Acetylene containing 2-(2-hydrazinyl)thiazole derivatives: design, synthesis, and in vitro and in silico evaluation of antimycobacterial activity against Mycobacterium tuberculosis.

Mycobacterium tuberculosis resistance to commercially available drugs is increasing day by day. To address this issue, various strategies were planned and are being implemented. However, there is a need for new drugs and rapid diagnostic methods. For this endeavour, in this paper, we present the synthesis of acetylene containing 2-(2-hydrazinyl) thiazole derivatives and in vitro evaluation against the H37Rv strain of Mycobacterium tuberculosis. Among the developed 26 acetylene containing 2-(2-hydrazinyl) thiazole derivatives, eight compounds inhibited the growth of Mycobacterium tuberculosis with MIC values ranging from 100 µg ml-1 to 50 µg ml-1. The parent acetylene containing thiosemicarbazones showed promising antimycobacterial activity by inhibiting up to 75% of the Mycobacterium at 50 µg ml-1. In addition, in silico studies were employed to understand the binding mode of all the novel acetylene-containing derivatives against the KasA protein of the Mycobacterium. Interestingly, the KasA protein interactions with the compounds were similar to the interactions of KasA protein with thiolactomycin and rifampicin. Cytotoxicity study results indicate that the compounds tested are non-toxic to human embryonic kidney cells.

PRMT3 interacts with ALDH1A1 and regulates gene-expression by inhibiting retinoic acid signaling.

Protein arginine methyltransferase 3 (PRMT3) regulates protein functions by introducing asymmetric dimethylation marks at the arginine residues in proteins. However, very little is known about the interaction partners of PRMT3 and their functional outcomes. Using yeast-two hybrid screening, we identified Retinal dehydrogenase 1 (ALDH1A1) as a potential interaction partner of PRMT3 and confirmed this interaction using different methods. ALDH1A1 regulates variety of cellular processes by catalyzing the conversion of retinaldehyde to retinoic acid. By molecular docking and site-directed mutagenesis, we identified the specific residues in the catalytic domain of PRMT3 that facilitate interaction with the C-terminal region of ALDH1A1. PRMT3 inhibits the enzymatic activity of ALDH1A1 and negatively regulates the expression of retinoic acid responsive genes in a methyltransferase activity independent manner. Our findings show that in addition to regulating protein functions by introducing methylation modifications, PRMT3 could also regulate global gene expression through protein-protein interactions.

The ribosomal protein eL21 interacts with the protein lysine methyltransferase SMYD2 and regulates its steady state levels.

The protein lysine methyltransferase, SMYD2 is involved in diverse cellular events by regulating protein functions through lysine methylation. Though several substrate proteins of SMYD2 are well-studied, only a limited number of its interaction partners have been identified and characterized. Here, we performed a yeast two-hybrid screening of SMYD2 and found that the ribosomal protein, eL21 could interact with SMYD2. SMYD2-eL21 interaction in the human cells was confirmed by immunoprecipitation methods. In vitro pull-down assays revealed that SMYD2 interacts with eL21 directly through its SET and MYND domain. Computational mapping, followed by experimental studies identified that Lys81 and Lys83 residues of eL21 are important for the SMYD2-eL21 interaction. Evolutionary analysis showed that these residues might have co-evolved with the emergence of SMYD2. We found that eL21 regulates the steady state levels of SMYD2 by promoting its transcription and inhibiting its proteasomal degradation. Importantly, SMYD2-eL21 interaction plays an important role in regulating cell proliferation and its dysregulation might lead to tumorigenesis. Our findings highlight a novel extra-ribosomal function of eL21 on regulating SMYD2 levels and imply that ribosomal proteins might regulate wide range of cellular functions through protein-protein interactions in addition to their core function in translation.

The uncharacterized protein FAM47E interacts with PRMT5 and regulates its functions.

Protein arginine methyltransferase 5 (PRMT5) symmetrically dimethylates arginine residues in various proteins affecting diverse cellular processes such as transcriptional regulation, splicing, DNA repair, differentiation, and cell cycle. Elevated levels of PRMT5 are observed in several types of cancers and are associated with poor clinical outcomes, making PRMT5 an important diagnostic marker and/or therapeutic target for cancers. Here, using yeast two-hybrid screening, followed by immunoprecipitation and pull-down assays, we identify a previously uncharacterized protein, FAM47E, as an interaction partner of PRMT5. We report that FAM47E regulates steady-state levels of PRMT5 by affecting its stability through inhibition of its proteasomal degradation. Importantly, FAM47E enhances the chromatin association and histone methylation activity of PRMT5. The PRMT5-FAM47E interaction affects the regulation of PRMT5 target genes expression and colony-forming capacity of the cells. Taken together, we identify FAM47E as a protein regulator of PRMT5, which promotes the functions of this versatile enzyme. These findings imply that disruption of PRMT5-FAM47E interaction by small molecules might be an alternative strategy to attenuate the oncogenic function(s) of PRMT5.

In situ formation, structural, mechanical and in vitro analysis of ZrO2/ZnFe2O4 composite with assorted composition ratios.

The investigation underline the in situ formation of ZrO2/ZnFe2O4 composites and the resultant structural, morphological, mechanical and magnetic properties. The characterization results ensured the crystallization of tetragonal ZrO2 (t-ZrO2) and ZnFe2O4 phases at 900 °C. Depending on Zn2+/Fe3+ content, the composite system revealed a gradual increment in the phase yield of ZnFe2O4. The significance of monoclinic ZrO2 (m-ZrO2) is also evident in all the systems at 900 °C; however, the incremental heat treatment to 1300 °C indicated its corresponding loss, thus indicating the reverse m- â†’ t-ZrO2 transition. The crystallization of ZnFe2O4 as a secondary phase in the t-ZrO2 matrix is also affirmed from the morphological analysis. Mechanical studies accomplished good uniformity in all the investigated compositions despite the variation in the phase content of ZnFe2O4 in composite system. All the t-ZrO2/ZnFe2O4 composites ensured strong ferrimagnetic features and moreover better biocompatibility and non-toxicity characteristics were displayed from in vitro tests.

SET7/9 interacts and methylates the ribosomal protein, eL42 and regulates protein synthesis.

Methylation of proteins is emerging to be an important regulator of protein function. SET7/9, a protein lysine methyltransferase, catalyses methylation of several proteins involved in diverse biological processes. SET7/9-mediated methylation often regulates the stability, sub-cellular localization and protein-protein interactions of its substrate proteins. Here, we aimed to identify novel biological processes regulated by SET7/9 by identifying new interaction partners. For this we used yeast two-hybrid screening and identified the large subunit ribosomal protein, eL42 as a potential interactor of SET7/9. We confirmed the SET7/9-eL42 interaction by co-immunoprecipitation and GST pulldown studies. The N-terminal MORN domain of SET7/9 is essential for its interaction with eL42. Importantly, we identified that SET7/9 methylates eL42 at three different lysines - Lys53, Lys80 and Lys100 through site-directed mutagenesis. By puromycin incorporation assay, we find that SET7/9-mediated methylation of eL42 affects global translation. This study identifies a new role of the functionally versatile SET7/9 lysine methyltransferase in the regulation of global protein synthesis.

2,4-Di-Tert-Butylphenol Isolated From an Endophytic Fungus, Daldinia eschscholtzii, Reduces Virulence and Quorum Sensing in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is among the top three gram-negative bacteria according to the WHO's critical priority list of pathogens against which newer antibiotics are urgently needed and considered a global threat due to multiple drug resistance. This situation demands unconventional antimicrobial strategies such as the inhibition of quorum sensing to alleviate the manifestation of classical resistance mechanisms. Here, we report that 2,4-di-tert-butylphenol (2,4-DBP), isolated from an endophytic fungus, Daldinia eschscholtzii, inhibits the quorum-sensing properties of P. aeruginosa. We have found that treating P. aeruginosa with 2,4-DBP substantially reduced the secretion of virulence factors as well as biofilm, and its associated factors that are controlled by quorum sensing, in a dose-dependent manner. Concomitantly, 2,4-DBP also significantly reduced the expression of quorum sensing-related genes, i.e., lasI, lasR, rhlI, and rhlR significantly. Importantly, 2,4-DBP restricted the adhesion and invasion of P. aeruginosa to the A549 lung alveolar carcinoma cells. In addition, bactericidal assay with 2,4-DBP exhibited synergism with ampicillin to kill P. aeruginosa. Furthermore, our computational studies predicted that 2,4-DBP could bind to the P. aeruginosa quorum-sensing receptors LasR and RhlR. Collectively, these data suggest that 2,4-DBP can be exploited as a standalone drug or in combination with antibiotic(s) as an anti-virulence and anti-biofilm agent to combat the multidrug resistant P. aeruginosa infection.

Structure, luminescence, mechanical and in vitro behavior of zirconia toughened alumina due to terbium substitutions.

The significance of Tb3+ inclusions at the zirconia toughened alumina (ZTA) structure was explored. The influence of Tb3+ content at the crystal structures of ZrO2 and Al2O3 and the resultant optical, mechanical, magnetic and cytotoxicity properties were deliberated. The critical role of Tb3+ to attain a structurally stable ZTA until 1500 °C is ensured. Depending on the Tb3+ content, either tetragonal zirconia (t-ZrO2) or cubic zirconia (c-ZrO2) structures were stabilized while the propensity of Tb3+ reaction with Al2O3 to yield TbAlO3 is transpired only after exceeding the occupancy limit in ZrO2. The green emission and paramagnetic features are imparted by the Tb3+ inclusions at the ZTA structure. Dense and pore free microstructures with a direct impact on the improved mechanical features of ZTA is empowered by the presence of Tb3+. Further, the results from MTT assay and live/dead cell staining ensured the negligence of Tb3+ contained ZTA systems to induce toxicity.

Structural and bio-mineralization features of alumina zirconia composite influenced by the combined Ca2+ and PO43- additions.

The structural and bioactivity features of alumina zirconia composite (AZC) due to Ca2+ and PO43- additions are demonstrated. An in situ synthetic approach, starting from the solution precursors is devised for the powder synthesis in which the assorted range of Ca2+ and PO43- additions were done to the equimolar concentrations of Al3+ and Zr4+ precursors. The results witnessed the unique crystallization of tetragonal zirconia (t-ZrO2) at 1100 °C while Ca2+, PO43- and Al2O3 remained in their amorphous state in the system. On further heat treatment, α-Al2O3 crystallized at 1200 °C, which enforced t- → m-ZrO2 transformation while Ca2+ and PO43- still retained their amorphous state. The immersion tests in simulated body fluid (SBF) solution validated the enhanced bio-mineralization activity of AZC due to Ca2+ and PO43- additions. The results from the indentation tests demonstrated good uniformity in the elastic modulus and hardness data of the investigated specimens. Further, in vitro cell culture tests ascertained the bioactivity of all the AZC compositions.