The aldehyde dehydrogenase AldA contributes to the hypochlorite defense and is redox-controlled by protein S-bacillithiolation in Staphylococcus aureus

Staphylococcus aureus produces bacillithiol (BSH) as major low molecular weight (LMW) thiol which functions in thiol-protection and redox-regulation by protein S-bacillithiolation under hypochlorite stress. The aldehyde dehydrogenase AldA was identified as S-bacillithiolated at its active site Cys279 under NaOCl stress in S. aureus. Here, we have studied the expression, function, redox regulation and structural changes of AldA of S. aureus. Transcription of aldA was previously shown to be regulated by the alternative sigma factor SigmaB. Northern blot analysis revealed SigmaB-independent induction of aldA transcription under formaldehyde, methylglyoxal, diamide and NaOCl stress. Deletion of aldA resulted in a NaOCl-sensitive phenotype in survival assays, suggesting an important role of AldA in the NaOCl stress defense. Purified AldA showed broad substrate specificity for oxidation of several aldehydes, including formaldehyde, methylglyoxal, acetaldehyde and glycol aldehyde. Thus, AldA could be involved in detoxification of aldehyde substrates that are elevated under NaOCl stress. Kinetic activity assays revealed that AldA is irreversibly inhibited under H2O2 treatment in vitro due to overoxidation of Cys279 in the absence of BSH. Pre-treatment of AldA with BSH prior to H2O2 exposure resulted in reversible AldA inactivation due to S-bacillithiolation as revealed by activity assays and BSH-specific Western blot analysis. Using molecular docking and molecular dynamic simulation, we further show that BSH occupies two different positions in the AldA active site depending on the AldA activation state. In conclusion, we show here that AldA is an important target for S-bacillithiolation in S. aureus that is up-regulated under NaOCl stress and functions in protection under hypochlorite stress

Modeling allosteric signal propagation using protein structure networks

Allosteric communication in proteins can be induced by the binding of effective ligands, mutations or covalent modifications that regulate a site distant from the perturbed region. To understand allosteric regulation, it is important to identify the remote sites that are affected by the perturbation-induced signals and how these allosteric perturbations are transmitted within the protein structure. In this study, by constructing a protein structure network and modeling signal transmission with a Markov random walk, we developed a method to estimate the signal propagation and the resulting effects. In our model, the global perturbation effects from a particular signal initiation site were estimated by calculating the expected visiting time (EVT), which describes the signal-induced effects caused by signal transmission through all possible routes. We hypothesized that the residues with high EVT values play important roles in allosteric signaling. We applied our model to two protein structures as examples, and verified the validity of our model using various types of experimental data. We also found that the hot spots in protein binding interfaces have significantly high EVT values, which suggests that they play roles in mediating signal communication between protein domains

Modulation of host cell processes by T3SS effectors

Two of the enteric Escherichia coli pathotypes-enteropathogenic E. coli (EPEC) and enterohaemorrhagic E. coli (EHEC)-have a conserved type 3 secretion system which is essential for virulence. The T3SS is used to translocate between 25 and 50 bacterial proteins directly into the host cytosol where they manipulate a variety of host cell processes to establish a successful infection. In this chapter, we discuss effectors from EPEC/EHEC in the context of the host proteins and processes that they target-the actin cytoskeleton, small guanosine triphosphatases and innate immune signalling pathways that regulate inflammation and cell death. Many of these translocated proteins have been extensively characterised, which has helped obtain insights into the mechanisms of pathogenesis of these bacteria and also understand the host pathways they target in more detail. With increasing knowledge of the positive and negative regulation of host signalling pathways by different effectors, a future challenge is to investigate how the specific effector repertoire of each strain cooperates over the course of an infection

Проектирование пользовательского взаимодействия в иммерсивных системах нового поколения

This article explores the issues of designing intelligent systems using augmented reality (AR) and virtual reality (VR) technologies. These technologies are based on the use of three-dimensional representation of the surrounding real or pre-designed virtual scene, the capabilities of these systems for determining the orientation of user and objects in space, intelligent analysis algorithms and machine learning. The complexity of developing individual algorithms, the need to take into account the specifics of the subject area for each applied system, interactive display based on threedimensional and other types of representations necessitates the development of new approaches and design principles. The article focuses on the capabilities of OSTIS Technology for use in these tasks, and also provides examples of building a system for technical description of equipment and a guided quest in augmented reality

