Model syntézy enzýmu podľa Goodwina (1965) je autorom tohoto prekladaného príspevku rozšírený v dvoch bodoch (obr. 1): a) Génový kompartment sa definoval ako dynamický blok. b) Pre transport aktivačného represora z ribozómu do bunkového jadra sa navrhlo časové oneskorenie. Použitím Perturbačnej analýzy (Pertubation Analysis) ako aj Ljapunovho teorému prvej aproximácie (aby sa linearizoval systém) a Nyquistovho kritéria stability sa dokázalo, že rovnovážny stav sa vyznačuje lokálnou asymptotickou stabilitou, bez ohľadu na to, aké sú parametre systému. Bola tak preukázala zaujímavá skutočnosť. Že v živej prírode sa vyvinul dynamický systém s robustnou stabilitou. Následne sa použilo kruhové kriterium (Circle Criterion) ako veľmi slabá verzia testu pre stabilitu nelineárneho systému, aby sa určili podmieky pre globálnu asymptotická stabilitu. Výsledky sú veľmi presvedčivé. Simulačně experimenty potvrdili globálnu stabilitu, a to pri všetkých podmienkach, pri ktorých sa urobiti simulácie. Systém má teda vlastnosti, ktoré naznačujú tzv. hyperstabilitu nelineárneho systému.

A model for enzyme synthesis, due to Goodwin (1965), is extended in two ways: dynamics are assigned to th the gene compartment and time delay is introduced for transport of activated repressor from ribosome to cell nucleus. Using Perturbation Analysis, Lyapunov s theorem of the First Approximation to linearize the system and the Nyquist Stability Criterion, it is shown that the equilibrium state has at least local asymptotic stability, irrespective of the values of the system parameters. It seems to be a very marvellous circumstance that nature has designed a system with such a robust stability property. A very weak version of a nonlinear stability property test, the Circle Criterion, is then invoked to establish a sufficient condition for global asymptotic stability. This result is certainly very conservative: simulation results point to global asymptotic stability under all circumstances.

• MeSH
• enzymy biosyntéza   MeSH
• lidé MeSH
• stabilita enzymů MeSH
• teoretické modely MeSH
• Check Tag
• lidé MeSH

• Konspekt
• Geriatrie
• NLK Obory
• geriatrie
• biologie

• Klíčová slova
• RNA,
• MeSH
• messenger RNA metabolismus   MeSH
• regulace genové exprese MeSH
• stabilita RNA MeSH
• Publikační typ
• monografie MeSH

• Konspekt
• Biologické vědy
• NLK Obory
• biologie

• MeSH
• biologická terapie MeSH
• buněčná a tkáňová terapie MeSH
• genetická terapie MeSH
• imunoterapie MeSH
• klinické lékařství MeSH
• Publikační typ
• abstrakty MeSH
• kongresy MeSH
• programy MeSH

• Konspekt
• Patologie. Klinická medicína
• NLK Obory
• biomedicínské inženýrství
• genetika, lékařská genetika
• biologie
• alergologie a imunologie

Nitrilases are highly conserved proteins with catabolic activity but much less understood functions in cell division and apoptosis. To elucidate the biological functions of Arabidopsis NITRILASE1, we characterized its molecular forms, cellular localization and involvement in cell proliferation and plant development. We performed biochemical and mass spectrometry analyses of NITRILASE1 complexes, electron microscopy of nitrilase polymers, imaging of developmental and cellular distribution, silencing and overexpression of nitrilases to study their functions. We found that NITRILASE1 has an intrinsic ability to form filaments. GFP-NITRILASE1 was abundant in proliferating cells, distributed in cytoplasm, in the perinuclear area and associated with microtubules. As cells exited proliferation and entered differentiation, GFP-NITRILASE1 became predominantly nuclear. Nitrilase silencing dose-dependently compromised plant growth, led to loss of tissue organization and sustained proliferation. Cytokinesis was frequently aborted, leading to enlarged polyploid cells. In reverse, independently transformed cell lines overexpressing GFP-NITRILASE1 showed slow growth and increased rate of programmed cell death. Altogether, our data suggest that NITRILASE1 homologues regulate the exit from cell cycle and entry into differentiation and simultaneously are required for cytokinesis. These functions are essential to maintain normal ploidy, genome stability and tissue organization.

