Protein kinase B controls Mycobacterium tuberculosis growth via phosphorylation of the transcriptional regulator Lsr2 at threonine 112.

Mycobacterium tuberculosis (Mtb) is able to persist in the body through months of multi-drug therapy. Mycobacteria possess a wide range of regulatory proteins, including the protein kinase B (PknB) which controls peptidoglycan biosynthesis during growth. Here, we observed that depletion of PknB resulted in specific transcriptional changes that are likely caused by reduced phosphorylation of the H-NS-like regulator Lsr2 at threonine 112. The activity of PknB towards this phosphosite was confirmed with purified proteins, and this site was required for adaptation of Mtb to hypoxic conditions, and growth on solid media. Like H-NS, Lsr2 binds DNA in sequence-dependent and non-specific modes. PknB phosphorylation of Lsr2 reduced DNA binding, measured by fluorescence anisotropy and electrophoretic mobility shift assays, and our NMR structure of phosphomimetic T112D Lsr2 suggests that this may be due to increased dynamics of the DNA-binding domain. Conversely, the phosphoablative T112A Lsr2 had increased binding to certain DNA sites in ChIP-sequencing, and Mtb containing this variant showed transcriptional changes that correspond with the change in DNA binding. In summary, PknB controls Mtb growth and adaptations to the changing host environment by phosphorylating the global transcriptional regulator Lsr2

Nitric oxide induces the distinct invisibility phenotype of Mycobacterium tuberculosis

During infection Mycobacterium tuberculosis (Mtb) forms physiologically distinct subpopulations that are recalcitrant to treatment and undetectable using standard diagnostics. These difficult to culture or differentially culturable (DC) Mtb are revealed in liquid media, their revival is often stimulated by resuscitation-promoting factors (Rpf) and prevented by Rpf inhibitors. Here, we investigated the role of nitric oxide (NO) in promoting the DC phenotype. Rpf-dependent DC Mtb were detected following infection of interferon-γ-induced macrophages capable of producing NO, but not when inducible NO synthase was inactivated. After exposure of Mtb to a new donor for sustained NO release (named NOD), the majority of viable cells were Rpf-dependent and undetectable on solid media. Gene expression analyses revealed a broad transcriptional response to NOD, including down-regulation of all five rpf genes. The DC phenotype was partially reverted by over-expression of Rpfs which promoted peptidoglycan remodelling. Thus, NO plays a central role in the generation of Rpf-dependent Mtb, with implications for improving tuberculosis diagnostics and treatments

EFFECT OF COMPLEX ADDITIONAL AND FLYING ASH ON CEMENT PROPERTIES

This article presents the study and improvement of the properties of the chemical complex additive C3 and mineral additive industrial waste by adding ash to the cement composition

Dimethyl fumarate eliminates differentially culturable Mycobacterium tuberculosis in an intranasal murine model of tuberculosis

Tuberculosis (TB) claims nearly 1.5 million lives annually. Current TB treatment requires a combination of several drugs administered for at least 6 months. Mycobacterium tuberculosis (Mtb), the causative agent of TB, can persist in infected humans and animals for decades. Moreover, during infection, Mtb produces differentially culturable bacteria (DCB) that do not grow in standard media but can be resuscitated in liquid media supplemented with sterile Mtb culture filtrates or recombinant resuscitation-promoting factors (Rpfs). Here, we demonstrate that, in an intranasal murine model of TB, Mtb DCB are detectable in the lungs after 4 weeks of infection, and their loads remain largely unchanged during a further 8 weeks. Treatment of the infected mice with dimethyl fumarate (DMF), a known drug with immunomodulatory properties, for 8 weeks eliminates Mtb DCB from the lungs and spleens. Standard TB treatment consisting of rifampicin, isoniazid, and pyrazinamide for 8 weeks reduces Mtb loads by nearly four orders of magnitude but does not eradicate DCB. Nevertheless, no DCB can be detected in the lungs and spleens after 8 weeks of treatment with DMF, rifampicin, isoniazid, and pyrazinamide. Our data suggest that addition of approved anti-inflammatory drugs to standard treatment regimens may improve TB treatment and reduce treatment duration.</p

Role of regional faults in the formation and placement of gold ore objects in western Uzbekistan

The results of the study, the relationship of regional faults with the processes of formation and mixing of gold ore occurrences in Western Uzbekistan are presented. According to researchers, the gold deposits of the region were formed under the influence of regional northwestern and transverse northeastern faults. So that the results of their research could serve as search signs and predictive criteria, they have not been studied enough to date. It is shown that the analysis of data on the location of gold ore occurrences in the network of regional faults in the region showed that 32% of gold ore objects were formed in the zones of northwestern faults. It is noted that the inter-fault space plays an important role in the placement of gold ore manifestations. Another important feature of the relationship between gold manifestations and regional faults has been revealed - this is the morphological feature (curvature area) of the faults, which determines the saturation of the inter-fault zone with manifestations of gold mineralization. The identified features of the relationship between regional faults and ring structures with gold occurrences are recommended to be used in forecasting and prospecting for gold

Efficient Protein Digestion at Elevated Temperature in the Presence of SDS and Calcium Ions for Membrane Proteomics.

