RESUMO
3D bioprinting has become a flexible technical means used in many fields. Currently, research on 3D bioprinting is mainly focused on the use of mammalian cells to print organ and tissue models, which has greatly promoted progress in the fields of tissue engineering, regenerative medicine, and pharmaceuticals. In recent years, bacterial bioprinting has gradually become a rapidly developing research fields, with a wide range of potential applications in basic research, biomedicine, bioremediation, and other field. Here, this works reviews new research on bacterial bioprinting, and discuss its future research direction.
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Bioimpressão , Animais , Impressão Tridimensional , Engenharia Tecidual , Medicina Regenerativa , Bactérias , Alicerces Teciduais , MamíferosRESUMO
Soy whey is an abundant, nutrient-rich and safe wastewater produced in tofu processing, so it is necessary to valorize it instead of discarding it as sewage. Whether soy whey can be used as a fertilizer substitute for agricultural production is unclear. In this study, the effects of soy whey serving as a nitrogen source to substitute urea on soil NH3 volatilization, dissolved organic matter (DOM) components and cherry tomato qualities were investigated by soil column experiment. Results showed that the soil NH4+-N concentrations and pH values of the 50% soy whey fertilizer combined with 50% urea (50%-SW) and 100% soy whey fertilizer (100%-SW) treatments were lower than those of 100% urea treatment (CKU). Compared with CKU, 50%-SW and 100%-SW treatments increased the abundance of ammonia oxidizing bacteria (AOB) by 6.52-100.89%, protease activity by 66.22-83.78%, the contents of total organic carbon (TOC) by 16.97-35.64%, humification index (HIX) of soil DOM by 13.57-17.99%, and average weight per fruit of cherry tomato by 13.46-18.56%, respectively. Moreover, soy whey as liquid organic fertilizer reduced the soil NH3 volatilization by 18.65-25.27% and the fertilization cost by 25.94-51.87% compared with CKU. This study provides a promising option with economic and environmental benefits for soy whey utilization and cherry tomato production, which contributes to the win-win effectiveness of sustainable production for both the soy products industry and agriculture.
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Solanum lycopersicum , Alimentos de Soja , Solo/química , Amônia/química , Soro do Leite/química , Volatilização , Fertilizantes/análise , Ureia , Frutas/química , Agricultura/métodos , Nitrogênio/análise , Proteínas do Soro do LeiteRESUMO
Genistin is one of the bioactive isoflavone glucosides found in legumes, which have great nutraceutical and pharmaceutical significance. The market available isoflavones are currently produced by direct plant extraction. However, its low abundance in plant and structural complexity hinders access to this phytopharmaceutical via plant extraction or chemical synthesis. Here, the E. coli cell factory for sustainable production of genistin from glycerol was constructed. First, we rebuilt the precursor genistein biosynthesis pathway in E. coli, and its titer was then increased by 668% by identifying rate-limiting steps and applying an artificial protein scaffold system. Then de novo production of genistin from glycerol was achieved by functional screening and introduction of glycosyl-transferases, UDP-glucose pathway and specific genistin efflux pumps, and 48.1 mg/L of genistin was obtained. A further engineered E. coli strain equipped with an improved malonyl-CoA pathway, alternative glycerol-utilization pathways, acetyl-CoA carboxylase (ACC), and CRISPR interference (CRISPRi) mediated regulation produced up to 137.8 mg/L of genistin in shake flask cultures. Finally, 202.7 mg/L genistin was achieved through fed-batch fermentation in a 3-L bioreactor. This study represents the de novo genistin production from glycerol for the first time and will lay the foundation for low-cost microbial production of glucoside isoflavones. In addition, the multiphase workflow may provide a reference for engineering the biosynthetic pathways in other microbial hosts as well, for green manufacturing of complex natural products.
