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A protein modification strategy was developed based on a thiol-yne click reaction using an electron-deficient yne reagent. This approach demonstrated exceptional selectivity towards thiols and exhibited rapid kinetics, resulting in conjugates with superior acid stability. The conjugation of IgG with an indole-derived fluorophore was achieved for the imaging of PD-L1 in cancer cells.
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As a kind of biosurfactants, iturin A has attracted people's wide attentions due to their features of biodegradability, environmentally friendly, etc.; however, high production cost limited its extensive application, and the aim of this research wants to improve iturin A production in Bacillus amyloliquefaciens. Firstly, dual promoter was applied to strengthen iturin A synthetase expression, and its yield was increased to 1.25 g/L. Subsequently, original 5'-UTRs of downstream genes (ituA, ituB, and ituC) in iturin A synthetase cluster were optimized, which significantly increased mRNA secondary stability, and iturin A yield produced by resultant strain HZ-T3 reached 2.32 g/L. Secondly, synthetic pathway of α-glucosidase inhibitor 1-deoxynojirimycin was blocked to improve substrate corn starch utilization, and iturin A yield was increased by 34.91% to 3.13 g/L. Thirdly, efficient precursor (fatty acids, Ser, and Pro) supplies were proven as the critical role in iturin A synthesis, and 5.52 g/L iturin A was attained by resultant strain, through overexpressing yngH, serC, and introducing ocD. Meanwhile, genes responsible for poly-γ-glutamic acid, extracellular polysaccharide, and surfactin syntheses were deleted, which led to a 30.98% increase of iturin A yield. Finally, lipopeptide transporters were screened, and iturin A yield was increased by 17.98% in SwrC overexpression strain, reached 8.53 g/L, which is the highest yield of iturin A ever reported. This study laid a foundation for industrial production and application development of iturin A, and provided the guidance of metabolic engineering breeding for efficient production of other metabolites synthesized by non-ribosomal peptide synthetase. KEY POINTS: ⢠Optimizing 5'-UTR is an effective tactics to regulate synthetase cluster expression. ⢠Blocking 1-DNJ synthesis benefited corn starch utilization and iturin A production. ⢠The iturin A yield attained in this work was the highest yield reported so far.
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Bacillus amyloliquefaciens , Engenharia Metabólica , Tensoativos , Bacillus amyloliquefaciens/genética , Bacillus amyloliquefaciens/metabolismo , Engenharia Metabólica/métodos , Tensoativos/metabolismo , Peptídeos Cíclicos/biossíntese , Peptídeos Cíclicos/genética , Peptídeos Cíclicos/metabolismo , Regiões Promotoras Genéticas , Ligases/genética , Ligases/metabolismoRESUMO
Proteolysis targeting chimeras (PROTACs) have shifted the paradigm for drug development via target protein degradation. However, PROTACs may exhibit systemic toxicity to normal cells due to indiscriminate degradation and the utility of inhibitors as a warhead for protein targeting. Here, we propose a new strategy for developing activatable PROTACs for cell-specific degradation of histone deacetylase (HDAC) with minimal side effects via caging of the warhead. Molecular docking reveals that the hydroxyl group of the HDAC inhibitor is crucial for targeting. An enzyme-activatable PROTAC is designed by caging the hydroxyl group with the substrate for NAD(P)H: quinone oxidoreductase 1 (NQO1) overexpressed in cancer cells. We demonstrate that the caged PROTAC can be converted to its active form in response to NQO1. The enzyme-activatable PROTAC allows the efficient and specific degradation of HDAC6 and exerts antiproliferative activity in NQO1-positive cells. The generalizability of the design is further demonstrated by engineering a H2O2-responsive PROTAC for specific degradation of HDAC6 in cells with elevated H2O2. The strategy of caging the ligand for target proteins would afford a new dimension for developing activatable PROTACs with high specificity and minimal side effects.
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Histona Desacetilases , Quimera de Direcionamento de Proteólise , Peróxido de Hidrogênio , Simulação de Acoplamento Molecular , Proteólise , NADRESUMO
Ulcerative colitis (UC) is a chronic inflammatory disease significantly impacting patients' lives. This study aimed to elucidate the alleviating effect of ethyl acetate extract (TBEA) from Terminalia bellirica fruit on UC and to explore its mechanism. TBEA was the fraction with the best anti-inflammatory activity screened using in vitro anti-inflammatory assays, and HPLC initially characterized its composition. The mice model of ulcerative colitis was established after free drinking of 2.5% dextran sulfate sodium for six days, and the experimental group was treated with 50 mg/kg and 100 mg/kg TBEA for seven days. We found that TBEA significantly alleviated symptoms in UC mice, including a physiologically significant reduction in disease activity index and pathological damage to colonic tissue. TBEA dramatically slowed down oxidative stress and inflammatory process in UC mice, as evidenced by decreasing myeloperoxidase and malondialdehyde activities and increasing glutathione and catalase levels by reducing the concentrations of IL-6, IL-1ß, TNF-α, and NO in UC mice, as well as by regulating key proteins in the IL-6/JAK2/STAT3 pathway. Meanwhile, TBEA maintained intestinal homeostasis by regulating intestinal flora structure. Our study provides new ideas for developing TBEA into a new drug to treat UC.
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Colite Ulcerativa , Colite , Microbioma Gastrointestinal , Terminalia , Animais , Camundongos , Colite Ulcerativa/induzido quimicamente , Colite Ulcerativa/tratamento farmacológico , Colite Ulcerativa/metabolismo , Citocinas/metabolismo , Terminalia/metabolismo , Mediadores da Inflamação/metabolismo , Interleucina-6/metabolismo , Frutas/metabolismo , Colo/metabolismo , Anti-Inflamatórios/uso terapêutico , Sulfato de Dextrana/efeitos adversos , Camundongos Endogâmicos C57BL , Modelos Animais de Doenças , Colite/tratamento farmacológicoRESUMO
This research aimed to investigate natamycin's antifungal effect and its mechanism against the chestnut pathogen Neofusicoccum parvum. Natamycin's inhibitory effects on N. parvum were investigated using a drug-containing plate culture method and an in vivo assay in chestnuts and shell buckets. The antifungal mechanism of action of natamycin on N. parvum was investigated by conducting staining experiments of the fungal cell wall and cell membrane. Natamycin had a minimum inhibitory concentration (MIC) of 100 µg/mL and a minimum fungicidal concentration (MFC) of 200 µg/mL against N. parvum. At five times the MFC, natamycin had a strong antifungal effect on chestnuts in vivo, and it effectively reduced morbidity and extended the storage period. The cell membrane was the primary target of natamycin action against N. parvum. Natamycin inhibits ergosterol synthesis, disrupts cell membranes, and causes intracellular protein, nucleic acid, and other macromolecule leakages. Furthermore, natamycin can cause oxidative damage to the fungus, as evidenced by decreased superoxide dismutase and catalase enzyme activity. Natamycin exerts a strong antifungal effect on the pathogenic fungus N. parvum from chestnuts, mainly through the disruption of fungal cell membranes.
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Ascomicetos , Natamicina , Natamicina/farmacologia , Antifúngicos/farmacologia , Testes de Sensibilidade MicrobianaRESUMO
MicroRNAs (miRNAs) play essential roles in regulating gene expression and cell fate. However, it remains a great challenge to image miRNAs with high accuracy in living cells. Here, we report a novel genetically encoded dual-color light-up RNA sensor for ratiometric imaging of miRNAs using Mango as an internal reference and SRB2 as the sensor module. This genetically encoded sensor is designed by expressing a splittable fusion of the internal reference and sensor module under a single promoter. This design strategy allows synchronous expression of the two modules with negligible interference. Live cell imaging studies reveal that the genetically encoded ratiometric RNA sensor responds specifically to mir-224. Moreover, the sensor-to-Mango fluorescence ratios are linearly correlated with the concentrations of mir-224, confirming their capability of determining mir-224 concentrations in living cells. Our genetically encoded light-up RNA sensor also enables ratiometric imaging of mir-224 in different cell lines. This strategy could provide a versatile approach for ratiometric imaging of intracellular RNAs, affording powerful tools for interrogating RNA functions and abundance in living cells.
