Phosphate Synthase Research Articles

Molecular descriptors and in silico studies of 4-((5-(decylthio)-4-methyl-4n-1,2,4-triazol-3-yl)methyl)morpholine as a potential drug for the treatment of fungal pathologies

The article explores the polypharmacological profiling of 4-((5-(decylthio)-4-methyl-4H-1,2,4-triazole-3-yl)methyl)morpholine as a potential antimicrobial agent. The study utilized 15148 electronic pharmacophore models of organisms, ranked by the Tversky index. Detailed analysis revealed classical bonding patterns with selected enzymes, identifying key amino acid residues involved in complex formation. Protein target prediction was conducted through various stages using the Galaxy web service, including ligand structure creation, pharmacophore alignment, and target ranking. The activities of the molecules against 1G6C, 2W6O, 3G7F, 3OWU, 4IVR, and 4TZT proteins were compared. Docking studies with PyMOL and Discovery Studio Visualizer revealed binding to thymidine kinase, thiamine phosphate synthase, and biotin carboxylase with promising binding affinities. These interactions suggest potential antibacterial and antiviral effects, warranting further virtual screening and in-depth studies for the development of effective antimicrobial drugs. Calculations of the molecules were made with the gaussian package program. Calculations were made on the 6-31++g** basis set at B3LYP, HF, and M062X levels with Gaussian software. Afterwards, the 0–100 ns interaction of the molecule with the highest activity was examined.

Hydrogen sulfide alleviates pericarp browning in lichi fruit by modulating energy and sugar metabolisms.

Postharvest litchi is susceptible to browning that limits the development of litchi industry. Hydrogen sulfide (H2S) is an important bioactive molecule that can regulate many physiological processes. This study examined the effects of exogenous H2S on pericarp browning and related physiological mechanisms in postharvest litchi. The results exhibited that exogenous H2S treatment delayed the browning of litchi pericarp and reduced the damage to cell membrane integrity during storage. This treatment inhibited the energy losses of litchi fruit by increasing the activities of H+-ATPase, Ca2+- ATPase, cytochrome C oxidase (CCO) and succinate dehydrogenase (SDH) and regulating the expression of energy metabolism-related genes, including LcAtpB, LcSnRK2, LcAAC1, LcAOX1 and LcUCP1. In addition, H2S treatment increased the levels of fructose, glucose, sucrose, inositol, galactose and sorbose in litchi fruit, and promoted sucrose synthesis by regulating the activities of sucrose phosphate synthase (SPS), sucrose synthase (SS), acid invertase (AI) and neutral invertase (NI). Based on the current findings, we suggest that exogenous H2S enhances the energy supply and antioxidant activity of litchi by modulating energy and sugar metabolism, thereby inhibiting fruit browning and senescence. These results indicated that H2S treatment is an effective approach to maintaining the quality of litchi fruit and extending its shelf life.

The Effect of Pro- and Eukaryotic Inductors on the Synthesis and Antimicrobial Activity of Acinetobacter calcoaceticus IMV B-7241 Surfactants