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Escherichia coli , Isoflavonas , Escherichia coli/genética , Escherichia coli/metabolismo , Engenharia Metabólica , Glicerol/metabolismo , Isoflavonas/metabolismo , GlucosídeosRESUMO
PURPOSE: Soy whey is a byproduct generated from the processing of several soybean products. Its valorization has continued to attract significant research interest in recent times due to the nutritional and bioactive potency of its chemical composition. Herein, the neuroprotective potency of a soy whey fermented by Cordyceps militaris SN-18 against hydrogen peroxide (H2O2)-induced oxidative injury in PC12 cells was investigated. METHODS: The phenolic compositions were analyzed by high-performance liquid chromatography. Antioxidant activities were assessed by ABTSâ¢+ scavenging assay, DPPH radical scavenging assay, reducing power assay, and ferric reducing antioxidant power assay. The neuroprotective effects of fermented soy whey (FSW) were investigated based on the oxidative injury model in PC12 cells. RESULTS: FSW possessed higher total phenolic content and antioxidant activities compared with unfermented soy whey (UFSW) and that most of the isoflavone glycosides were hydrolyzed into their corresponding aglycones during fermentation. The extract from FSW exhibited a greater protective effect on PC12 cells against oxidative injury by promoting cell proliferation, restoring cell morphology, inhibiting lactic dehydrogenase leakage, reducing reactive oxygen species levels, and enhancing antioxidant enzyme activities compared with that from UFSW. Additionally, cell apoptosis was significantly inhibited by FSW through down-regulation of caspase-3, caspase-9, and Bax and up-regulation of Bcl-2 and Bcl-xL. S-phase cell arrest was attenuated by FSW through increasing cyclin A, CDK1 and CDK2, and decreasing p21 protein. CONCLUSION: Fermentation with C. militaris SN-18 could significantly improve the bioactivity of soy whey by enhancing the ability of nerve cells to resist oxidative damage.
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Cordyceps , Fármacos Neuroprotetores , Animais , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Apoptose , Cordyceps/metabolismo , Peróxido de Hidrogênio/toxicidade , Fármacos Neuroprotetores/farmacologia , Estresse Oxidativo , Células PC12 , Ratos , Glycine max/metabolismo , Soro do Leite/metabolismoRESUMO
Fermented soybean products have attracted great attention due to their health benefits. In the present study, the hypoxia-injured PC12 cells induced by cobalt chloride (CoCl2) were used to evaluate the neuroprotective potency of tofu fermented by Actinomucor elegans (FT). Results indicated that FT exhibited higher phenolic content and antioxidant activity than tofu. Moreover, most soybean isoflavone glycosides were hydrolyzed into their corresponding aglycones during fermentation. FT demonstrated a significant protective effect on PC12 cells against hypoxic injury by maintaining cell viability, reducing lactic dehydrogenase leakage, and inhibiting oxidative stress. The cell apoptosis was significantly attenuated by the FT through down-regulation of caspase-3, caspases-8, caspase-9, and Bax, and up-regulation of Bcl-2 and Bcl-xL. S-phase cell arrest was significantly inhibited by the FT through increasing cyclin A and decreasing the p21 protein level. Furthermore, treatment with the FT activated autophagy, indicating that autophagy possibly acted as a survival mechanism against CoCl2-induced injury. Overall, FT offered a potential protective effect on nerve cells in vitro against hypoxic damage.
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Cobalto/toxicidade , Mucorales/metabolismo , Fármacos Neuroprotetores/farmacologia , Alimentos de Soja , Animais , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Hipóxia Celular/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Fermentação , Estresse Oxidativo/efeitos dos fármacos , Células PC12 , Fenóis/química , RatosRESUMO
Glutinous rice-based foods have a long history are consumed worldwide. They are also in great demand for the pursuit of novel sensory and natural health benefits. In this study, we developed a novel fermented glutinous rice product with the supplementation of Fu brick tea. Using in vitro antioxidant evaluation and phenolic compounds analysis, fermentation with Fu brick tea increased the total phenolic content and enhanced the antioxidant activity of glutinous rice, including scavenging of 1,1-Diphenyl-2-picryl-hydrazyl (DPPH) radical, 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) radical, and hydroxyl radical, ferric-reducing antioxidant power, and ferric ion reducing power and iron chelating capability. Besides, compared with traditional fermented glutinous rice, this novel functional food exhibited a stronger activity for protecting DNA against hydroxyl radical-induced oxidation damage. Quantitative analysis by HPLC identified 14 compounds covering catechins and phenolic acids, which were considered to be positively related to the enhanced antioxidant capability. Furthermore, we found that 80% ethanol was a suitable extract solvent compared with water, because of its higher extraction efficiency and stronger functional activities. Our results suggested that this novel fermented glutinous rice could serve as a nutraceutical food/ingredient with special sensory and functional activities.