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Proteínas Luminescentes/genética , MicroRNAs/química , Imagem Óptica/métodos , RNA/química , Técnicas Biossensoriais , Linhagem Celular , Engenharia Genética/métodos , Humanos , Imagem Molecular/métodosRESUMO
We report herein the synthesis of novel ring-fused pyrazoloamino pyridine/pyrimidine derivatives as potential FAK inhibitors and the evaluation of pharmaceutical activity against five cancer cell lines (MDA-MB-231, BXPC-3, NCI-H1975, DU145 and 786O). Generally, the majority of compounds displayed strong anti-FAK enzymatic potencies (IC50 < 1 nM) and could effectively inhibit several class of cancer cell lines within the concentration of 3 µM in comparison with GSK2256098 as a reference. Among them, compound 4o is considered to be the most effective due to high sensitivity in antiproliferation. In culture, 4o could not only inhibit FAK Y397 phosphorylation in MDA-MB-231 cell line, but also trigger apoptosis in a dose-dependent manner. Furthermore, computational docking analysis also suggested that 4o and TAE-226 displayed the similar interaction with FAK kinase domain.
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Antineoplásicos/farmacologia , Desenho de Fármacos , Quinase 1 de Adesão Focal/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Pirimidinas/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Quinase 1 de Adesão Focal/metabolismo , Humanos , Estrutura Molecular , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Piridinas/química , Pirimidinas/química , Relação Estrutura-AtividadeRESUMO
An effective and practical CuI-catalyzed methodology toward N-alkyl or N-methyl phosphonamides and phosphinamides was herein demonstrated. The transformation took place readily under the oxidative conditions, and plenty of N-alkylated (methylated) amides (30 examples) were successfully furnished in high efficiency (up to 92% yields). Dicumyl peroxide was considered to act either as the oxidant for the alkylation reaction or as methyl donator for the methylation protocol.
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A general and direct N-arylation of sulfonamides and NH-sulfoximines by sodium arylsulfinates through a desulfitative pathway was herein demonstrated. The reaction proceeded with catalytic loadings of Cu(II)-catalysts without any external ligands. And the novel arylation protocol featured for high efficiency (up to 93% yields) and good substituent tolerance (up to 53 examples). Moreover, a plausible reaction mechanism was also discussed.
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Rhodium-catalyzed ortho-amidations of sulfoximines lead to key intermediates for the preparation of thiadiazine 1-oxides. Following a straightforward protocol, a variety of synthetically valuable compounds can be obtained, thus circumventing common multistep approaches towards potentially bioactive products.
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Rhodium-catalyzed directed CH-functionalizations have been used in hydroarylations of heterobicyclic alkenes with NH-sulfoximines. Unexpectedly, the bicyclic framework is retained, resulting in the formation of addition products being attractive intermediates for functionalized molecules that are difficult to prepare by other means.
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Near-infrared (NIR) chromophores with analyte tunable emission and absorption properties are highly desirable for developing activatable fluorescence and photoacoustic (PA) probes for bioimaging and disease diagnosis. Here we engineer a class of new chromophores by extending the π-conjugation system of a xanthene scaffold at position 7 with different electron withdrawing groups. It is demonstrated that these chromophores exhibit pH-dependent transition from a spirocyclic "closed" form to a xanthene "open" form with remarkable changes in spectral properties. We further develop fluorescence and PA probes by caging the NIR xanthene chromophores with a dipeptidyl peptidase 4 (DPPIV) substrate. In vitro and live cell studies show that these probes allow activatable fluorescence and PA detection and imaging of DPPIV activity with high sensitivity, high specificity and fast response. Moreover, these two probes allow high-contrast and highly specific imaging of DPPIV activity in a tumour-bearing mouse model in vivo via systemic administration. This study highlights the potential of a xanthene scaffold as a versatile platform for developing high-contrast fluorescence and PA molecular probes.