• MeSH
• aminohydrolasy chemie   genetika   metabolismus   ultrastruktura   MeSH
• Arabidopsis cytologie   genetika   růst a vývoj   MeSH
• buněčná diferenciace genetika   MeSH
• buněčná smrt genetika   MeSH
• buněčný cyklus genetika   MeSH
• cytoplazma metabolismus   MeSH
• cytoskelet genetika   metabolismus   MeSH
• hydrolasy působící na anhydridy kyselin genetika   MeSH
• nádorové proteiny genetika   MeSH
• nestabilita genomu * MeSH
• proliferace buněk MeSH
• regulace genové exprese u rostlin MeSH
• RNA interference MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The determination of a suitable buffer environment for a protein of interest is not an easy task. The requirements of advanced techniques, the demands on the biological material and the researcher time needed for buffer optimization, as well as personal inflexibility, lead frequently to the use of sub-optimal buffers. Here, we demonstrate the design of a 48-condition buffer screen that can be used to determine an appropriate environment for downstream studies. By the combination of several techniques (differential scanning fluorimetry, dynamic light scattering, and bio-layer interferometry), we are able to assess the protein stability, homogeneity and binding activity across the screen with less than half a milligram of protein in 1 day. The application of this screen helps to avoid unsuitable conditions, to explain problems observed upon protein analysis and to choose the most suitable buffers for further research. The screen can be routinely used as a primary screen for buffer optimization in labs and facilities.

• MeSH
• dynamický rozptyl světla MeSH
• fluorometrie MeSH
• proteiny MeSH
• pufry MeSH
• stabilita proteinů * MeSH
• Publikační typ
• časopisecké články MeSH

The ribosome, owing to its exceptional conservation, harbours a remarkable molecular fossil known as the protoribosome. It surrounds the peptidyl transferase center (PTC), responsible for peptide bond formation. While previous studies have demonstrated the PTC activity in RNA alone, our investigation reveals the intricate roles of the ribosomal protein fragments (rPeptides) within the ribosomal core. This research highlights the significance of rPeptides in stability and coacervation of two distinct protoribosomal evolutionary stages. The 617nt 'big' protoribosome model, which associates with rPeptides specifically, exhibits a structurally defined and rigid nature, further stabilized by the peptides. In contrast, the 136nt 'small' model, previously linked to peptidyltransferase activity, displays greater structural flexibility. While this construct interacts with rPeptides with lower specificity, they induce coacervation of the 'small' protoribosome across a wide concentration range, which is concomitantly dependent on the RNA sequence and structure. Moreover, these conditions protect RNA from degradation. This phenomenon suggests a significant evolutionary advantage in the RNA-protein interaction at the early stages of ribosome evolution. The distinct properties of the two protoribosomal stages suggest that rPeptides initially provided compartmentalization and prevented RNA degradation, preceding the emergence of specific RNA-protein interactions crucial for the ribosomal structural integrity.

• MeSH
• konformace nukleové kyseliny MeSH
• molekulární modely MeSH
• peptidy chemie   metabolismus   MeSH
• peptidyltransferasy metabolismus   chemie   MeSH
• ribozomální proteiny * metabolismus   chemie   MeSH
• ribozomy * metabolismus   MeSH
• RNA metabolismus   chemie   MeSH
• stabilita RNA MeSH
• Publikační typ
• časopisecké články MeSH

Haloalkane dehalogenases are microbial enzymes with a wide range of biotechnological applications, including biocatalysis. The use of organic co-solvents to solubilize their hydrophobic substrates is often necessary. In order to choose the most compatible co-solvent, the effects of 14 co-solvents on activity, stability and enantioselectivity of three model enzymes, DbjA, DhaA, and LinB, were evaluated. All co-solvents caused at high concentration loss of activity and conformational changes. The highest inactivation was induced by tetrahydrofuran, while more hydrophilic co-solvents, such as ethylene glycol and dimethyl sulfoxide, were better tolerated. The effects of co-solvents at low concentration were different for each enzyme-solvent pair. An increase in DbjA activity was induced by the majority of organic co-solvents tested, while activities of DhaA and LinB decreased at comparable concentrations of the same co-solvent. Moreover, a high increase of DbjA enantioselectivity was observed. Ethylene glycol and 1,4-dioxane were shown to have the most positive impact on the enantioselectivity. The favorable influence of these co-solvents on both activity and enantioselectivity makes DbjA suitable for biocatalytic applications. This study represents the first investigation of the effects of organic co-solvents on the biocatalytic performance of haloalkane dehalogenases and will pave the way for their broader use in industrial processes.