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Antioxidantes/química , Quelantes de Ferro/química , Oryza/química , Fenóis/química , Benzotiazóis/química , Compostos de Bifenilo/química , Catecóis/química , China , Cromatografia Líquida de Alta Pressão , Dano ao DNA/efeitos dos fármacos , Fermentação , Sequestradores de Radicais Livres/química , Humanos , Radical Hidroxila/química , Oxirredução , Picratos/química , Ácidos Sulfônicos/química , Triterpenos/químicaRESUMO
Medium-chain fatty acids (MCFAs, 6-10 carbons) are valuable precursors to many industrial biofuels and chemicals, recently engineered reversal of the ß-oxidation (r-BOX) cycle has been proposed as a potential platform for efficient synthesis of MCFAs. Previous studies have made many exciting achievements on functionally characterizing four core enzymes of this r-BOX cycle. However, the information about bottleneck nodes in this cycle is elusive. Here, a quantitative assessment of the inherent limitations of this cycle was conducted to capitalize on its potential. The selection of the core ß-oxidation reversal enzymes in conjunction with acetyl-CoA synthetase endowed the ability to synthesize about 1g/L MCFAs. Furthermore, a gene dosage experiment was developed to identify two rate-limiting enzymes (acetyl-CoA synthetase and thiolase). The de novo pathway was then separated into two modules at thiolase and MCFA production titer increased to 2.8g/L after evaluating different construct environments. Additionally, the metabolism of host organism was reprogrammed to the desired biochemical product by the clustered regularly interspaced short palindromic repeats interference system, resulted in a final MCFA production of 3.8g/L. These findings described here identified the inherent limitations of r-BOX cycle and further unleashed the lipogenic potential of this cycle, thus paving the way for the development of a bacterial platform for microbial production of high-value oleo-chemicals from low-value carbons in a sustainable and environmentally friendly manner.
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Acetato-CoA Ligase , Proteínas de Escherichia coli , Escherichia coli , Ácidos Graxos , Engenharia Metabólica , Acetato-CoA Ligase/biossíntese , Acetato-CoA Ligase/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/biossíntese , Proteínas de Escherichia coli/genética , Ácidos Graxos/biossíntese , Ácidos Graxos/genética , OxirreduçãoRESUMO
Previous studies have made many exciting achievements on pushing the functional reversal of beta-oxidation cycle (r-BOX) to more widespread adoption for synthesis of a wide variety of fuels and chemicals. However, the redox cofactor requirement for the efficient operation of r-BOX remains unclear. In this work, the metabolic efficiency of r-BOX for medium-chain fatty acid (C6-C10, MCFA) production was optimized by redox cofactor engineering. Stoichiometric analysis of the r-BOX pathway and further experimental examination identified NADH as a crucial determinant of r-BOX process yield. Furthermore, the introduction of formate dehydrogenase from Candida boidinii using fermentative inhibitor byproduct formate as a redox NADH sink improved MCFA titer from initial 1.2g/L to 3.1g/L. Moreover, coupling of increasing the supply of acetyl-CoA with NADH to achieve fermentative redox balance enabled product synthesis at maximum titers. To this end, the acetate re-assimilation pathway was further optimized to increase acetyl-CoA availability associated with the new supply of NADH. It was found that the acetyl-CoA synthetase activity and intracellular ATP levels constrained the activity of acetate re-assimilation pathway, and 4.7g/L of MCFA titer was finally achieved after alleviating these two limiting factors. To the best of our knowledge, this represented the highest titer reported to date. These results demonstrated that the key constraint of r-BOX was redox imbalance and redox engineering could further unleash the lipogenic potential of this cycle. The redox engineering strategies could be applied to acetyl-CoA-derived products or other bio-products requiring multiple redox cofactors for biosynthesis.