By now, the mixed cultivation ("co-cultivation") of antimicrobial compound producers with other microorganisms or the introduction of biological inductors in different physiological states (live and inactivated cells, as well as the corresponding supernatants) into the culture medium is a simple, cheap, and effective way to increase the synthesis of practically important microbial metabolites and regulate their biological activity. In most studies, researchers use bacterial strains of various species as inductors, however, in recent years, there have been an increasing number of publications reporting the use of eukaryotic inductors, in response to which there's observed an increase in the synthesis of antimicrobial compounds by the bacterial producers. In addition, the effectiveness of biological inductors depends on the conditions of their cultivation and physiological state. Aim. To study the effect of the methods of preparation and physiological state of biological inductors (gram-negative bacteria Enterobacter cloacae C-8 and yeast Saccharomyces cerevisiae BTM-1) on the activity of biosynthetic enzymes and antimicrobial activity of Acinetobacter calcoaceticus IMV B-7241 surfactants. Methods. Purified glycerol and crude glycerol in equimolar carbon concentration were used as a substrate for cultivation of A. calcoaceticus IMV B-7241. The microbial inductors were grown on both agar and liquid media with glucose as a carbon source. Live or inactivated cells of S. cerevisiae BTM-1 or E. cloacae C-8, as well as the corresponding supernatant, were added to the medium in an amount of 2.5–10 % (v/v). The extracellular surfactants were obtained from the supernatant of the culture liquid by extraction with a mixture of chloroform and methanol (2:1). The antimicrobial activity of surfactants against bacterial (Bacillus subtilis BT-2, Escherichia coli IEM-1, Staphylococcus aureus BMS-1, Pseudomonas sp. MI-2) and yeast (Candida albicans D-6 and Candida tropicalis PE-2) test cultures was determined by the indicator of the minimum inhibitory concentration. The activity of enzymes for the biosynthesis of surface-active glyco- (phosphoenolpyruvate carboxylase, phosphoenolpyruvate synthetase, phosphoenolpyruvate carboxykinase, trehalose phosphate synthase) and aminolipids (NADP+-dependent glutamate dehydrogenase) was analyzed in cell-free extracts obtained after sonication of cells. Results. The introduction into the culture medium of A. calcoaceticus of IMV B-7241 with glycerol of various purification degrees, both pro- and eukaryotic inductors in different physiological states was accompanied by the synthesis of surfactants, the antimicrobial activity of which against the test cultures was higher by one to two orders of magnitude compared to preparations obtained without inductors. It was found that E. cloacae C-8 cells grown in liquid medium were slightly more effective as inductors than those grown in agar medium: the minimum inhibitory concentrations against bacterial and yeast cultures of surfactants synthesised in their presence were 1–6 and 2.5–8 μg/mL, respectively. When live S. cerevisiae BTM-1 or E. cloacae C-8 cells were used as inductors, the production of microbial surfactants with higher antimicrobial activity than those synthesized in the presence of inactivated cells or supernatants was observed: the minimum inhibitory concentrations against the test cultures were in the range of 0.85–16, 2–20 and 1.5–22 μg/mL, respectively. The higher antimicrobial activity of surfactants synthesized in the presence of a pro- or eukaryotic inductor in the medium with purified glycerol may be caused by an increase of aminolipids in their composition, as evidenced by a 1.6–2.1-fold increase in NADP+-dependent glutamate dehydrogenase activity in A. calcoaceticus IMV B-7241 cells compared to the values of cultivation without inductors. The same level of activity of this enzyme during the cultivation of the IMV B-7241 strain in the medium with crude glycerol in the presence of inductors and without them may indicate the synthesis under such conditions of other, than aminolipids, metabolites with antimicrobial activity. Conclusions. As a result of this study, it was established that the antimicrobial activity of A. calcoaceticus IMV B-7241 surfactants can be increased by introducing pro- and eukaryotic inductors in the form of live or inactivated cells, as well as the corresponding supernatants into the medium with glycerol of different degrees of purification.

Conformational Modulation of a Mobile Loop Controls Catalysis in the (βα)8-Barrel Enzyme of Histidine Biosynthesis HisF.

The overall significance of loop motions for enzymatic activity is generally accepted. However, it has largely remained unclear whether and how such motions can control different steps of catalysis. We have studied this problem on the example of the mobile active site β1α1-loop (loop1) of the (βα)8-barrel enzyme HisF, which is the cyclase subunit of imidazole glycerol phosphate synthase. Loop1 variants containing single mutations of conserved amino acids showed drastically reduced rates for the turnover of the substrates N'-[(5'-phosphoribulosyl) formimino]-5-aminoimidazole-4-carboxamide ribonucleotide (PrFAR) and ammonia to the products imidazole glycerol phosphate (ImGP) and 5-aminoimidazole-4-carboxamide-ribotide (AICAR). A comprehensive mechanistic analysis including stopped-flow kinetics, X-ray crystallography, NMR spectroscopy, and molecular dynamics simulations detected three conformations of loop1 (open, detached, closed) whose populations differed between wild-type HisF and functionally affected loop1 variants. Transient stopped-flow kinetic experiments demonstrated that wt-HisF binds PrFAR by an induced-fit mechanism whereas catalytically impaired loop1 variants bind PrFAR by a simple two-state mechanism. Our findings suggest that PrFAR-induced formation of the closed conformation of loop1 brings active site residues in a productive orientation for chemical turnover, which we show to be the rate-limiting step of HisF catalysis. After the cyclase reaction, the closed loop conformation is destabilized, which favors the formation of detached and open conformations and hence facilitates the release of the products ImGP and AICAR. Our data demonstrate how different conformations of active site loops contribute to different catalytic steps, a finding that is presumably of broad relevance for the reaction mechanisms of (βα)8-barrel enzymes and beyond.