• MeSH
• bakteriální proteiny chemie   metabolismus   MeSH
• cirkulární dichroismus MeSH
• furany MeSH
• hydrolasy chemie   metabolismus   MeSH
• organické látky chemie   MeSH
• rekombinantní fúzní proteiny chemie   metabolismus   MeSH
• rozpouštědla chemie   MeSH
• stabilita enzymů MeSH
• vysoká teplota MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The RNA chaperone Hfq plays crucial roles in bacterial gene expression and is a major facilitator of small regulatory RNA (sRNA) action. The toroidal architecture of the Hfq hexamer presents three well-characterized surfaces that allow it to bind sRNAs to stabilize them and engage target transcripts. Hfq-interacting sRNAs are categorized into two classes based on the surfaces they use to bind Hfq. By characterizing a systematic alanine mutant library of Hfq to identify amino acid residues that impact survival of Escherichia coli experiencing nitrogen (N) starvation, we corroborated the important role of the three RNA-binding surfaces for Hfq function. We uncovered two, previously uncharacterized, conserved residues, V22 and G34, in the hydrophobic core of Hfq, to have a profound impact on Hfq's RNA-binding activity in vivo. Transcriptome-scale analysis revealed that V22A and G34A Hfq mutants cause widespread destabilization of both sRNA classes, to the same extent as seen in bacteria devoid of Hfq. However, the alanine substitutions at these residues resulted in only modest alteration in stability and structure of Hfq. We propose that V22 and G34 have impact on Hfq function, especially critical under cellular conditions when there is an increased demand for Hfq, such as N starvation.

• MeSH
• bakteriální RNA * metabolismus   genetika   chemie   MeSH
• dusík metabolismus   MeSH
• Escherichia coli * genetika   metabolismus   MeSH
• hydrofobní a hydrofilní interakce * MeSH
• konzervovaná sekvence MeSH
• malá nekódující RNA * metabolismus   genetika   chemie   MeSH
• mutace MeSH
• protein hostitelského faktoru 1 * metabolismus   genetika   chemie   MeSH
• proteiny z Escherichia coli * metabolismus   genetika   chemie   MeSH
• regulace genové exprese u bakterií MeSH
• stabilita RNA * genetika   MeSH
• stanovení celkové genové exprese MeSH
• transkriptom genetika   MeSH
• vazba proteinů MeSH
• Publikační typ
• časopisecké články MeSH

The majority of naturally occurring proteins have evolved to function under mild conditions inside the living organisms. One of the critical obstacles for the use of proteins in biotechnological applications is their insufficient stability at elevated temperatures or in the presence of salts. Since experimental screening for stabilizing mutations is typically laborious and expensive, in silico predictors are often used for narrowing down the mutational landscape. The recent advances in machine learning and artificial intelligence further facilitate the development of such computational tools. However, the accuracy of these predictors strongly depends on the quality and amount of data used for training and testing, which have often been reported as the current bottleneck of the approach. To address this problem, we present a novel database of experimental thermostability data for single-point mutants FireProtDB. The database combines the published datasets, data extracted manually from the recent literature, and the data collected in our laboratory. Its user interface is designed to facilitate both types of the expected use: (i) the interactive explorations of individual entries on the level of a protein or mutation and (ii) the construction of highly customized and machine learning-friendly datasets using advanced searching and filtering. The database is freely available at https://loschmidt.chemi.muni.cz/fireprotdb.

• MeSH
• anotace sekvence MeSH
• bodová mutace * MeSH
• databáze proteinů * MeSH
• datové soubory jako téma MeSH
• internet MeSH
• molekulární modely MeSH
• proteiny chemie   genetika   MeSH
• software MeSH
• stabilita proteinů MeSH
• strojové učení statistika a číselné údaje   MeSH
• výpočetní biologie metody   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH