Synthesis, Crystal Structure, Hirshfeld Surface Analysis, DFT, and DNA-Binding Studies of ( E )-2-(3-Hydroxy-4-Methoxybenzylidene)Hydrazinecarbothioamide Abstract (E)-2-(3-Hydroxy-4-methoxybenzylidene)hydrazinecarbothioamide 3 was synthesized by reacting thiosemicarbazide with 2-hydorxy-3-methoxybenzaldehyde in dry ethanol. The structure was elucidated by spectroscopic (FT-IR, 1H NMR, and 13C NMR) and single crystal X-ray diffraction techniques. A detailed analysis of the intermolecular interactions has been performed based on the Hirshfeld surfaces and their associated two-dimensional fingerprint plots. DFT, spectroscopic, and electrochemical DNA-binding analysis confirmed that the compound is reactive to bind with DNA. Viscometric studies suggested that compound 3 has a mixed mode of interaction and intercalated into the DNA base pairs predominantly along with the possibility of electrostatic interactions. Graphical Abstract

Isolation of Bisphenol A-Tolerating/degrading Shewanella haliotis Strain MH137742 from an Estuarine Environment Abstract The human exposure to bisphenol A (BPA) occurs frequently. Once, this compound was one of the highest volume chemicals produced worldwide and used as a plasticizer in many products. However, even at low concentration, it can cause severe damage to the endocrine system because of its endocrine disruptor activity. Thus, mitigation studies to remove or reduce this contaminant from the environment are essential. An alternative method of removing it from the environment is the use of bioremediation processes to the selected bacteria isolated from a BPA-impacted area. In this work, four halotolerant strains were isolated from the Santos Estuary System, one of the most important Brazilian examples of environmental degradation. In the present work, one strain presented strong BPA tolerance and high BPA-degrading activity and could grow in a minimum medium containing BPA as the main carbon source. Strain MH137742 was identified as Shewanella haliotis, based on 16S rRNA gene sequencing and mass spectrometry identification by MALDI-TOF Biotyper. Shewanella haliotis was able to tolerate up to 150 mg L−1 of BPA and biotransform 75 mg L−1 in 10 h in a liquid culture medium. Based on the analysis of the produced metabolites by LC-MS, it was possible to predict the metabolic pathway used by this microorganism to degrade the BPA.

Amino Acids as Additives against Amorphous Aggregation: In Vitro and In Silico Study on Human Lysozyme Abstract The effect of 16 amino acids (AA) with various physicochemical properties was investigated on human lysozyme (HL) heat-induced amorphous aggregation. UV-Visible spectrophotometry was used to monitor the kinetics of aggregation in the absence and presence of AA, and transmission electron microscopy (TEM) images were taken from the aggregates. To conduct in silico experiments, Autodock vina was used for docking of AA into protein (via YASARA interface), and FTmap information was checked for an insight onto putative binding sites. Prediction of aggregation-prone regions of lysozyme was made by AGGRESCAN and Tango. Among all tested AA, phenylalanine had the best anti-aggregation effect, followed by lysine. In addition, based on in silico tests, Trp 109 and Val 110 of lysozyme are suggested to be of importance in the aggregation process of the enzyme. In conclusion, phenylalanine, arginine, and lysine were found to affect the nucleation phase of lysozyme aggregation and could be considered as suitable stabilizing structures for this enzyme.

Establishment of CRISPR/Cas9-Mediated Knock-in System for Porcine Cells with High Efficiency Abstract Since the birth of clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9, the new genome engineering technology has become a hot topic in the scientific community. However, for swine, the system of pig cells’ homology directed repair (HDR) is generally unstable and costly. Here, we aim to make knock-in of porcine cells more realizable. The Rosa26 locus was chosen for gene editing. Through the optimization of strategy, an efficient sgRNA was selected by TIDE analysis. Correspondingly, a vector system was constructed for gene insertion in pRosa26 locus by homologous recombination. A large percentage of cells whose gene is edited easily result in apoptosis. To improve the positive rate, culturing systems have been optimized. Sequence alignment and nuclear transfer confirmed that we got two knock-in cell lines and transgene primary porcine fetal fibroblasts (PFFs) successfully. Results showed that the gene editing platform we used can obtain genetically modified pig cells stably and efficiently. This system can contribute to pig gene research and production of transgenic pigs.