The lipid-metabolism enzyme ECI2 reduces neutrophil extracellular traps formation for colorectal cancer suppression

Abnormalities in ether lipid metabolism as well as the formation of neutrophil extracellular traps have recently been recognized as detrimental factors affecting tumorigenesis and progression. However, the role of abnormal ether lipid metabolism in colorectal cancer (CRC) evolution has not been reported. Here we show that the lipid metabolism-related gene enoyl-CoA δ-isomerase 2 (ECI2) plays a tumor-suppressor role in CRC and is negatively associated with poor prognosis in CRC patients. We mechanistically demonstrate that ECI2 reduces ether lipid-mediated Interleukin 8 (IL-8) expression leading to decreased neutrophil recruitment and neutrophil extracellular traps formation for colorectal cancer suppression. In particular, ECI2 inhibits ether lipid production in CRC cells by inhibiting the peroxisomal localization of alkylglycerone phosphate synthase (AGPS), the rate-limiting enzyme for ether lipid synthesis. These findings not only deepen our understanding of the role of metabolic reprogramming and neutrophil interactions in the progression of CRC, but also provide ideas for identifying potential diagnostic markers and therapeutic targets for CRC.

24-epibrassinolide enhances drought tolerance in grapevine (Vitis vinifera L.) by regulating carbon and nitrogen metabolism.

Exogenous application of 24-epibrassinolide can alleviate oxidative damage, improve photosynthetic capacity, and regulate carbon and nitrogen assimilation, thus improving the tolerance of grapevine (Vitis vinifera L.) to drought stress. Brassinosteroids (BRs) are a group of plant steroid hormones in plants and are involved in regulating plant tolerance to drought stress. This study aimed to investigate the regulation effects of BRs on the carbon and nitrogen metabolism in grapevine under drought stress. The results indicated that drought stress led to the accumulation of superoxide radicals and hydrogen peroxide and an increase in lipid peroxidation. A reduction in oxidative damage was observed in EBR-pretreated plants, which was probably due to the improved antioxidant concentration. Moreover, exogenous EBR improved the photosynthetic capacity and sucrose phosphate synthase activity, and decreased the sucrose synthase, acid invertase, and neutral invertase, resulting in improved sucrose (190%) and starch (17%) concentrations. Furthermore, EBR pretreatment strengthened nitrate reduction and ammonium assimilation. A 57% increase in nitrate reductase activity and a 13% increase in glutamine synthetase activity were observed in EBR pretreated grapevines. Meanwhile, EBR pretreated plants accumulated a greater amount of proline, which contributed to osmotic adjustment and ROS scavenging. In summary, exogenous EBR enhanced drought tolerance in grapevines by alleviating oxidative damage and regulating carbon and nitrogen metabolism.

Carbamoyl phosphate synthase 1 (CPS1), a mitochondrial enzyme of the urea cycle, is involved in LDL receptor modulation: In vitro studies

Carbamoyl phosphate synthase 1 (CPS1), a mitochondrial enzyme of the urea cycle, is involved in LDL receptor modulation: In vitro studies

Protein kinase MeSnRK2.3 positively regulates starch biosynthesis by interacting with the transcription factor MebHLH68 in cassava.