Growing significance of membrane proteins inspires continuous development and improvement of methods for robust membrane proteomics. Here, we developed a very simple and efficient method for membrane protein digestion using an ionic detergent SDS at high temperature, conditions where trypsin is normally inactivated. Our results suggest that trypsin can be stabilized by a combination of calcium ions and sodium chloride which enables protein digestion at elevated temperature in the presence of SDS. Application of this method resulted in higher efficiency of digestion and improved identification of membrane proteins, offering novel opportunities for advancement in membrane proteomics

Mycobacterial phosphatase PstP regulates global serine threonine phosphorylation and cell division.

Protein phosphatase PstP is conserved throughout the Actinobacteria in a genetic locus related to cell wall synthesis and cell division. In many Actinobacteria it is the sole annotated serine threonine protein phosphatase to counter the activity of multiple serine threonine protein kinases. We used transcriptional knockdown, electron microscopy and comparative phosphoproteomics to investigate the putative dual functions of PstP as a specific regulator of cell division and as a global regulator of protein phosphorylation. Comparative phosphoproteomics in the early stages of PstP depletion showed hyperphosphorylation of protein kinases and their substrates, confirming PstP as a negative regulator of kinase activity and global serine and threonine phosphorylation. Analysis of the 838 phosphorylation sites that changed significantly, suggested that PstP may regulate diverse phosphoproteins, preferentially at phosphothreonine near acidic residues, near the protein termini, and within membrane associated proteins. Increased phosphorylation of the activation loop of protein kinase B (PknB) and of the essential PknB substrate CwlM offer possible explanations for the requirement for pstP for growth and for cell wall defects when PstP was depleted

Structural and optical properties of sol-gel synthesized TiO2 nanocrystals: Effect of Ni and Cr (co)doping

Nickel and chromium metal ions have been utilized to dope and co-dope titanium dioxide nanocrystals, thereby broadening the light absorption range of titania into the visible light spectrum. The doped and co-doped TiO2 nanocrystals were prepared using sol-gel techniques, with a varied doping concentration that extended from 0.25 to 10.0 wt%.These modified materials underwent comprehensive analysis using standard analytical tools such as X-ray diffraction, Raman spectroscopy, BET surface area measurement, Fourier transform infrared spectroscopy, UV–vis diffuse reflectance spectroscopy, and fluorescence spectroscopy. Powder XRD technique reveals that the modified catalyst majorly contains the anatase polymorph with the transition metal ion either substituting Ti or located as interstitials in the lattice of TiO2. Raman and UV–Vis absorption spectra of the doped and codoped catalyst a show λmax shift towards longer wavelength when in the metal ion concentration is increased from 0.25 to 10 wt%. FTIR patterns show the stretching and vibration patterns of hydroxyl radicals present in the nanocrystals. The BET surface areas of the doped and codoped TiO2 nanocrystals have substantially higher surface areas compared with that of the undoped TiO2. Electronic structural studies of the TiO2, (Ti,Cr)O2, (Ti,Ni)O2, and (Ti,Ni,Cr)O2 crystals using density functional theory indicated a reduction in the band gap width (Eg) of pure TiO2 when doped with transition metals. Specifically, the indirect bandgap values for Ni(10%), Cr(10%) and NiCr(5% + 5%) doping were 2.96, 2.83, and 2.7 eV, respectively. Furthermore, the photoluminescence intensity substantially decreased with the incorporation of transition metal ions in the TiO2 nanocrystal

A Mycobacterium tuberculosis Effector Targets Mitochondrion, Controls Energy Metabolism, and Limits Cytochrome c Exit

Host metabolism reprogramming is a key feature of Mycobacterium tuberculosis (Mtb) infection that enables the survival of this pathogen within phagocytic cells and modulates the immune response facilitating the spread of the tuberculosis disease. Here, we demonstrate that a previously uncharacterized secreted protein from Mtb, Rv1813c, manipulates the host metabolism by targeting mitochondria. When expressed in eukaryotic cells, the protein is delivered to the mitochondrial intermembrane space and promotes the enhancement of host ATP production by boosting the oxidative phosphorylation metabolic pathway. Furthermore, the release of cytochrome c from mitochondria, an early apoptotic event in response to short-term oxidative stress, is delayed in Rv1813c-expressing cells. This study reveals a novel class of mitochondria targeting effectors from Mtb that might participate in host cell metabolic reprogramming and apoptosis control during Mtb infections. IMPORTANCE In this article, using a combination of techniques (bioinformatics, structural biology, and cell biology), we identified and characterized a new class of effectors present only in intracellular mycobacteria. These proteins specifically target host cell mitochondria when ectopically expressed in cells. We showed that one member of this family (Rv1813c) affects mitochondria metabolism in a way that might twist the immune response. This effector also inhibits the cytochrome c exit from mitochondria, suggesting that it might alter normal host cell apoptotic capacities, one of the first defenses of immune cells against Mtb infection