the insoluble (non-extractable) proteins have been sparsely studied. We studied the EDTA-insoluble proteinaceous film from the cuticle layer of eggshell. This film consists of three main areas: spots (cca 300 μm diameter), blotches (small spots with diameter only tens of μm), and the surroundings (i.e., the area without spots and blotches) where spots contain a visible accumulation of pigment. These areas were cut out of the membrane by laser microdissection, proteins were cleavaged by trypsin, and the peptides were analyzed by nLC/MS (Q-TOF). This study has identified 29 proteins and a further eight were determined by less specific "cleavage" with semitrypsin. The relative abundances of these proteins were determined using the exponentially modified protein abundance index (emPAI) where the most dominant proteins were eggshell-specific ones, such as ovocleidin-17 and ovocleidin-116. Individual areas of the cuticle membrane differ in their relative proportions of 14 proteins, where significant differences between the three quantification criteria (direct, after normalization to ovocledin-17, or to ovocledin-116) were observed in four proteins.

• MeSH
• Chromatography, Liquid methods   MeSH
• Mass Spectrometry methods   MeSH
• Chickens MeSH
• Membrane Proteins metabolism   MeSH
• Proteome metabolism   MeSH
• Proteomics methods   MeSH
• Egg Proteins metabolism   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Wheat belongs to six major food allergens inducing IgE-mediated hypersensitivity reaction manifesting as cutaneous, gastrointestinal, and respiratory symptoms. Although cereals are a staple food item in most diets, only a few wheat proteins causing hypersensitivity have been identified. To characterize wheat allergens, salt-soluble wheat extracts were separated by 1-DE and 2-DE and IgE-binding proteins were detected by immunoblotting using sera of patients with allergy to ingested wheat. Proteins, frequently recognized by IgE on 2-DE were analyzed by MALDI-TOF and QTOF and their spectrum was completed by 1-DE and LCQ(DECA) nLC-MS/MS IT technique. Using all three techniques we identified 19 potential wheat allergens such as alpha-amylase inhibitors, beta-amylase, profilin, serpin, beta-D-glucan exohydrolase, and 27K protein. Employing newly developed ELISA, levels of IgE Abs against Sulamit wheat extract and alpha-amylase inhibitors type 1 and 3 were quantified and shown to be significantly elevated in sera of allergic patients compared to those of healthy controls. The level of IgE Abs against alpha-amylase inhibitor type 3 was lower, slightly above the cut-off value in the majority of patients' sera. Our findings contribute to the identification of wheat allergens aimed to increase the specificity of serum IgE and cell activation diagnostic assays.

• MeSH
• Electrophoresis, Gel, Two-Dimensional MeSH
• Allergens immunology   MeSH
• Wheat Hypersensitivity diagnosis   immunology   MeSH
• alpha-Amylases antagonists & inhibitors   MeSH
• Child MeSH
• Adult MeSH
• Electrophoresis, Polyacrylamide Gel MeSH
• Enzyme-Linked Immunosorbent Assay MeSH
• Financing, Organized MeSH
• Immunoblotting MeSH
• Immunoglobulin E blood   MeSH
• Enzyme Inhibitors immunology   metabolism   MeSH
• Humans MeSH
• Adolescent MeSH
• Peptide Fragments analysis   metabolism   MeSH
• Flow Cytometry MeSH
• Triticum immunology   metabolism   adverse effects   MeSH
• Plant Proteins immunology   metabolism   MeSH
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization MeSH
• Check Tag
• Child MeSH
• Adult MeSH
• Humans MeSH
• Adolescent MeSH