Starch biosynthesis involves numerous enzymes and is a crucial metabolic activity in plant storage organs. Sucrose non-fermenting related protein kinase 2 (SnRK2) is an abscisic acid (ABA)-dependent kinase and a significant regulatory enzyme in the ABA signaling pathway. However, whether SnRK2 kinases regulate starch biosynthesis is unclear. In this study, we identified that MeSnRK2.3, an ABA-dependent kinase, was highly expressed in the storage roots of cassava and was induced by ABA. Overexpression of MeSnRK2.3 in cassava significantly increased the starch content in the storage roots and promoted plant growth. MeSnRK2.3 was further found to interact with the cassava basic helix-loop-helix 68 (MebHLH68) transcription factor in vivo and in vitro. MebHLH68 directly bound to the promoters of sucrose synthase 1 (MeSUS1), granule-bound starch synthase I a (MeGBSSIa), and starch-branching enzyme 2.4 (MeSBE2.4), thereby upregulating their transcriptional activities. Additionally, MebHLH68 negatively regulated the transcriptional activity of sucrose phosphate synthase B (MeSPSB). Moreover, phosphorylated MebHLH68 by MeSnRK2.3 up-regulated the transcription activity of MeSBE2.4. These findings demonstrated that the MeSnRK2.3-MebHLH68 module connects the ABA signaling pathway and starch biosynthesis in cassava, thereby providing direct evidence of ABA-mediated participation in the sucrose metabolism and starch biosynthesis pathway.

Sucrose Phosphate Synthase Genes in Plants: Its Role and Practice for Crop Improvement.

Plants convert solar energy and carbon dioxide into organic compounds through photosynthesis. Sucrose is the primary carbonate produced during photosynthesis. Sucrose phosphate synthase (SPS) is the key enzyme controlling sucrose biosynthesis in plants. There are at least three SPS gene families in higher plants, named A, B, and C. However, in monocotyledonous plants from Poaceae, there are at least five SPS gene families, named A, B, C, DIII, and DIV. Each family of SPS genes in different plants shows a divergent expression pattern. So different families of SPS genes participate in diverse biological functions, including sucrose accumulation, plant growth and production, and abiotic stress tolerance. SPS activity in plants is regulated by exogenous factors through gene expression and reversible protein phosphorylation. It is a practicable way to improve crop traits through SPS gene transformation. This work analyzes the cloning, phylogeny, and regulatory mechanism of the SPS gene in plants, reviews its biological function as well as its role in crop improvement, and discusses the challenges and future perspectives. This paper can serve as a reference for further study on plant SPS genes and eventually for crop improvement.

Foliar application of sodium selenite affects the growth, antioxidant system, and fruit quality of strawberry.

Selenium (Se) plays a vital role in various physiological processes in plants and is regarded as an essential micronutrient for human health as well. In this study, sodium selenite solution at 10, 40, 70, and 100 mg·L-1 concentrations was foliar sprayed, and the strawberry plant growth, antioxidant system, and fruit quality with an emphasis on sugar and acid content were assessed. The results showed that 10 mg·L-1 of sodium selenite treatment promoted plant growth, while all the treated concentrations could enhance photosynthesis, the antioxidant system in leaves, the content of Se, and ascorbic acid in fruits. More importantly, 40 mg·L-1 sodium selenite treatment significantly increased fruit weight, total soluble solid, total phenolic content, and anthocyanins, as well as improved the shape index. Furthermore, it decreased the total flavonoid and proanthocyanidin content. Particularly, sodium selenite treatment at 40 and 70 mg·L-1 largely increased the ratio of soluble sugars to titratable acid. The changes of predominant sugars and organic acids during fruit development were further investigated. The sucrose, fructose, and glucose content was upregulated by sodium selenite treatment through upregulating the activities of sucrose phosphate synthase (SPS) and acid invertase, as well as the FaSPS expression. In addition, sodium selenite treatment inhibited the activity of citrate synthase and phosphoenolpyruvate carboxylase, rather than modulating their transcript levels to reduce the citric acid content. This work presented a potentially efficient approach to enhance plant growth and fruit quality and supplement Se in strawberry, providing insights into the mechanism of regulating sugar and acid metabolism by Se.