Extracellular Expression of L-Aspartate-α-Decarboxylase from Bacillus tequilensis and Its Application in the Biosynthesis of β-Alanine Abstract L-aspartate-α-decarboxylase was extracellularly expressed to enhance its production for β-alanine biosynthesis. L-aspartate-α-decarboxylase and cutinase were coexpressed in Escherichia coli; more than 40% of the L-aspartate-α-decarboxylase was secreted into the medium. Selection of best conditions among tested variables enhanced L-aspartate-α-decarboxylase production by the recombinant strain. The total L-aspartate-α-decarboxylase activity reached 20.3 U/mL. Analysis of the enzymatic properties showed that the optimum temperature and pH for L-aspartate-α-decarboxylase were 60 °C and 7.5, respectively. Enzyme activity was stable at pH 4.0–8.5 and displayed sufficient thermal stability at temperatures < 50 °C. In addition, enzymatic synthesis of β-alanine was performed using extracellularly expressed L-aspartate-α-decarboxylase, and a mole conversion rate of > 99% was reached with a substrate concentration of 1.5 M. Extracellular expression of L-aspartate-α-decarboxylase resulted in increased enzyme production, indicating its possible application in the enzymatic synthesis of β-alanine.

Use of an (Hemi) Cellulolytic Enzymatic Extract Produced by Aspergilli Species Consortium in the Saccharification of Biomass Sorghum Abstract This study evaluated the production of lignocellulose-degrading enzymes by solid-state fermentation (SSF) using a microbial consortium of Aspergillus fumigatus SCBM6 and A. niger SCBM1 (AFN extract). The fungal strains were cultivated in sugarcane bagasse (SCB) and wheat bran (WB) as lignocellulosic substrates for 7 days at 30 °C. After SSF, the highest peaks of enzyme production were 150 and 80 U g−1 for β-xylosidase and β-glucosidase at 48 h, 375 U g−1 for xylanase at 96 h, and 80 U g−1 for endoglucanase and 4 U g−1 for cellulase activity on filter paper (FPase) at 144 h. The efficiency of the produced AFN extract was investigated in the enzymatic hydrolysis of crude biomass sorghum (BS) and after the removal of extractives (ES). After saccharification, the glucose and xylose concentrations were 10× superior in ES than in BS hydrolysate (2.5 g L−1 after 12 h). The presence of inhibitors of alcoholic fermentation, such as formic acid, was also reduced in ES hydrolysates, indicating that the removal of extractives positively contributed to the effectiveness of enzymatic hydrolysis of biomass sorghum using AFN extract.

Gaseous Formaldehyde Degrading by Methylobacterium sp. XJLW Abstract Formaldehyde is harmful to human beings. It is widely used in chemical industry, medicine, and agriculture and is frequently discharged into the sewage. Microbial metabolism of formaldehyde has attracted increasing attention for its potential application in formaldehyde removal, especially for indoor gaseous formaldehyde degradation. Methylobacterium sp. XJLW capable of degrading formaldehyde was isolated and exhibited a strong activity for liquid formaldehyde degradation. In the present study, the survival rate of XJLW was evaluated under drought, 30 °C, 4 °C, 15 °C, 35 °C, and 40 °C. After 4 days, the average survival rate under 30°C is the greatest (83.97%) among the five temperatures. Whether the temperature was above or below 30°C, the average survival rate decreased significantly. However, the resistance of XJLW to reduced temperatures seemed better than that to increased temperatures. The average survival rate under 15°C and 4°C was 71.1% and 58.67%, while that under 35 °C and 40 °C was 49.47% and 0.1%. Two batches of gaseous formaldehyde treatments were carried out in an analog device with super absorbent polymer (SAP) as the carrier materials of XJLW. The results showed that XJLW could effectively degrade gaseous formaldehyde in the analog device for a long period.