Kristallstruktur und Intramolekularer Elelktronentransfer

Title page Table of contents Theoretical part Materials and Methods Results Discussion Conclusion ReferencesThe main aim of the present doctoral work was to show that Adx can be photoreduced without presence of AR, and hence, to probe the "shuttle" mechanism. In order to achieve the aim, the truncated form of Adx, Adx(1-108), was covalently modified with Ru(bpy)2(mbpy). Ru(bpy)2(mbpy)-Adx(1-108) was characterized by X-ray crystallography, biophysical and biochemical methods. Possible electron-transfer pathways within the covalent 1:1 Ru(bpy)2(mbpy)-Adx(1-108) complex were predicted. Photoreduction of the [2Fe- 2S] cluster of adrenodoxin was confirmed by EPR measurements. The electron- transfer rate is concentration dependent and indicates that intermolecular electron transfer takes place. The monomolecular intramolecular rate was derived at zero concentration of the Ru(bpy)2(mbpy)-Adx(1-108) complex. Comparative investigation of the covalent 1:1 Ru(bpy)2(mbpy)-Adx(1-108) complex with the wild-type Adx and unmodified Adx(1-108) showed that, in general, reduction of Adx, one-electron carrier, is accompanied by a transfer of two electrons. However, spin quantification of the reduced [2Fe-2S] cluster indicated that only one of two irons is in Fe2+ state. At present, there is no clear explanation for this unusual feature and it needs further investigation. In general, the combination of theory and present experimental data provides a basis for further investigations of the mechanism of the electron transfer in the P450-system.Das Hauptziel der vorliegenden Arbeit war, mittels einer Photoreduktion des Adrenodoxins zu zeigen, dass das Eisen-Schwefel-Protein ohne Adrenodoxinreduktase reduziert und damit das Shuttle-Modell für Adrenodoxin überprüft werden kann. Um dieses Hauptziel zu erreichen, wurde eine verkürzte Form des Adrenodoxins, Adx(1-108), mit dem Ru(bpy)2(mbpy)-Komplex kovalent modifiziert. Ru(bpy)2(mbpy)-modifiziertes Adx(1-108) wurde mittels Röntgenstrukturanalyse, biophysikalischer und biochemischer Methoden untersucht. Anhand der Struktur des 1:1 Ru(bpy)2(mbpy)-Adx(1-108)-Komplexes wurden mögliche Elektronentransferwege innerhalb des Komplexes vorgeschlagen. Photoreduktion des Eisen-Schwefel-Zentrums des Adrenodoxins wurde anhand von EPR-Messungen nachgewiesen. Die Elektronentransferrate ist konzentrationsabhängig und zeigt, dass intermolekulare Elektronenübertragung stattfindet. Die intramolekulare Rate wurde bei unendlicher Verdünnung des Ru(bpy)2(mbpy)-Adx(1-108)-Komplexes berechnet. Bei der vergleichenden Untersuchung des 1:1 Ru(bpy)2(mbpy)-Adx(1-108)-Komplexes mit dem Wildtyp- Adrenodoxin und freiem Adx(1-108) wurde festgestellt, dass Adrenodoxin als Ein-Elektrontransferprotein eine Tendenz zeigt, bei Übertragung von zwei Elektronen reduziert zu werden, obwohl die Spinquantifizierung deutlich zeigt, dass nur ein Eisenatom den Fe2+-Zustand einnimmt. Die Ursache für das ungewöhnliche Verhalten des Proteins ist unklar und muss weiter untersucht werden. Die Kombination der theoretischen Kenntnisse mit experimentellen Daten erlaubt einen Einblick in den Mechanismus des Elektronentransports im P450-System. Damit wurde die Grundlage für weiterführende Untersuchungen zum Elektronentransport im Cytochrom P450-System geschaffen

3D-представление объектов в интеллектуальных компьютерных системах нового поколения

This article is dedicated to the issues of constructing and using a three-dimensional representation in various tasks of applied intelligent systems, as well as corresponding systems of spatial positioning and orientation. The description of the representation itself, as well as the principles of its construction, is implemented within the knowledge base of the OSTIS system, which allows for deep integration of various tasks and methods, and also subsequently leads to an increased degree of convergence of various research domains

Интероперабельность как важнейший компонент интеллектуальной образовательной среды в средней школе

The paper presents some results of an analysis of the role of the development of interoperability, cognitive abilities and emotional intelligence in children in a modern school. The importance and ways of introducing technological tools with capabilities for interaction and data exchange to optimize the educational process are discussed. The signifi cance of the development of cognitive abilities and emotional intelligence of students and the impact of this on their academic achievements and social adaptation are also considered

Coiled-coil dimerization of the LOV2 domain of the blue-light photoreceptor phototropin 1 from<i>Arabidopsis thaliana</i>

A key role in signal transduction and dimerization mediated by Per–Arnt–Sim (PAS) domains is played by α-helical linkers that flank the structurally similar α/β cores of these domains. However, crystal-packing forces and the different construct lengths and sequences of the PAS domains influence the final length and orientation of the linkers relative to the core and create uncertainty in the exact mechanism of the linker function. Thus, structural characterization and comparison of the linkers within isolated PAS-domain constructs and/or full-length PAS-containing proteins is important for clarification of the mechanism. The plant blue-light photoreceptors phototropins possess two N-terminal flavin mononucleotide-based light, oxygen or voltage (LOV) domains (LOV1 and LOV2) that comprise a subclass of the PAS family and one C-terminal serine/threonine kinase domain whose enzymatic activity is regulated by blue light. The dark-adapted state crystal structures of the Arabidopsis thaliana phototropin 1 and phototropin 2 LOV1-domain constructs flanked by an N-terminal A′α helix and the structure of the phototropin 2 core LOV2 domain are known. Here, the crystal structure of the A. thaliana phototropin 1 LOV2 domain has been determined in its dark-adapted state. The core is flanked by an N-terminal A′α helix and a C-terminal Jα helix similar to those in the previously reported structure of Avena sativa phototropin 1 LOV2. In contrast to the monomeric A. sativa LOV2, A. thaliana LOV2 is a dimer in which two A′α helices adopt a scissor-like orientation at the dimer interface and form a short α-helical coiled coil. The Jα helix predominantly interacts with the β-sheet and plays a role in coiled-coil formation and dimerization