Contents of paeoniflorin and albiflorin in two Korean landraces of Paeonia lactiflora and characterization of paeoniflorin biosynthesis genes in peony.

Paeoniflorin and albiflorin are monoterpene glycosides that exhibit various medicinal properties in Paeonia species. This study explored the terpene biosynthesis pathway and analyzed the distribution of these compounds in different tissues of two Korean landraces of Paeonia lactiflora to gain insights into the biosynthesis of monoterpene glycosides in P. lactiflora and their potential applications. Two Korean landraces, Hongcheon var. and Hwacheon var, of P. lactiflora were used for the analyses. Contents of the paeoniflorin and albiflorin were analyzed using HPLC. RNA was extracted, sequenced, and subjected to transcriptome analysis. Differential gene expression, KEGG, and GO analyses were performed. Paeoniflorin biosynthesis genes were isolated from the transcriptomes using the genes in Euphorbia maculata with the NBLAST program. Phylogenetic analysis of of 1-Deoxy-D-xylulose 5-phosphate synthase (DOXPS), geranyl pyrophosphate synthase (GPPS), and pinene synthase (PS) was carried out with ClustalW and MEGA v5.0. Analysis of paeoniflorin and albiflorin content in different tissues of the two P. lactiflora landraces revealed significant variation. Transcriptome analysis yielded 36,602 unigenes, most of which were involved in metabolic processes. The DEG analysis revealed tissue-specific expression patterns with correlations between landraces. The isolation of biosynthetic genes identified 173 candidates. Phylogenetic analysis of the key enzymes in these pathways provides insights into their evolutionary relationships. The sequencing and analysis of DOXPS, GPPS, PS revealed distinct clades and subclades, highlighting their evolutionary divergence and functional conservation. Our findings highlight the roots as the primary sites of paeoniflorin and albiflorin accumulation in P. lactiflora, underscoring the importance of tissue-specific gene expression in their biosynthesis. this study advances our understanding of monoterpene glycoside production and distribution in Paeonia, thereby guiding further plant biochemistry investigations.

Cryo-EM structure of 1-deoxy-D-xylulose 5-phosphate synthase DXPS from Plasmodium falciparum reveals a distinct N-terminal domain

Plasmodium falciparum is the main causative agent of malaria, a deadly disease that mainly affects children under five years old. Artemisinin-based combination therapies have been pivotal in controlling the disease, but resistance has arisen in various regions, increasing the risk of treatment failure. The non-mevalonate pathway is essential for the isoprenoid synthesis in Plasmodium and provides several under-explored targets to be used in the discovery of new antimalarials. 1-deoxy-D-xylulose-5-phosphate synthase (DXPS) is the first and rate-limiting enzyme of the pathway. Despite its importance, there are no structures available for any Plasmodium spp., due to the complex sequence which contains large regions of high disorder, making crystallisation a difficult task. In this manuscript, we use cryo-electron microscopy to solve the P. falciparum DXPS structure at a final resolution of 2.42 Å. Overall, the structure resembles other DXPS enzymes but includes a distinct N-terminal domain exclusive to the Plasmodium genus. Mutational studies show that destabilization of the cap domain interface negatively impacts protein stability and activity. Additionally, a density for the co-factor thiamine diphosphate is found in the active site. Our work highlights the potential of cryo-EM to obtain structures of P. falciparum proteins that are unfeasible by means of crystallography.

Resistance to protoporphyrinogen oxidase inhibitors in giant ragweed (Ambrosia trifida).

