Capture-and-release of a sulfoquinovose-binding protein on sulfoquinovose-modified agarose.

The solute-binding protein (SBP) components of periplasmic binding protein-dependent ATP-binding cassette (ABC)-type transporters often possess exquisite selectivity for their cognate ligands. Maltose binding protein (MBP), the best studied of these SBPs, has been extensively used as a fusion partner to enable the affinity purification of recombinant proteins. However, other SBPs and SBP-ligand based affinity systems remain underexplored. The sulfoquinovose-binding protein SmoF, is a substrate-binding protein component of the ABC transporter cassette in Agrobacterium tumefaciens involved in importing sulfoquinovose (SQ) and its derivatives for SQ catabolism. Here, we show that SmoF binds with high affinity to the octyl glycoside of SQ (octyl-SQ), demonstrating remarkable tolerance to extension of the anomeric substituent. The 3D X-ray structure of the SmoF·octyl-SQ complex reveals accommodation of the octyl chain, which projects to the protein surface, providing impetus for the synthesis of a linker-equipped SQ-amine using a thiol-ene reaction as a key step, and its conjugation to cyanogen bromide modified agarose. We demonstrate the successful capture and release of SmoF from SQ-agarose resin using SQ as competitive eluant, and selectivity for release versus other organosulfonates. We show that SmoF can be captured and purified from a cell lysate, demonstrating the utility of SQ-agarose in capturing SQ binding proteins from complex mixtures. The present work provides a pathway for development of 'capture-and-release' affinity resins for the discovery and study of SBPs.

Anti-Obesity Effect of Lactobacillus acidophilus DS0079 (YBS1) by Inhibition of Adipocyte Differentiation through Regulation of p38 MAPK/PPARγ Signaling.

Obesity is spawned by an inequality between the portion of energy consumed and the quantity of energy expended. Disease entities such as cardiovascular disease, arteriosclerosis, hypertension, and cancer, which are correlated with obesity, influence society and the economy. Suppression of adipogenesis, the process of white adipocyte generation, remains a promising approach for treating obesity. Oil Red O staining was used to differentiate 3T3-L1 cells for screening 20 distinct Lactobacillus species. Among these, Lactobacillus acidophilus DS0079, referred to as YBS1, was selected for further study. YBS1 therapy decreased 3T3-L1 cell development. Triglyceride accumulation and mRNA expression of the primary adipogenic marker, peroxisome proliferator-activated receptor gamma (PPARγ), including its downstream target genes, adipocyte fatty acid binding protein 4 and adiponectin, were almost eliminated. YBS1 inhibited adipocyte differentiation at the early stage (days 0-2), but no significant difference was noted between the mid-stage (days 2-4) and late-stage (days 4-6) development. YBS1 stimulated the activation of p38 mitogen-activated protein kinase (p38 MAPK) during the early stages of adipogenesis; however, this effect was eliminated by the SB203580 inhibitor. The data showed that YBS1 administration inhibited the initial development of adipocytes via stimulation of the p38 MAPK signaling pathway, which in turn controlled PPARγ expression. In summary, YBS1 has potential efficacy as an anti-obesity supplement and requires further exploration.

2,4'-Dihydroxybenzophenone: A Promising Anti-Inflammatory Agent Targeting Toll-like Receptor 4/Myeloid Differentiation Factor 2-Mediated Mitochondrial Reactive Oxygen Species Production during Lipopolysaccharide-Induced Systemic Inflammation.

The biochemical properties of 2,4'-dihydroxybenzophenone (DHP) have not been extensively studied. Therefore, this study aimed to investigate whether DHP could alleviate inflammatory responses induced by lipopolysaccharide (LPS) and endotoxemia. The results indicated that DHP effectively reduced mortality and morphological abnormalities, restored heart rate, and mitigated macrophage and neutrophil recruitment to inflammatory sites in LPS-microinjected zebrafish larvae. Additionally, the expression of pro-inflammatory mediators, including inducible nitric oxide synthase (iNOS), tumor necrosis factor-α (TNF-α), and interleukin-12 (IL-12), was significantly reduced in the presence of DHP. In RAW 264.7 macrophages, DHP inhibited the LPS-induced inflammatory response by downregulating pro-inflammatory mediators and decreasing the expression of myeloid differentiation primary response 88 (MyD88), phosphorylation of IL-1 receptor-associated protein kinase-4 (p-IRAK4), and nuclear factor-κB (NF-κB). Molecular docking analysis demonstrated that DHP occupies the hydrophobic pocket of myeloid differentiation factor 2 (MD2) and blocks the dimerization of Toll-like receptor 4 (TLR4). A molecular dynamics simulation confirmed that DHP stably bound to the hydrophobic pocket of MD2. Furthermore, the DHP treatment inhibited mitochondrial reactive oxygen species (mtROS) production during the LPS-induced inflammatory response in both RAW 264.7 macrophages and zebrafish larvae, which was accompanied by stabilizing mitochondrial membrane potential. In conclusion, our study highlights the therapeutic potential of DHP in alleviating LPS-induced inflammation and endotoxemia. The findings suggest that DHP exerts its anti-inflammatory effects by inhibiting the TLR4/MD2 signaling pathway and reducing the level of mtROS production. These results contribute to a better understanding of the biochemical properties of DHP and support its further exploration as a potential therapeutic agent for inflammatory conditions and endotoxemia.

