Fermentation pattern and yields of microbial protein were investigated in cultures of the rabbit caecal contents supplied with glucose, xylose, starch, pectin and xylan. Rabbits at the age of 4 weeks (before weaning) and 3 months were slaughtered, their caecal contents added at 1.1% to growth media and incubated anaerobically at 39 degrees C for 18 h. Caecal microorganisms of 4-week-old rabbits produced no methane and caproate, less butyrate, but more propionate than microorganisms of 3-month-old rabbits. In both groups of rabbits, fermentation of xylose produced significantly more propionate and less butyrate than fermentation of glucose. More propionate and less acetate was formed from starch than from pectin. In caecal cultures from 4-week-old rabbits with pectin, the molar percentages of acetate was significantly higher and percentages of other short-chain fatty acids (SCFA) lower than in cultures with starch or xylan. In cultures from 3-month-old rabbits, fermentation of pectin and xylan produced similar SCFA profiles, different from SCFA molar composition in cultures with starch. Average production of microbial protein was 129 mg per 1 g of carbohydrate digested (range 110 to 141 mg/g). Protein yields were the same on glucose and xylose, but nonsignificantly higher on starch than on pectin and xylan. It can be concluded that the characteristics of substrate affected fermentation pattern in mixed cultures of rabbit caecal microorganisms. Substrate effects on protein yields were not statistically significant, due to high variation.

• MeSH
• anaerobní bakterie metabolismus   MeSH
• cékum metabolismus   mikrobiologie   MeSH
• fermentace MeSH
• králíci MeSH
• metabolismus sacharidů * MeSH
• proteiny metabolismus   MeSH
• sacharidy aplikace a dávkování   MeSH
• substrátová specifita MeSH
• techniky in vitro MeSH
• věkové faktory MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• proteiny MeSH
• sacharidy MeSH

Collagen, myosin and albumin were incubated for 7 days at 20 degrees C with fructose, ribose or glyceraldehyde. For thus-formed glycated proteins, quantities were determined by the Conway microdiffusion technique and by the colorimetric method based on Coomassie brilliant blue G-250 colour yield. It was found that when albumin was glycated with increasing amounts of glyceraldehyde, the colour yield was decreased by 7-33%. In collagen, myosin and albumin incubated with 0.5 mol/l fructose, 0.5 mol/l ribose or 0.1 mol/l glyceraldehyde, protein concentration was not changed, as proved by the Conway microdiffusion technique; the Coomassie brilliant blue G-250 colour yield was up to 50% lower, depending on the protein used, and was decreased much less when proteins were incubated with less sugar.

• MeSH
• albuminy analýza   chemie   MeSH
• fruktosa chemie   MeSH
• glyceraldehyd chemie   MeSH
• glykoproteiny analýza   MeSH
• kolagen analýza   chemie   MeSH
• kolorimetrie MeSH
• myosiny analýza   chemie   MeSH
• ribosa chemie   MeSH
• rosanilinová barviva * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• albuminy MeSH
• coomassie Brilliant Blue MeSH Prohlížeč
• fruktosa MeSH
• glyceraldehyd MeSH
• glykoproteiny MeSH
• kolagen MeSH
• myosiny MeSH
• ribosa MeSH
• rosanilinová barviva * MeSH

Our previous studies have shown physiological and yield intensification of selected crops with the application of nanoparticles (NPs). However, the impact on the quantitative, qualitative, and yield parameters of maize (Zea mays L.) in field conditions remains highly debated. This study aimed to evaluate the effects of zinc oxide (ZnO-NPs), gold NPs anchored to meso-biosilica (Au-NP-bioSi), and titanium dioxide (TiO2-NPs) as biological stimulants under field conditions during the vegetation season of 2021 in the Central European region. The study assessed the effects on the number of plants, yield, yield components, and nutritional quality, including mineral nutrients, starch, and crude protein levels. The potential translocation of these chemically-physically stable NPs, which could pose a hazard, was also investigated. The results indicate that Au-NP-bioSi and ZnO-NPs-treatments were the most beneficial for yield and yield components at a statistically significant level. Mineral nutrient outcomes were varied, with the NP-free variant performing the best for phosphorus-levels, while Au-NP-bioSi and ZnO-NPs were optimal for crude protein. Starch content was comparable across the TiO2-NPs, Au-NP-bioSi, and control variants. Importantly, we observed no hazardous translocation of NPs or negative impacts on maize grain quality. This supports the hypothesis that NPs can serve as an effective tool for precise and sustainable agriculture.