Giant ragweed (Ambrosia trifida L.) is one of the most troublesome weed species in corn (Zea mays L.) and soybean [Glycine max (L.) Merr.] cropping systems. Following numerous reports in 2018 of suspected herbicide resistance in several Ambrosia trifida populations from Wisconsin, our objective was to characterize the response of these accessions to acetolactate synthase (ALS), enolpyruvyl shikimate phosphate synthase (EPSPS), and protoporphyrinogen oxidase (PPO) inhibitors applied POST. Four accessions (AT1, AT4, AT6, and AT10) exhibited ≥ 50% plant survival after exposure to the cloransulam 3× rate. Two accessions (AT8 and AT10) and one accession (AT2) exhibited ≥ 50% plant survival after exposure to glyphosate and fomesafen 1× rates, respectively. The AT10 accession exhibited multiple resistance to cloransulam and glyphosate. The AT12 accession was 28.8-fold resistant to fomesafen and 3.7-fold resistant to lactofen. A codon change in PPX2 conferring a R98L substitution was identified as the most likely mechanism conferring PPO-inhibitor resistance. To our knowledge, this is the first confirmed case of PPO-inhibitor resistance in Ambrosia trifida globally and we identified the genetic mutation likely conferring resistance. Proactive and diversified integrated weed management strategies are of paramount importance for sustainable long-term Ambrosia trifida management. © 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Effects of Drought Stress on Key Enzymes in Soybean Carbon and Nitrogen Metabolism

Background: The synthesis and catalysis of enzymes are inherently linked to the presence of water and drought-induced water deficit in plants can adversely impact enzyme activity. Drought severely impairs the growth and development of crops, with the extent of its impact varying according to the severity of the water scarcity. Drought also regulates the activity of carbon and nitrogen metabolizing enzymes in plants. Methods: To investigate the effects of varying degrees of drought on soybeans, this study selected HN44 (drought-resistant variety) and HN65 (drought-sensitive variety) as subjects through sand culture and assessed the changes in key enzymes involved in carbon and nitrogen metabolism within the soybean leaves in 2022. Result: The results indicated that short-term drought stress increased the activities of sucrose synthase (SUS), Sucrose Phosphate Synthase (SPS) and Glutamine Synthetase (GS), while decreasing the activity of nitrate reductase (NR). In contrast, prolonged drought reduced the activities of SUS and GS. This study provides a theoretical foundation for enhancing the drought stress resistance of soybeans.

ALA Promotes Sucrose Accumulation in Early Peach Fruit by Regulating SPS Activity.

5-Aminolevulinic acid (ALA), as a novel plant growth regulator, is a critical precursor for the biosynthesis of porphyrin compounds in all organisms. Many studies have reported that exogenous ALA treatment could improve fruit sweetness. However, the mechanism by which ALA promotes the increase in sugar content in fruit remains unclear. In this study, we found that ALA significantly promoted sucrose accumulation and SPS (sucrose phosphate synthase) activity in peach fruit. At 14, 28, 42, 50 and 60 days after ALA treatment, sucrose content of fruit was increased by 23%, 43%, 37%, 40% and 16%, respectively, compared with control treatment, and SPS enzyme activity was increased by 21%, 28%, 47%, 37% and 29%, respectively. Correlation analysis showed that the sucrose content of peach fruit under ALA treatment was significantly positively correlated with SPS activity. Subsequently, bioinformatics was used to identify SPS gene family members in peach fruit, and it was found that there were four members of the PpSPS gene family, distributed on chromosomes 1, 7 and 8, named PpSPS1, PpSPS2, PpSPS3 and PpSPS4, respectively. The results of qRT-PCR showed that PpSPS2 and PpSPS3 were highly expressed in response to ALA during fruit development, and the expression of PpSPS2 was positively correlated with SPS activity and sucrose accumulation in peach fruit. The results of tobacco subcellular localization showed that PpSPS2 was mainly distributed in the cytoplasm and nucleus, while PpSPS3 was mainly distributed in the nucleus. The results of this study will lay the foundation for further study on the functions of PpSPS and the regulation of sugar metabolism during the development and ripening of peach fruit by ALA.

Selenium Nanoparticle and Melatonin Treatments Improve Melon Seedling Growth by Regulating Carbohydrate and Polyamine.