Assuntos
Acetilcoenzima A , Candida , Coenzimas , Ácidos Graxos , NADP , Acetilcoenzima A/genética , Acetilcoenzima A/metabolismo , Candida/genética , Candida/metabolismo , Coenzimas/genética , Coenzimas/metabolismo , Ácidos Graxos/biossíntese , Ácidos Graxos/genética , NADP/genética , NADP/metabolismo , OxirreduçãoRESUMO
KEY MESSAGE: Overexpression of OsNRT2.3b in rice can increase Pi uptake and accumulation through advanced root system, enhanced OsPT and OsPHR genes expression, and the phloem pH homeostasis. Nitrogen (N) and phosphorus (P) are two essential macronutrients for plants. Overexpression of the rice nitrate transporter, OsNRT2.3b, can improve rice grain yield and nitrogen use efficiency (NUE). Here, OsNRT2.3b overexpression resulted in increased grain yield, straw yield, and grain:straw ratio, accompanied by increased P concentrations in the leaf blade, leaf sheath, culm, and unfilled rice hulls. Overexpression of OsNRT2.3b significantly increased 33Pi uptake compared with WT under 300-µM Pi but not 10-µM Pi condition in 24 h. Moreover, the OsNRT2.3b-overexpressing rice lines showed increased root and shoot biomass, root:shoot ratio, total root length root surface area and N, P accumulation under 300- and 10-µM Pi supply in hydroponic solution. The levels of OsPT2, OsPT8, and OsPHR2 expression in roots and of OsPT1 and OsPHR2 in shoots were upregulated in OsNRT2.3b-overexpressing rice. These results indicated that OsNRT2.3b overexpression can improve rice P uptake and accumulation, partially through the advanced root system, enhanced gene expression, and the phloem pH regulation function.
Assuntos
Proteínas de Transporte de Ânions/metabolismo , Oryza/metabolismo , Fósforo/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Proteínas de Transporte de Ânions/genética , Transporte Biológico/genética , Transporte Biológico/fisiologia , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Transportadores de Nitrato , Oryza/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/genéticaRESUMO
Plant proteins belonging to the NPF (formerly NRT1/PTR) family are well represented in every genome and function in transporting a wide variety of substrates. In this study, we showed that rice OsNPF2.4 is located in the plasma membrane and is expressed mainly in the epidermis, xylem parenchyma, and phloem companion cells. Functional analysis in oocytes showed that OsNPF2.4 is a pH-dependent, low-affinity NO3â» transporter. Short-term (¹5NO3â») influx rate, long-term NO3â» acquisition by root, and upward transfer from root to shoot were decreased by disruption of OsNPF2.4 and increased by OsNPF2.4 overexpression under high NO3â» supply. Moreover, the redistribution of NO3â» in the mutants in comparison with the wild type from the oldest leaf to other organs, particularly to N-starved roots, was dramatically changed. Knockout of OsNPF2.4 decreased rice growth and potassium (K) concentration in xylem sap, root, culm, and sheath, but increased the shoot:root ratio of tissue K under higher NO3â». We conclude that OsNPF2.4 functions in acquisition and long-distance transport of NO3â» , and that altering its expression has an indirect effect on K recycling between the root and shoot.
Assuntos
Proteínas de Transporte de Ânions/genética , Proteínas de Transporte de Ânions/metabolismo , Regulação da Expressão Gênica de Plantas , Nitratos/metabolismo , Oryza/crescimento & desenvolvimento , Oryza/genética , Transportadores de Nitrato , Oryza/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Potássio/metabolismoRESUMO
The active metabolite (S)-equol, derived from daidzein by gut microbiota, exhibits superior antioxidative activity compared with its precursor and plays a vital role in human health. As only 25% to 50% of individuals can naturally produce equol when supplied with isoflavone, we engineered probiotic E. coli Nissle 1917 (EcN) to convert dietary isoflavones into (S)-equol, thus offering a strategy to mimic the gut phenotype of natural (S)-equol producers. However, co-fermentation of EcN-eq with fecal bacteria revealed that gut microbial metabolites decreased NADPH levels, hindering (S)-equol production. Transcriptome analysis showed that the quorum-sensing (QS) transcription factor SdiA negatively regulates NADPH levels and (S)-equol biosynthesis in EcN-eq. Screening AHLs showed that SdiA binding to C10-HSL negatively regulates the pentose phosphate pathway, reducing intracellular NADPH levels in EcN-eq. Molecular docking and dynamics simulations investigated the structural disparities in complexes formed by C10-HSL with SdiA from EcN or E. coli K12. Substituting sdiA_EcN in EcN-eq with sdiA_K12 increased the intracellular NADPH/NADP+ ratio, enhancing (S)-equol production by 47%. These findings elucidate the impact of AHL-QS in the gut microbiota on EcN NADPH metabolism, offering insights for developing (S)-equol-producing EcN probiotics tailored to the gut environment.