• Klíčová slova
• biosilica, gold, grain quality, maize, nanofertilizers, nanoparticles, spray application, titanium dioxide, yield, zinc oxide,
• Publikační typ
• časopisecké články MeSH

Expression systems based on high selectivity and activity of T7 RNA polymerase and presence of a strong T7 promoter have been commonly used for cloning and expression of various recombinant proteins in Escherichia coli. When the expression system is designed in such a way that the produced protein is not being transferred into periplasm, bacterial cells must be lysed in order to isolate and purify the protein. The final yield and quality of the synthesized protein then depend on various factors, protein size, amino acid sequence, solubility in cytoplasm, and folding requirements among them. The yield in the T7 RNA polymerase/promoter system can be positively influenced by use of rifampicin. In this report we demonstrate usefulness of the antibiotic in detail. We describe rifampicin-enhanced expression of a plant cytokinin-specific beta-glucosidase. Two bacterial cultures are compared, one expressing the enzyme without and one in the presence of rifampicin. The antibiotic not only increased the yield of the recombinant protein, which seems to be a general phenomenon, but also favored the final assembly of the protein's subunits into a catalytically active dimer form.

• MeSH
• beta-glukosidasa chemie   imunologie   metabolismus   MeSH
• cytokininy chemie   MeSH
• dimerizace MeSH
• DNA řízené RNA-polymerasy metabolismus   MeSH
• Escherichia coli enzymologie   MeSH
• konformace proteinů MeSH
• rekombinantní proteiny chemie   metabolismus   MeSH
• rifampin farmakologie   MeSH
• sbalování proteinů MeSH
• virové proteiny MeSH
• western blotting MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bacteriophage T7 RNA polymerase MeSH Prohlížeč
• beta-glukosidasa MeSH
• cytokinin-beta-glucosidase MeSH Prohlížeč
• cytokininy MeSH
• DNA řízené RNA-polymerasy MeSH
• rekombinantní proteiny MeSH
• rifampin MeSH
• virové proteiny MeSH

Intercropping is an alternative farming method that maximizes crop yield and resource usage effectiveness, especially in low-input agricultural systems. Legume-based intercropping systems can effectively boost the quality and wheat yield by promoting soil functions and microbial activities. However, changes in the types of legumes and field management can alter the response of crop functions. A three-year field study was conducted on intercropping cultivation of winter wheat variety (Butterfly and Lorien) and legume species (faba bean, incarnate clover, spring pea, winter pea) to assess grain yield and wheat quality in organic farming. Based on the results, Butterfly showed higher grain quality but lower grain yield and yield components than Lorien. Mixtures of legume crops with winter wheat did not significantly differ in wheat grain yield, but grain quality variables were significantly affected. Protein content (PC) was significantly higher in wheat and legume mixtures than in sole wheat by 4 %. PC in wheat + winter pea (Wheat + Wi) and wheat + faba bean (Wheat + Fa) were higher than wheat sown alone. Wet gluten (WG) was higher in Wheat + Wi than in sole wheat and wheat + incarnate clover mixtures (Wheat + In). The rheological parameters evaluated by the Mixolab showed greater wheat quality in Butterfly and legume mixtures. Mixed and row-row intercropping of wheat and legume species did not significantly influence rheological properties. To conclude, customizing wheat yield and grain quality under the effect of winter wheat and legume mixtures requires considering the optimal solution based on different cultivates, wheat varieties and legume species to achieve the desired response.

• Klíčová slova
• Baking quality, Crop productivity, Mixolab, Rheological properties, Wheat yield,
• Publikační typ
• časopisecké články MeSH

Engineered small non-antibody protein scaffolds are a promising alternative to antibodies and are especially attractive for use in protein therapeutics and diagnostics. The advantages include smaller size and a more robust, single-domain structural framework with a defined binding surface amenable to mutation. This calls for a more systematic approach in designing new scaffolds suitable for use in one or more methods of directed evolution. We hereby describe a process based on an analysis of protein structures from the Protein Data Bank and their experimental examination. The candidate protein scaffolds were subjected to a thorough screening including computational evaluation of the mutability, and experimental determination of their expression yield in E. coli, solubility, and thermostability. In the next step, we examined several variants of the candidate scaffolds including their wild types and alanine mutants. We proved the applicability of this systematic procedure by selecting a monomeric single-domain human protein with a fold different from previously known scaffolds. The newly developed scaffold, called ProBi (Protein Binder), contains two independently mutable surface patches. We demonstrated its functionality by training it as a binder against human interleukin-10, a medically important cytokine. The procedure yielded scaffold-related variants with nanomolar affinity.

• Klíčová slova
• computational saturation, directed evolution, interleukin-10, protein engineering, protein scaffold, ribosome display,
• MeSH
• databáze proteinů MeSH
• interleukin-10 metabolismus   MeSH
• konformace proteinů MeSH
• počítačová simulace MeSH
• proteinové inženýrství MeSH
• proteiny chemie   genetika   metabolismus   MeSH
• rekombinantní proteiny chemie   genetika   metabolismus   MeSH
• ribozomy metabolismus   MeSH
• řízená evoluce molekul metody   MeSH
• sekvence aminokyselin MeSH
• stabilita proteinů MeSH
• vazba proteinů MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• interleukin-10 MeSH
• proteiny MeSH
• rekombinantní proteiny MeSH

Protein solubility is an attractive engineering target primarily due to its relation to yields in protein production and manufacturing. Moreover, better knowledge of the mutational effects on protein solubility could connect several serious human diseases with protein aggregation. However, we have limited understanding of the protein structural determinants of solubility, and the available data have mostly been scattered in the literature. Here, we present SoluProtMutDB - the first database containing data on protein solubility changes upon mutations. Our database accommodates 33 000 measurements of 17 000 protein variants in 103 different proteins. The database can serve as an essential source of information for the researchers designing improved protein variants or those developing machine learning tools to predict the effects of mutations on solubility. The database comprises all the previously published solubility datasets and thousands of new data points from recent publications, including deep mutational scanning experiments. Moreover, it features many available experimental conditions known to affect protein solubility. The datasets have been manually curated with substantial corrections, improving suitability for machine learning applications. The database is available at loschmidt.chemi.muni.cz/soluprotmutdb.

• Klíčová slova
• Machine learning, Mutational database, Protein aggregation, Protein engineering, Protein yield, Soluble expression,
• Publikační typ
• časopisecké články MeSH

Macroautophagy/autophagy is a catabolic process by which cytosolic content is engulfed, degraded and recycled. It has been implicated as a critical pathway in advanced stages of cancer, as it maintains tumor cell homeostasis and continuous growth by nourishing hypoxic or nutrient-starved tumors. Autophagy also supports alternative cellular trafficking pathways, providing a mechanism of non-canonical secretion of inflammatory cytokines. This opens a significant therapeutic opportunity for using autophagy inhibitors in cancer and acute inflammatory responses. Here we developed a high throughput compound screen to identify inhibitors of protein-protein interaction (PPI) in autophagy, based on the protein-fragment complementation assay (PCA). We chose to target the ATG12-ATG3 PPI, as this interaction is indispensable for autophagosome formation, and the analyzed structure of the interaction interface predicts that it may be amenable to inhibition by small molecules. We screened 41,161 compounds yielding 17 compounds that effectively inhibit the ATG12-ATG3 interaction in the PCA platform, and which were subsequently filtered by their ability to inhibit autophagosome formation in viable cells. We describe a lead compound (#189) that inhibited GFP-fused MAP1LC3B/LC3B (microtubule associated protein 1 light chain 3 beta) puncta formation in cells with IC50 value corresponding to 9.3 μM. This compound displayed a selective inhibitory effect on the growth of autophagy addicted tumor cells and inhibited secretion of IL1B/IL-1β (interleukin 1 beta) by macrophage-like cells. Compound 189 has the potential to be developed into a therapeutic drug and its discovery documents the power of targeting PPIs for acquiring specific and selective compound inhibitors of autophagy.Abbreviations: ANOVA: analysis of variance; ATG: autophagy related; CQ: chloroquine; GFP: green fluorescent protein; GLuc: Gaussia Luciferase; HEK: human embryonic kidney; IL1B: interleukin 1 beta; LPS: lipopolysaccharide; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; PCA: protein-fragment complementation assay; PDAC: pancreatic ductal adenocarcinoma; PMA: phorbol 12-myristate 13-acetate; PPI: protein-protein interaction. VCL: vinculin.

• Klíčová slova
• Autophagy inhibition, LC3B, cancer, drug screen, pancreatic cancer, protein-fragment complementation assay, small molecules,
• MeSH
• Atg12 MeSH
• autofagie * MeSH
• interleukin-1beta farmakologie   MeSH
• lidé MeSH
• nádory slinivky břišní * MeSH
• proteiny asociované s mikrotubuly metabolismus   MeSH
• proteiny spojené s autofagií MeSH
• ubikvitin konjugující enzymy metabolismus   MeSH
• zelené fluorescenční proteiny metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ATG12 protein, human MeSH Prohlížeč
• Atg12 MeSH
• ATG3 protein, human MeSH Prohlížeč
• interleukin-1beta MeSH
• proteiny asociované s mikrotubuly MeSH
• proteiny spojené s autofagií MeSH
• ubikvitin konjugující enzymy MeSH
• zelené fluorescenční proteiny MeSH

Mitochondrial uncoupling protein (UcP) contains two tryptophans buried in transmembrane alpha-helices: Trp-173 at the matrix end of fourth alpha-helix and Trp-280 on the sixth alpha-helix. However, the steady-state emission of isolated UcP exhibited properties unusual for alpha-helices: maximum close to that of free tryptophan emission and low quantum yield of 0.04. The former suggests prevailing tryptophan contacts with hydrophilic residues and confirms Trp-173 proximity to the water/membrane interface and Trp-280 location near a water-filled nucleotide-binding-site cavity. The latter might indicate that transmembrane segments are not true alpha-helices. Measured depolarization factor of 0.6 suggests also their "breathing". Analysis of UcP emission decays, measured by time-correlated-single-photon-counting, yielded components 0.4-0.6 ns, 2.2-3 ns and 9-10 ns (or alternatively 0.1, 1.5, 4.3 and 12.2 ns; or 0.1-0.3, 1.2, 3.7 and 10.5 ns), very similar to those of free tryptophan in water, where the longest component belongs to anionic form. Hence, such an "anionic" conformation must exist in UcP, perhaps as a consequence of charge-transfer complexes between Trp-173 & Lys-174 and Trp-280 & Arg-276. Moreover, N-ethylmaleimide modification, known to induce conformational changes, prolonged the "10 ns" component, decreased quantum yield to 0.03 without changes in emission spectra, while slightly shifting absorption to red and increasing tyrosine exposure to water.

• MeSH
• biofyzika MeSH
• biofyzikální jevy MeSH
• ethylmaleimid farmakologie   MeSH
• fluorescenční polarizace MeSH
• fluorescenční spektrometrie MeSH
• hnědá tuková tkáň chemie   MeSH
• iontové kanály MeSH
• konformace proteinů účinky léků   MeSH
• křečci praví MeSH
• křeček rodu Mesocricetus MeSH
• membránové proteiny chemie   MeSH
• mitochondriální proteiny MeSH
• mitochondrie chemie   MeSH
• molekulární modely MeSH
• molekulární sekvence - údaje MeSH
• rozpřahující látky chemie   MeSH
• sekundární struktura proteinů MeSH
• sekvence aminokyselin MeSH
• techniky in vitro MeSH
• transportní proteiny chemie   MeSH
• tryptofan chemie   MeSH
• uncoupling protein 1 MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ethylmaleimid MeSH
• iontové kanály MeSH
• membránové proteiny MeSH
• mitochondriální proteiny MeSH
• rozpřahující látky MeSH
• transportní proteiny MeSH
• tryptofan MeSH
• uncoupling protein 1 MeSH

INTRODUCTION: Soybean is an important legume crop and a leading source of dietary protein and oil in animal feed, as well as an important food for human consumption. The objective of our research was to study soybean genetic resources in context of future protein self-sufficiency both in human and animal nutrition. METHODS: Collection of 360 different accessions from various regions worldwide was evaluated across four European locations during two consecutive years in phenotyping trials. The five most important traits of soybean - plant emergence, plant length, protein content, seed yield, and R8 stage - were carefully analysed, revealing significant variability. RESULTS: Ten exceptionally stable genotypes were identified based on their protein content and yield, presenting promising candidates for breeding programs. DISCUSSION: Our findings underscore the importance of integrating genotype-environment interaction analyses into breeding initiatives, considering the observed variability in phenotypic traits across diverse environments and genotypes.

• Klíčová slova
• Glycine max, field trials, genotype-environment interactions, morphological and phenological traits, weather data,
• Publikační typ
• časopisecké články MeSH