Bio-stimulants, such as selenium nanoparticles and melatonin, regulate melon growth. However, the effects of individual and combined applications of selenium nanoparticles and melatonin on the growth of melon seedlings have not been reported. Here, two melon cultivars were sprayed with selenium nanoparticles, melatonin, and a combined treatment, and physiological and biochemical properties were analyzed. The independent applications of selenium nanoparticles, melatonin, and their combination had no significant effects on the plant heights and stem diameters of Jiashi and Huangmengcui melons. Compared with the controls, both selenium nanoparticle and melatonin treatments increased soluble sugars (6-63%) and sucrose (11-88%) levels, as well as the activity of sucrose phosphate synthase (171-237%) in melon leaves. The phenylalanine ammonia lyase (29-95%), trans cinnamate 4-hydroxylase (32-100%), and 4-coumaric acid CoA ligase (26-113%), as well as mRNA levels, also increased in the phenylpropanoid metabolism pathway. Combining the selenium nanoparticles and melatonin was more effective than either of the single treatments. In addition, the levels of superoxide dismutase (43-130%), catalase (14-43%), ascorbate peroxidase (44-79%), peroxidase (25-149%), and mRNA in melon leaves treated with combined selenium nanoparticles and melatonin were higher than in controls. The results contribute to our understanding of selenium nanoparticles and melatonin as bio-stimulants that improve the melon seedlings' growth by regulating carbohydrate, polyamine, and antioxidant capacities.

Establishment of Physalis alkekengi cell suspension culture: time-dependent behavior of genes related to the steroidal compounds, key enzymes, and physalins under static magnetic field.

Cell suspension culture has the potential to be a valuable source for the bioactive compound productions. In this study, an optimized procedure was established for callus and cell suspension culture of Physalis alkekengi for the first time, and the impact of static magnetic field (SMF, 6 mT) was studied on the high-value metabolic compounds through investigation of signaling molecules and gene expressions at the late log-to-stationary phase. Results showed that the growth regulators of 6-benzyl amino purine (BAP, 1.5mg-1 L) and 1-naphthaleneacetic acid (NAA, 0.4mg-1 L) induced the highest fresh weight, callus rate, callus index, and total withanolides. Cell suspension culture was established in the liquid MS medium supplied with BAP (1.5mg-1 L) and NAA (0.1mg-1 L). SMF application decreased slightly the cell growth and viability and enhanced the number of round-shaped cells. The hydrogen peroxide (H2O2) and nitric oxide (NO) levels increased at an all-time series after SMF exposure, and their maximum contents were observed after 12h. A significant alteration of malondialdehydecontent was also identified after 12h of SMF exposure. The expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR), 1-deoxyD-xylulose 5-phosphate synthase (DXS), squalene synthase (SQS), sterol Δ7-reductase (DWF5), and C-7,8 sterol isomerase (HYD1) genes was upregulated significantly after 24 and 48h. An increase in the total withanolides was related to more activity of HMGR and DXS enzymes in SMF-exposed cells and the maximum physalin A (12.8mgg-1 DW) and physalin B (1.92mgg-1 DW) obtained after 24h compared to controls. Findings suggest that SMF can play a supportive factor in inducing steroidal compounds in P. alkekengi through modulating H2O2 and NO levels and the related-gene expressions.

Reconstruction and identification of the native PLP synthase complex from Methanosarcina acetivorans lysate.

Many protein-protein interactions behave differently in biochemically purified forms as compared to their in vivo states. As such, determining native protein structures may elucidate structural states previously unknown for even well-characterized proteins. Here we apply the bottom-up structural proteomics method, cryoID , toward a model methanogenic archaeon. While they are keystone organisms in the global carbon cycle and active members of the human microbiome, there is a general lack of characterization of methanogen enzyme structure and function. Through the cryoID approach, we successfully reconstructed and identified the native Methanosarcina acetivorans pyridoxal 5'-phosphate (PLP) synthase (PdxS) complex directly from cryogenic electron microscopy (cryoEM) images of fractionated cellular lysate. We found that the native PdxS complex exists as a homo-dodecamer of PdxS subunits, and the previously proposed supracomplex containing both the synthase (PdxS) and glutaminase (PdxT) was not observed in cellular lysate. Our structure shows that the native PdxS monomer fashions a single 8α/8β TIM-barrel domain, surrounded by seven additional helices to mediate solvent and interface contacts. A density is present at the active site in the cryoEM map and is interpreted as ribose 5-phosphate. In addition to being the first reconstruction of the PdxS enzyme from a heterogeneous cellular sample, our results reveal a departure from previously published archaeal PdxS crystal structures, lacking the 37 amino acid insertion present in these prior cases. This study demonstrates the potential of applying the cryoID workflow to capture native structural states at atomic resolution for archaeal systems, for which traditional biochemical sample preparation is nontrivial.