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Botryosphaeria dothidea, a notorious plant pathogen, is responsible for causing chestnut rot during postharvest storage. This research aimed to assess the antifungal properties of perilla essential oil (PEO) and perillaldehyde (PAE) against B. dothidea. PEO's and PAE's inhibitory effects on B. dothidea were investigated using an agar dilution method, a fumigation method, and an in vivo assay in chestnuts and shell buckets. Based on the results of gas chromatography-mass spectrometry, it was confirmed that the main component of PEO was elemicin. The antifungal mechanism of PEO and PAE against B. dothidea was investigated by conducting staining experiments of the fungal cell wall and cell membrane. PEO and PAE strongly inhibit the mycelial growth of B. dothidea in a dose-dependent manner. The inhibitory mechanism is mainly related to the destruction of the integrity of the fungal cell wall and plasma membrane. Notably, PEO retains its antifungal efficacy against B. dothidea in chestnuts, effectively prolonging their storage life. These findings indicate that PEO and PAE are nontoxic, eco-friendly botanical fungicides, holding promise for controlling postharvest chestnut rot.
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Here we develop a near infrared (NIR) fluorogenic probe for carbon monoxide (CO) detection and imaging based on palladium-mediated carbonylation using a NIR boron-dipyrromethene difluoride as a fluorophore and tetraethylene glycols as aqueous moieties. The probe is utilized to image exogenous and endogenous CO under different stimulated conditions in live cells.
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Compostos de Boro , Monóxido de Carbono , Paládio , Corantes Fluorescentes , Espectroscopia de Luz Próxima ao InfravermelhoRESUMO
Revitalization by oxygen: A rhodium(III)-catalyzed oxidative CH/NH activation/annulation sequence provided access to a variety of substituted 1,2-benzothiazine derivatives from readily available NH-sulfoximines and alkynes (see scheme; Cp*=C5 Me5 ). The oxidation system consisted of molecular oxygen in combination with a catalytic amount of Fe(OAc)2 .
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Activation and turning CO2 into value added products is a promising orientation to address environmental issues caused by CO2 emission. Currently, electrocatalysis has a potent well-established role for CO2 reduction with fast electron transfer rate; but it is challenged by the poor selectivity and low faradic efficiency. On the other side, biocatalysis, including enzymes and microbes, has been also employed for CO2 conversion to target Cn products with remarkably high selectivity; however, low solubility of CO2 in the liquid reaction phase seriously affects the catalytic efficiency. Therefore, a new synergistic role in bioelectrocatalysis for CO2 reduction is emerging thanks to its outstanding selectivity, high faradic efficiency, and desirable valuable Cn products under mild condition that are surveyed in this review. Herein, we comprehensively discuss the results already obtained for the integration craft of enzymatic-electrocatalysis and microbial-electrocatalysis technologies. In addition, the intrinsic nature of the combination is highly dependent on the electron transfer. Thus, both direct electron transfer and mediated electron transfer routes are modeled and concluded. We also explore the biocompatibility and synergistic effects of electrode materials, which emerge in combination with tuned enzymes and microbes to improve catalytic performance. The system by integrating solar energy driven photo-electrochemical technics with bio-catalysis is further discussed. We finally highlight the significant findings and perspectives that have provided strong foundations for the remarkable development of green and sustainable bioelectrocatalysis for CO2 reduction, and that offer a blueprint for Cn valuable products originate from CO2 under efficient and mild conditions.
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Dióxido de Carbono , Técnicas Eletroquímicas , Transporte de Elétrons , Biocatálise , CatáliseRESUMO
Background: The Chinese pharmacopeia records Terminalia chebula as effective in treating prolonged diarrhea and dysentery, blood in the stool, and prolapse. Modern pharmacological research proves it has multiple pharmacological benefits, including antioxidant, anti-inflammatory, analgesic, hepatoprotective, neuroprotective, and other properties. Objectives: This study aims to clarify the role of Terminalia chebula's ethyl acetate extract (TCEA) on ulcerative colitis (UC) induced by dextran sodium sulfate (DSS) in mice, as well as explore the potential mechanism of action. Materials and methods: The variation of different extracts of T. chebula was detected using the HPLC technique, and the main components in TCEA were identified. DSS was used to establish a mouse model to mimic the physiological state of UC in humans; the alleviating effect of TCEA and positive control 5-ASA on UC mice were evaluated by gavage treatment. Disease progression was assessed by monitoring the mouse's weight change and disease activity index (DAI). The changes in colon tissue were estimated by measuring colon length, HE, and AB-PAS staining and detecting oxidative stress parameters. The results draw from Western blot and real-time PCR showed the TLR4/MyD88/NF-κB pathway may involve in the anti-inflammatory activity of TCEA. Furthermore, the gut flora sequencing technique was employed to monitor the differentiation of intestinal microbiota of mice induced by DSS and TCEA treatment. Results: TCEA significantly lowered DAI scores and inhibited the weight loss and colonic shortening induced by DSS. The colon histomorphology and oxidative stress levels were enhanced after TCEA treatment compared with DSS induced UC group. TCEA attenuated the inflammatory response by regulating TLR4/MyD88/NF-κB pathway activation. Intestinal flora sequencing showed that DSS and TCEA greatly impacted mice's composition and diversity of intestinal microorganisms. But TCEA increased the abundance of Bacteroidetes and decreased the abundance of Firmicutes and Proteobacteria compared with the DSS group, which contributed a lot to returning the intestinal flora to a balanced state. Conclusion: This study confirms the alleviating effect of TCEA on UC and provides new ideas for developing TCEA into a new drug to treat UC.