2,4'-Dihydroxybenzophenone Exerts Bone Formation and Antiosteoporotic Activity by Stimulating the ß-Catenin Signaling Pathway.

2,4'-Dihydroxybenzophenone (DHP) is an organic compound derived from Garcinia xanthochymus, but there have been no reports on its biochemical functions and bioavailability. In this study, we evaluated whether DHP affects osteoblast differentiation and activation in MC3T3-E1 preosteoblast cells, as well as antiosteoporotic activity in zebrafish larvae. Nontoxic concentrations of DHP-treated MC3T3-E1 preosteoblast cells increased alkaline phosphatase (ALP) activation and mineralization in a concentration-dependent manner, accompanied by higher expression of osteoblast-specific markers, including Runt-related transcription factor 2 (RUNX2), osterix, and ALP. Consistent with the data in MC3T3-E1 preosteoblast cells, DHP upregulated osteoblast-specific marker genes in zebrafish larvae and simultaneously enhanced vertebral formation. We also revealed that DHP increased the phosphorylation of glycogen synthase kinase-3ß (GSK-3ß) at Ser9 and the total expression of ß-catenin in the cytosol and markedly increased the localization of ß-catenin into the nucleus. Furthermore, DHP restored the prednisolone (PDS)-induced marked decrease in ALP activity and mineralization, as well as osteoblast-specific marker expression. In PDS-treated zebrafish, DHP also alleviated PDS-induced osteoporosis by restoring vertebral formation and osteoblast-related gene expression. Taken together, these results suggest that DHP is a potential osteoanabolic candidate for treating osteoporosis by stimulating osteoblast differentiation.

Characterization of Bacillus velezensis TJS119 and its biocontrol potential against insect pathogens.

Introduction: The white-spotted flower chafer (Protaetia brevitarsis seulensis), which is widely distributed in Asian countries, is traditionally used in oriental medicine. However, its larvae are prone to severe damage by green muscardine disease (caused by Metarhizium anisopliae) during breeding. The aim of this study was to characterize Bacillus velezensis TJS119, which has been isolated from freshwater, and investigate its potential as a biocontrol agent against M. anisopliae in insects. Methods: TJS119 was obtained from freshwater samples in the Republic of Korea and was classified as B. velezensis. We evaluated its in vitro antifungal effect, sequenced the bacterial whole genome, mined genes responsible for the synthesis of secondary metabolites, performed secondary metabolite analysis Ultra performance liquid chromatography-mass spectrometry (UPLC-MS/MS), and conducted bioassays for determining green muscardine disease control ability. Results: Bacillus velezensis TJS119 inhibited the mycelial growth of M. anisopliae in vitro. The size of the B. velezensis TJS119 genome was estimated to be 3,890,913 bp with a GC content of 46.67% and 3,750 coding sequences. Biosynthetic gene clusters for secondary metabolites with antifungal activity were identified in the genome. Lipopeptides, including fengycin secreted by TJS119 exhibit antifungal activity. Application of TJS119 for the biocontrol against green muscardine disease increased the viability of white-spotted flower chafer by 94.7% compared to the control. Discussion: These results indicate that B. velezensis TJS119 is a potential biocontrol agent for insect pathogens.

Metabolomics Reveals Lysinibacillus capsici TT41-Induced Metabolic Shifts Enhancing Drought Stress Tolerance in Kimchi Cabbage (Brassica rapa L. subsp. pekinensis).

Climate change has increased variable weather patterns that affect plants. To address these issues, we developed a microbial biocontrol agent against drought stress in kimchi cabbage (Brassica rapa L. subsp. pekinensis). We selected three bacterial strains (Leifsonia sp. CS9, Bacillus toyonensis TSJ7, and Lysinibacillus capsici TT41) because they showed a survival rate of up to 50% and good growth rate when treated with 30% PEG 6000. The three strains were treated with kimchi cabbage to confirm their enhanced drought stress resistance under non-watering conditions. Among the three strains, the TT41 treated group showed a significant increase in various plant parameters compared with the negative control on the 7th day. We performed extensive profiling of primary and secondary metabolites from kimchi cabbage and the TT41 strain. Multivariate and pathway analyses revealed that only the TT41 group clustered with the well-watered group and showed almost the same metabolome on the 7th day. When treated with TT41, lactic acid was identified as an indicator metabolite that significantly improved drought stress tolerance. Furthermore, lactic acid treatment effectively induced drought stress tolerance in kimchi cabbage, similar to that achieved with the TT41 strain.

Correction: Molecular basis of sulfolactate synthesis by sulfolactaldehyde dehydrogenase from Rhizobium leguminosarum.

[This corrects the article DOI: 10.1039/D3SC01594G.].