Improvement of Methane Production from Sugar Beet Wastes Using TiO 2 and Fe 3 O 4 Nanoparticles and Chitosan Micropowder Additives Abstract An experimental study was performed to measure biogas production from sugar beet waste, which is, in fact, the chopped parts of the sugar beet not going through the sugar extraction process, at different additive concentrations. Medium molecular weight chitosan in microsize and TiO2 and Fe3O4 nanoparticles were added to ten experimental reactors to investigate their effect on the anaerobic digestion process. Three different concentrations of 0.01, 0.04, and 0.12% w/w were used for each additive. Biogas production and methane content were compared with a control sample containing no additive. Adding chitosan in powder form did not help the process nor improved methanogenic activities. The results showed no effect on anaerobic digestion by the addition of TiO2 nanoparticles in the mentioned concentrations, whereas adding Fe3O4 nanoparticles led to a slight increase in methane production and in volatile solid and total solid reduction. The maximum enhancement in methane and biogas production in the sample containing 0.04% Fe3O4, as compared with the control sample, reached 19.77% and 15.09%, respectively.

Utilization of a Silicone Rubber Membrane for Passive Oxygen Supply in a Microbial Fuel Cell Treating Carbon and Nitrogen from Synthetic Coke-Oven Wastewater Abstract This study firstly introduced a silicone rubber membrane (SRM) into microbial fuel cell (MFC) for passive oxygen supply to simultaneously remove phenol and nitrogen from synthetic coke-oven wastewater diluted with seawater. Passive oxygen transport with biofilm on the membrane was improved by ~ 18-fold in comparison with the one without a biofilm. In addition, although the oxygen supply was passive, nitrification accounted for 34% of those aeration conditions. It was also found that silicone rubber membrane can control NO2−–N and/or NO3−–N production. A dual-chamber MFC treating the synthetic coke-oven wastewater achieved a maximum power density of 54 mW m−2 with a coulombic efficiency of 2.7%. We conclude that silicone rubber membrane is effective for sustainable coke-oven wastewater treatment in MFCs.

Inhibitory Effects of Vanadium-Binding Proteins Purified from the Sea Squirt Halocynthia roretzi on Adipogenesis in 3T3-L1 Adipocytes Abstract The inhibitory effects of vanadium-binding proteins (VBPs) from the blood plasma and the intestine of sea squirt on adipogenesis in 3T3-L1 adipocytes were examined. 3T3L-1 cells treated with VBP blood plasma decreased markedly the lipid content in maturing pre-adipocytes in a dose-dependent manner, whereas VBP intestine did not show significant effects on lipid accumulation. Both VBPs did not have significant effect on cell viability. In order to demonstrate the anti-adipogenic effects of VBP blood plasma, the expressions of several adipogenic transcription factors and enzymes were investigated by Reverse Transcriptase-Polymerase Chain Reaction. VBP blood plasma down-regulated the expressions of transcription factors; PPAR-γ, C/EBP-α, SREBP1, and FAS, but did not have significant effects on the expressions of lipolytic enzymes; HSL and LPL. Both the crude and purified VBPs significantly increased the mRNA levels of Wnt10b, FZ1, LRP6, and β-catenin, while decreased the expression of GSK-3β. Hence, VBP blood plasma inhibited adipogenesis by activating WNT/β-catenin pathway via the activation of Wnt10b. Based on the findings, VBP blood plasma decreased lipid accumulation which was mediated by decreasing adipogenesis, not by lipolysis. Therefore, VBP blood plasma could be used to treat obesity.