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Medium-chain fatty acids (MCFAs) are essential chemical feedstocks. Microbial production of MCFAs offers an attractive alternative to conventional methods, but the costly media and external inducers limit its practical application. To address this issue and make MCFA production more cost-effective, an E.coli platform was developed using soy whey as a medium and galactose as an autoinducer. We first designed an efficient, stringent, homogeneous, and robust galactose-based autoinduction system for the expression of pathway enzymes by rationally engineering the promoter of the galactose-proton symporter (GalP). Subsequently, the intracellular acetyl-CoA availability and NADH regeneration were enhanced to improve the reversal of the ß-oxidation cycle. The resulting strain yielded 8.20 g/L and 16.42 g/L MCFA in pH-controlled batch fermentation and fed-batch fermentation with glucose added using soy whey as medium, respectively. This study provided a cost-effective and promising platform for MCFA production, as well as future strain development for other value-added chemicals production.
Assuntos
Escherichia coli , Ácidos Graxos , Escherichia coli/metabolismo , Ácidos Graxos/metabolismo , Galactose/metabolismo , Soro do Leite/metabolismo , Análise Custo-Benefício , Engenharia Metabólica/métodos , Proteínas do Soro do Leite/metabolismo , FermentaçãoRESUMO
Daidzein is a major isoflavone compound with an immense pharmaceutical value. This study applied a novel P450 CYP82D26 which can biosynthesize daidzein from (2S)-naringenin. However, the recombinant P450 systems often suffer from low coupling efficiency, leading to an electron transfer efficiency decrease and harmful reactive oxygen species release, thereby compromising their stability and catalytic efficiency. To address these challenges, the SH3-GBD-PDZ (SGP) protein scaffold was applied to assemble a multienzyme system comprising CYP82D26, P450 reductase, and NADP+-dependent aldehyde reductase in desired stoichiometric ratios. Results showed that the coupling efficiency of the P450 system was significantly increased, primarily attributed to the channeling effect of NADPH resulting from the proximity of tethered enzymes and the electrostatic interactions between NADPH and SGP. Assembling this SGP-scaffolded assembly system in Escherichia coli yielded a titer of 240.5 mg/L daidzein with an 86% (2S)-naringenin conversion rate, which showed a 9-fold increase over the free enzymes of the P450 system. These results underscore the potential application of the SGP-scaffolded multienzyme system in enhancing the coupling and catalytic efficiency of the P450 system.
Assuntos
Flavanonas , Isoflavonas , NADPH-Ferri-Hemoproteína Redutase , NADP/metabolismo , NADPH-Ferri-Hemoproteína Redutase/metabolismo , Proteínas , Isoflavonas/metabolismoRESUMO
This study investigated the effect of in situ produced water-soluble α-glucan (LcWSG) and water-insoluble α-glucan (LcWIG) from Leuconostoc citreum SH12 on the physicochemical properties of fermented soymilk. α-Glucans produced by Leuc. citreum SH12 improved water-holding capacity, viscosity, viscoelasticity and texture of fermented soymilk. Gtf1365 and Gtf836 of the five putative glucansucrases were responsible for synthesizing LcWSG and LcWIG during soymilk fermentation, respectively. Co-fermentation of soymilk with Gtf1365 and Gtf836 and non-exopolysaccharide-producing Lactiplantibacillus plantarum D1031 indicated that LcWSG effectively hindered the whey separation of fermented soymilk by increasing viscosity, while LcWIG improved hardness, springiness and accelerated protein coagulation. Fermented soymilk gel formation was mainly based on hydrogen bonding and hydrophobic interactions, which were promoted by both LcWSG and LcWIG. LcWIG has a greater effect on α-helix to ß-sheet translation in fermented soymilk, causing more rapid protein aggregation and thicker cross-linked gel network. Structure-based exploration of LcWSG and LcWIG from Leuc. citreum SH12 revealed their distinct roles in the physicochemical properties of fermented soymilk due to their different ratio of α-1,6 and α-1,3 glucosidic linkages and various side chain length. This study may guide the application of the water-soluble and water-insoluble α-glucans in fermented plant protein foods for their quality improvement.
Assuntos
Fermentação , Glucanos , Leuconostoc , Solubilidade , Leite de Soja , Água , Leuconostoc/metabolismo , Leite de Soja/química , Água/química , Viscosidade , Glucanos/química , Fenômenos QuímicosRESUMO
Calcium-binding peptides have gained significant attention due to their potential applications in various fields. In this study, we aimed to prepare, characterize, and evaluate the stability of calcium-binding peptides derived from chicken blood. Chicken hemoglobin peptides (CPs) were obtained by protease hydrolysis and were applied to prepare chicken hemoglobin peptide-calcium chelate (CP-Ca). The preparation conditions were optimized, and the characteristics and stability of CP-Ca were analyzed. The optimal chelating conditions were determined by single-factor and response surface tests, and the maximum calcium ion chelating rate was 77.54%. Amino acid analysis indicated that glutamic acid and aspartic acid motifs played an important role in the chelation of the calcium ions and CP. According to the characterization analysis, CP-Ca was a different substance compared with CP; calcium ions chelated CPs via the sites of carbonyl oxygen, carboxyl oxygen, and amino nitrogen groups; and after the chelation, the structure changed from a smooth homogeneous plate to compact granular. The stability analysis showed that CP-Ca was stable at different temperatures, pH, and gastrointestinal conditions. The study indicates that chicken blood is a promising source of peptide-calcium chelates, providing a theoretical basis for application in functional foods and improving the utilization value of chicken blood.
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Dietary fibers have attracted much attention due to their multiple benefits on gut health. In this work, the protective mechanism of dietary fiber from sweetpotato residues (SRDF) on the high-fat diet (HFD)-induced intestinal barrier injury was investigated using microbiome-metabolomics-based approach. The physicochemical property analysis demonstrated a thermal stability below 200 °C and porous pectin-polysaccharide structure of SRDF with high in vitro functional activities. The biochemical analysis indicated that SRDF significantly ameliorated intestinal barrier function by improving intestinal morphology and permeability and inhibiting inflammatory response. Microbiome analysis demonstrated that SRDF significantly reversed the HFD-induced dysbacteriosis, decreased the ratio of Firmicutes/Bacteroides and enhanced the relative abundance of probiotics, such as Muribaculaceae and Bifidobacteriaceae. Metabolomics analysis showed that SRDF also significantly altered the metabolic profile in the colon, wherein the differential metabolites were mainly involved in amino acid metabolism (especially tryptophan). Pearson correlation coefficient identified the beneficial relationship between intestinal microbiome and metabolome induced by SRDF. The limitation of this study was that the mouse model may not fully replicate the human intestinal responses due to the difference between the standard environmental conditions and natural world. Generally, our results implied the great potential of SRDF as a functional food ingredient.
Assuntos
Dieta Hiperlipídica , Fibras na Dieta , Microbioma Gastrointestinal , Ipomoea batatas , Metabolômica , Animais , Dieta Hiperlipídica/efeitos adversos , Microbioma Gastrointestinal/efeitos dos fármacos , Ipomoea batatas/química , Fibras na Dieta/farmacologia , Fibras na Dieta/metabolismo , Camundongos , Metabolômica/métodos , Masculino , Metaboloma/efeitos dos fármacos , Intestinos/patologia , Intestinos/microbiologia , Intestinos/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/patologia , Camundongos Endogâmicos C57BLRESUMO
(S)-Equol is one of the most bioactive metabolites of the isoflavones with immense nutritional and pharmaceutical value. Soy whey is the major liquid byproduct of the soy product processing industries that is rich in nutrients and (S)-equol biosynthetic precursor daidzin. However, it is usually disposed into the sewage, causing high environmental contamination. Herein, we constructed a recombinant Escherichia coli for the biosynthesis of (S)-equol from soy whey. First, we evaluated daidzin-specific transporters and optimized the anaerobically induced Pnar in the (S)-equol biosynthesis cassette to produce (S)-equol from daidzin. Then, sucrase and α-galactosidase were co-expressed to confer sucrose, stachyose, and raffinose utilization capacity on E. coli. Meanwhile, EIIBCAglc was inactivated to eliminate the daidzin transport inhibition induced by glucose. Finally, combining these strategies and optimizing the fermentation conditions, the optimal strain produced 91.5 mg/L of (S)-equol with a yield of 0.96 mol/mol substrates in concentrated soy whey. Overall, this new strategy is an attractive route to broaden the applications of soy whey and achieve the eco-friendly production of (S)-equol.
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A nitrate transporter, BcNRT1, was isolated from non-heading Chinese cabbage (Brassica campestris ssp. chinensis Makino) cultivar 'Suzhouqing'. The full-length cDNA was obtained using the rapid amplification of cDNA ends technique and contains an open reading frame of 1,770 bp that predicts a protein of 589 acid residues that possesses 12 putative transmembrane domains. Using the GUS marker gene driven by the BcNRT1 promoter, we found BcNRT1 expression to be concentrated in primary and lateral root tips and in shoots of transgenic Arabidopsis plants. The YFP fused to BcNRT1 and transformed into cabbage protoplasts indicated that BcNRT1 was localized to the plasma membrane. The expression of BcNRT1 in roots was induced by exposure to 25 mM nitrate, and the BcNRT1 cRNA heterologously expressed in Xenopus laevis oocytes showed nitrate conductance when nitrate was included in the medium. Moreover, mutant chl1-5 plants harboring 35S::BcNRT1 showed sensitivity to chlorate treatment and exhibited restored nitrate uptake. In conclusion, the results indicate that BcNRT1 functions as a low affinity nitrate transporter in non-heading Chinese cabbage.
Assuntos
Proteínas de Transporte de Ânions/genética , Proteínas de Transporte de Ânions/metabolismo , Brassica/genética , Brassica/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Sequência de Aminoácidos , Animais , Clonagem Molecular , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Teste de Complementação Genética , Glucuronidase/genética , Glucuronidase/metabolismo , Dados de Sequência Molecular , Transportadores de Nitrato , Nitratos/metabolismo , Nitratos/farmacologia , Oócitos/metabolismo , Filogenia , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas , Transporte Proteico , Protoplastos/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Homologia de Sequência de Aminoácidos , Xenopus laevisRESUMO
Isoflavones are a class of major phenolic compounds, derived from soybeans, that possess unique therapeutic and biological properties. The possible mechanisms of isoflavone-mediated protection of neuronal PC12 cells against hypoxic damage was investigated in this study. Isoflavones showed potential neuroprotective effects by increasing cell viability, decreasing the level of reactive oxygen species (ROS), and inhibiting apoptosis and cell cycle arrest in cobalt chloride (CoCl2)-induced hypoxic damage. A Western blot analysis indicated that isoflavones decreased apoptosis by up-regulating the Bcl-xL protein and down-regulating the Bax protein. They further reduced the S-phase fraction of the cell cycle by down-regulating the p21 protein and up-regulating the cyclin A protein levels. Additionally, isoflavones activated Nrf2 protein translocation and inhibited the p38 MAPK and AKT-mTOR pathways. A molecular docking analysis further revealed that isoflavones displayed a potential competitive interaction with the Nrf2 protein for Keap1. Our findings suggest that isoflavones could be a potent neuroprotective phytochemical in soybeans and their products.
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Soybean whey, as a byproduct of soybean industry, has caused considerable concern recently because of its abundant nutrients. To further utilize soybean whey, it was fermented with Weissella hellenica D1501, and the neuroprotective potency of this beverage was studied in the present work. The phenolic profile and antioxidant capacity of fermented soybean whey (FSBW) were analyzed. The neuroprotective effects were evaluated based on the hydrogen peroxide-stimulated oxidative damage model in a neural-like cell (PC12). Results demonstrated that soybean whey's phenolic contents and antioxidant activities were markedly improved after fermentation. Glycoside isoflavones were efficiently converted into aglycones by W. hellenica D1501. FSBW extract apparently increased cell viability, decreased reactive oxide species levels, and protected antioxidant enzymes in oxidative damage. Furthermore, FSBW effectively reduced apoptosis rate by inhibiting Bax protein and improving Bcl-2 and Bcl-xL proteins. FSBW ameliorated the cell cycle through the decrease of p21 protein and an increase of cyclin A protein. The findings of this study thus suggested that W. hellenica D1501-fermented soybean whey could potentially protect nerve cells against oxidative damage.