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An unprecedented strategy to access highly enantioenriched dihydropyrazoles is described. It involves formal [4+1] cycloadditions of in situ-derived azoalkenes and sulfur ylides catalyzed by a chiral copper/Tol-BINAP complex. A variety of synthetically and biologically important dihydropyrazoles have been obtained with high enantioselectivities (up to 97:3 er) in good yields (83-97%).
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Alcenos/química , Compostos Azo/química , Pirazóis/síntese química , Enxofre/química , Catálise , Cristalografia por Raios X , Ciclização , Modelos Moleculares , Estrutura Molecular , Compostos Organometálicos/química , Pirazóis/química , EstereoisomerismoRESUMO
Inorganic arsenic (iAs) is a group 1 carcinogen, and consumption of rice can be a significant pathway of iAs exposure in the food chain. Although there are regulations in place to control iAs for marketed rice in some countries, additional measures are explored to remove arsenic from rice. Due to the surface-bound and soluble nature of iAs, previous studies have shown that it can be removed to a significant extent using different cooking methods. Towards this goal we modified and tested the absorption method in combination with four home-friendly cooking treatments (UA = unwashed and absorbed, WA = washed and absorbed, PSA = pre-soaked and absorbed, and PBA = parboiled and absorbed) using both brown and white rice (3 types each). The nutrient elements were measured using ICP-MS and arsenic speciation was carried out using LC-ICP-MS. Overall, our results show that PBA was the optimum approach assessed, removing 54% and 73% of inorganic arsenic (iAs) for brown and white rice respectively, raising the margin of exposure (MOE) by 3.7 for white rice and 2.2 times for brown rice, thus allowing the consumption of rice more safely for infants, children and adults. Other cooking treatments were effective in reducing the iAs concentration from white rice only. Here we also report changes in selected nutrient elements (P, K, Mg, Zn and Mn) which are relatively abundant in rice. In general, the treatments retained more nutrients in brown rice than white rice. No significant loss of Zn was observed from both rice types and the loss of other nutrients was similar or less than in comparison to reported losses from rice cooked in excess water in the literature. We conclude that PBA is a promising technique and further research is needed by including different regional rice types and water quality levels.
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Arsênio , Oryza , Adulto , Arsênio/análise , Criança , Culinária , Contaminação de Alimentos/análise , Humanos , Lactente , NutrientesRESUMO
Synthesis of ultrathin metal-organic framework (MOF) nanosheets for highly efficient oxygen evolution reaction (OER) is prevalent, but still many challenges remain. Herein, a facile and efficient three-layer method is reported for the synthesis of NiCoFe-based trimetallic MOF nanosheets, which can be directly used for the oxygen evolution reaction in alkaline conditions. The physical characterization and morphology of trimetallic MOF nanosheets were characterized by powder X-ray diffraction (PXRD), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). By optimizing the molar ratio of Ni/Co/Fe atoms, a series of MOFs with different metal proportions were synthesized. Among them, the as-prepared (Ni3Co1)3Fe1-MOF nanosheets can deliver a current density of 10 mA cm-2 at a low overpotential of 245 mV with a small Tafel slope of 50.9 mV dec-1 in an alkaline electrolyte and exhibit excellent stability. More importantly, through the characterization of the intermediates in the OER process, the possible source of the catalytic active species is the electrochemically transformed metal hydroxides and oxyhydroxides.