The transcription factor ERF110 promotes cold tolerance by directly regulating sugar and sterol biosynthesis in citrus.

ERFs (ethylene-responsive factors) are known to play a key role in orchestrating cold stress signal transduction. However, the regulatory mechanisms and target genes of most ERFs are far from being well deciphered. In this study, we identified a cold-induced ERF, designated as PtrERF110, from trifoliate orange (Poncirus trifoliata L. Raf., also known as Citrus trifoliata L.), an elite cold-hardy plant. PtrERF110 is a nuclear protein with transcriptional activation activity. Overexpression of PtrERF110 remarkably enhanced cold tolerance in lemon (Citrus limon) and tobacco (Nicotiana tabacum), whereas VIGS (virus-induced gene silencing)-mediated knockdown of PtrERF110 drastically impaired the cold tolerance. RNA sequence analysis revealed that PtrERF110 overexpression resulted in global transcriptional reprogramming of a range of stress-responsive genes. Three of the genes, including PtrERD6L16 (early responsive dehydration 6-like transporters), PtrSPS4 (sucrose phosphate synthase 4), and PtrUGT80B1 (UDP-glucose: sterol glycosyltransferases 80B1), were confirmed as direct targets of PtrERF110. Consistently, PtrERF110-overexpressing plants exhibited higher levels of sugars and sterols compared to their wild type counterparts, whereas the VIGS plants had an opposite trend. Exogenous supply of sucrose restored the cold tolerance of PtrERF110-silencing plants. In addition, knockdown of PtrSPS4, PtrERD6L16, and PtrUGT80B1 substantially impaired the cold tolerance of P. trifoliata. Taken together, our findings indicate that PtrERF110 positively modulates cold tolerance by directly regulating sugar and sterol synthesis through transcriptionally activating PtrERD6L16, PtrSPS4, and PtrUGT80B1. The regulatory modules (ERF110-ERD6L16/SPS4/UGT80B1) unraveled in this study advance our understanding of the molecular mechanisms underlying sugar and sterol accumulation in plants subjected to cold stress.

Genome Sequence Resource of Bacillus mojavensis KRS009, a Rhizospheric Microorganism with Biocontrol Potential

Bacillus mojavensis KRS009 was identified as an antagonistic strain with a strong inhibitory effect on various phytopathogenic fungi. To provide further insight into its biocontrol mechanisms and ability to improve plant salt tolerance, the high quality complete genome of KRS009 was sequenced by PacBio platform and assembled using PacBio-HiFiReads. The KRS009 genome consists of one circular chromosome of 4,089,687 bp with 43.5% GC content and 4,062 open reading frames, of which 3,920 protein-coding genes, 86 transfer RNA, 10 ribosomal RNA, and 26 small RNA were identified in this genome. Among them, genes related to compatible solutes, including treC, galU, proX, and proW, are involved in the synthesis of trehalose, proline, and betaine metabolism and play an important role in relieving osmotic stress. In addition, the KRS009 genome contains genes associated with high salinity tolerance, including those responsible for Na+/H+ antiporters, K+ transporters, TrkH thiamin phosphate synthase, K+-sensing histidine kinase, aryl-phospho-beta-D-glucosidase, flavoprotein CzcO associated with the cation diffusion facilitator CzcD, and chaperonin GroEL. Together, the high-quality genome resource of strain KRS009 would provide a molecular basis for further research on its biocontrol and plant salt tolerance mechanisms. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .