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1.
New Phytol ; 242(1): 154-169, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38375601

RESUMO

Phloem sieve elements (PSE), the primary conduits collaborating with neighboring phloem pole pericycle (PPP) cells to facilitate unloading in Arabidopsis roots, undergo a series of developmental stages before achieving maturation and functionality. However, the mechanism that maintains the proper progression of these differentiation stages remains largely unknown. We identified a gain-of-function mutant altered phloem pole pericycle 1 Dominant (app1D), producing a truncated, nuclear-localized active form of NAC with Transmembrane Motif 1-like (NTL9). This mutation leads to ectopic expression of its downstream target CALLOSE SYNTHASE 8 (CalS8), thereby inducing callose accumulation, impeding SE differentiation, impairing phloem transport, and inhibiting root growth. The app1D phenotype could be reproduced by blocking the symplastic channels of cells within APP1 expression domain in wild-type (WT) roots. The WT APP1 is primarily membrane-tethered and dormant in the root meristem cells but entries into the nucleus in several cells in PPP near the unloading region, and this import is inhibited by blocking the symplastic intercellular transport in differentiating SE. Our results suggest a potential maintenance mechanism involving an APP1-CalS8 module, which induces CalS8 expression and modulates symplastic communication, and the proper activation of this module is crucial for the successful differentiation of SE in the Arabidopsis root.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Glucanos , Glucosiltransferases , Arabidopsis/metabolismo , Floema/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo
2.
J Enzyme Inhib Med Chem ; 38(1): 2163241, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36629443

RESUMO

In this work, a series of novel compounds Spartinin C1-C24 were screened, synthesised and evaluated for inhibiting xanthine oxidase thus lowering serum uric acid level. The backbones were derived from the components of coastal marine source Spartina alterniflora and marketed drugs. The top hits Spartinin C10 & C22 suggested high inhibition percentages (78.54 and 93.74) at 10 µM dosage, which were higher than the positive control Allopurinol. They were low cytotoxic onto human normal hepatocyte cells. Treatment with Spartinin C10 could lower the serum uric acid level to 440.0 µM in the hyperuricemic model mice (723.0 µM), comparable with Allopurinol (325.8 µM). Spartinin C10 was more appreciated than Allopurinol on other serum indexes. The preliminary pharmacokinetics evaluation indicated that the rapid absorption, metabolism and elimination of Spartinin C10 should be further improved. The discovery of pharmaceutical molecules from coastal marine source here might inspire the inter-disciplinary investigations on public health.


Assuntos
Alopurinol , Hiperuricemia , Humanos , Camundongos , Animais , Alopurinol/farmacologia , Alopurinol/uso terapêutico , Ácido Úrico/uso terapêutico , Ácidos Cumáricos , Hiperuricemia/tratamento farmacológico , Xantina Oxidase/metabolismo
3.
Analyst ; 147(15): 3534-3541, 2022 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-35792650

RESUMO

As a precursor of all reactive oxygen species (ROS), superoxide anions play an important role in organisms. However, excessive superoxide anions can cause various diseases. Thus, it is highly urgent to develop efficient tools for in situ superoxide anion detection. In this work, a novel boric acid-based, mitochondria-targeted fluorescent probe Mito-YX for superoxide anion detection was designed by regulating its intramolecular charge transfer (ICT) effect. The probe exhibited turn-on fluorescence enhancement within 4 min of reaction with the superoxide anion. In addition, Mito-YX also exhibited high selectivity and a low detection limit down to 0.24 µM with good mitochondrial targeting characteristics, which provided a necessary basis for in vivo detection of superoxide anions. What is more, Mito-YX was successfully applied for the in situ monitoring of superoxide anions in living MCF-7 cells, RAW 264.7 cells and a mouse model of lung inflammation stimulated by LPS. This work provided an important and promising tool for rapid in situ diagnosis and research of the progression of pneumonia.


Assuntos
Corantes Fluorescentes , Superóxidos , Animais , Corantes Fluorescentes/toxicidade , Humanos , Células MCF-7 , Camundongos , Mitocôndrias , Imagem Óptica
4.
Bioorg Med Chem Lett ; 73: 128907, 2022 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-35902063

RESUMO

In this work, hit compounds Spartinin F1-F20 sharing the Spartina alterniflora-sourced ferulic acid backbone were synthesized and evaluated on inhibiting xanthine oxidase and lowering uric acid level. The top hit Spartinin F2 exhibited inhibition percentages at 10 µM dosage as high as 84.48 (higher than that of the positive control allopurinol) and low cyto-toxicity. Spartinin F2 inferred potential efficiency in lowering the serum UA level (from 631.6 µM to 295.0 µM), which was comparable with allopurinol (to 309.2 µM). Spartinin F2 was also beneficial for other serum indexes. The bioavailability of Spartinin F2 was 63.71% from the preliminary pharmacokinetics test and the molecular docking simulation indicated that except for retaining the hydrogen bonds with the key residues such as THR 1010 and LYS 771, the introduction of the π-sulfur interactions via the sulfonate might also be beneficial for developing more potent XO inhibitors.


Assuntos
Alopurinol , Xantina Oxidase , Alopurinol/química , Alopurinol/farmacologia , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Simulação de Acoplamento Molecular , Relação Estrutura-Atividade , Ácido Úrico , Xantina Oxidase/metabolismo
5.
BMC Med Imaging ; 22(1): 21, 2022 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-35125096

RESUMO

OBJECTIVE: The purpose of this study was to compare imaging features between COVID-19 and mycoplasma pneumonia (MP). MATERIALS AND METHODS: The data of patients with mild COVID-19 and MP who underwent chest computed tomography (CT) examination from February 1, 2020 to April 17, 2020 were retrospectively analyzed. The Pneumonia-CT-LKM-PP model based on a deep learning algorithm was used to automatically quantify the number, volume, and involved lobes of pulmonary lesions, and longitudinal changes in quantitative parameters were assessed in three CT follow-ups. RESULTS: A total of 10 patients with mild COVID-19 and 13 patients with MP were included in this study. There was no difference in lymphocyte counts at baseline between the two groups (1.43 ± 0.45 vs. 1.44 ± 0.50, p = 0.279). C-reactive protein levels were significantly higher in MP group than in COVID-19 group (p < 0.05). The number, volume, and involved lobes of pulmonary lesions reached a peak in 7-14 days in the COVID-19 group, but there was no peak or declining trend over time in the MP group (p < 0.05). CONCLUSION: Based on the longitudinal changes of quantitative CT, pulmonary lesions peaked at 7-14 days in patients with COVID-19, and this may be useful to distinguish COVID-19 from MP and evaluate curative effects and prognosis.


Assuntos
COVID-19/diagnóstico por imagem , Pneumonia por Mycoplasma/diagnóstico por imagem , Tomografia Computadorizada por Raios X , Adulto , Estudos de Avaliação como Assunto , Feminino , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos
6.
Environ Microbiol ; 23(2): 1174-1185, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33215844

RESUMO

Fungi dominated the eukaryotic group in the anaerobic sedimentary environment below the ocean floor where they play an essential ecological role. However, the adaptive mechanism of fungi to these anaerobic environments is still unclear. Here, we reported the anaerobic adaptive mechanism of Schizophyllum commune 20R-7-F01, isolated from deep coal-bearing sediment down to ~2 km below the seafloor, through biochemical, metabolomic and transcriptome analyses. The fungus grows well, but the morphology changes obviously and the fruit body develops incompletely under complete hypoxia. Compared with aerobic conditions, the fungus has enhanced branched-chain amino acid biosynthesis and ethanol fermentation under anaerobic conditions, and genes related to these metabolisms have been significantly up-regulated. Additionally, the fungus shows novel strategies for synthesizing ethanol by utilizing both glycolysis and ethanol fermentation pathways. These findings suggest that the subseafloor fungi may adopt multiple mechanisms to cope with lack of oxygen.


Assuntos
Sedimentos Geológicos/microbiologia , Schizophyllum/isolamento & purificação , Schizophyllum/fisiologia , Água do Mar/microbiologia , Aminoácidos de Cadeia Ramificada/biossíntese , Anaerobiose , Carvão Mineral/análise , Etanol/metabolismo , Fermentação , Regulação Fúngica da Expressão Gênica , Genes Fúngicos/genética , Sedimentos Geológicos/química , Schizophyllum/genética , Schizophyllum/metabolismo , Água do Mar/química
7.
Environ Microbiol ; 23(11): 6940-6952, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34431210

RESUMO

Fungi have been reported to be the dominant eukaryotic group in anoxic sub-seafloor sediments, but how fungi subsist in the anoxic sub-marine sedimental environment is rarely understood. Our previous study demonstrated that the fungus, Schizophyllum commune 20R-7-F01 isolated from a ~2 km sediment below the seafloor, can grow and produce primordia in the complete absence of oxygen with enhanced production of branched-chain amino acids (BCAAs), but the primordia cannot be developed into fruit bodies without oxygen. Here, we present the individual and synergistic effects of oxygen and BCAAs on the fruit-body development of this strain. It was found that the fungus required a minimum oxygen concentration of 0.5% pO2 to generate primordia and 1% pO2 to convert primordia into mature fruit body. However, if BCAAs (20 mM) were added to the medium, the primordium could be developed into fruit body at a lower oxygen concentration up to 0.5% pO2 where genes fst4 and c2h2 playing an important role in compensating oxygen deficiency. Moreover, under hypoxic conditions, the fungus showed an increase in mitochondrial number and initiation of auto-phagocytosis. These findings suggest that the fruit-body formation of S. commune may have multiple mechanisms, including energy and amino acid metabolism in response to oxygen concentrations.


Assuntos
Schizophyllum , Aminoácidos de Cadeia Ramificada , Sedimentos Geológicos , Crescimento e Desenvolvimento , Oxigênio/metabolismo , Schizophyllum/metabolismo
8.
J Ind Microbiol Biotechnol ; 47(8): 573-583, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32885332

RESUMO

γ-Glutamyl compounds have unveiled their importance as active substances or precursors of pharmaceuticals. In this research, an approach for enzymatic synthesis of γ-glutamyl compounds was developed using γ-glutamylmethylamide synthetase (GMAS) from Methylovorus mays and polyphosphate kinase (PPK) from Corynebacterium glutamicum. GMAS and PPK were co-recombined in pETDuet-1 plasmid and co-expressed in E. coli BL21 (DE3), and the enzymatic properties of GMAS and PPK were investigated, respectively. Under the catalysis of the co-expression system, L-theanine was synthesized with 89.8% conversion when the substrate molar ratio of sodium glutamate and ethylamine (1:1.4) and only 2 mM ATP were used. A total of 14 γ-glutamyl compounds were synthesized by this one-pot method and purified by cation exchange resin and isoelectric point crystallization with a yield range from 22.3 to 72.7%. This study provided an efficient approach for the synthesis of γ-glutamyl compounds by GMAS and PPK co-expression system.


Assuntos
Carbono-Nitrogênio Ligases/metabolismo , Corynebacterium glutamicum/enzimologia , Escherichia coli/genética , Glutamatos/biossíntese , Methylophilaceae/enzimologia , Fosfotransferases (Aceptor do Grupo Fosfato)/metabolismo , Carbono-Nitrogênio Ligases/genética , Escherichia coli/enzimologia , Fermentação , Microrganismos Geneticamente Modificados , Ressonância Magnética Nuclear Biomolecular , Fosfotransferases (Aceptor do Grupo Fosfato)/genética
9.
Int J Syst Evol Microbiol ; 68(3): 947-950, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29458543

RESUMO

A Gram-stain-negative, rod-shaped, red-pigmented, aerobic bacterium, strain M2T, was isolated from a seawater sample collected from the western Pacific Ocean at a depth of 1000 m and characterized using polyphasic taxonomy. Strain M2T was catalase-positive and oxidase-negative. Cells grew at 4-33 °C (optimum, 25 °C), at pH 6-9 (optimum, 7) and with 0-4 % (w/v) (optimum, 1 %) NaCl. Phylogenetic trees based on 16S rRNA gene sequences showed that strain M2T was associated with the genus Hymenobacter. Strain M2T showed the highest 16S rRNA gene sequence similarities to Hymenobacter actinosclerus CCUG 39621T (95.7 %), Hymenobacter tibetensis XTM003T (95.6 %) and Hymenobacter psychrotolerans Tibet-IIU11T (95.2 %). The DNA G+C content was 59.98 mol%. Strain M2T contained C16 : 1ω5c (25.0 %), iso-C15 : 0 (23.9 %) and summed feature 3 (C16 : 1ω6c and/or C16 : 1ω7c, 20.4 %) as major cellular fatty acids. The major quinone of strain M2T was menaquinone 7 and the major polar lipid was phosphatidylethanolamine. The major polyamine of strain M2T was sym-homospermidine. The phylogenetic analysis and physiological and biochemical data showed that strain M2T should be classified as representing a novel species of the genus Hymenobacter, for which the name Hymenobacter profundi sp. nov. is proposed. The type strain is M2T (=CCTCC AB 2017185T=KCTC 62120T).

10.
Int J Syst Evol Microbiol ; 68(5): 1776-1780, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29620498

RESUMO

A Gram-stain-negative, aerobic bacterium, strain M5T, was isolated from a seawater sample collected from the western Pacific Ocean at a depth of 1000 m and characterized by using polyphasic taxonomy. Cells of the strain were rod-shaped and motile by a single polar flagellum. Cells grew at 4-40 °C (optimum, 25 °C), at pH 7-10 (optimum, 9) and with 0-10 % NaCl (optimum, 1-2 %). Phylogenetic trees based on 16S rRNA gene sequences showed that strain M5T was associated with the genus Pseudomonas, and showed highest similarities to Pseudomonas pelagia CL-AP6T (97.8 %) and Pseudomonas salina XCD-X85T (97.5 %) and Pseudomonas sabulinigri J64T (96.4 %). The average nucleotide identity scores for strains CL-AP6T and XCD-X85T were 74.6 % and 73.7 %, the Genome-to-Genome Distance Calculator scores were 15.8-19.5 % and 15.4-19.7 %, and the species identification scores were 92.3 % and 92.4 %. The major isoprenoid quinone of strain M5T was ubiquinone (Q-9) and the major cellular fatty acids were summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c; 33.2 %), summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c; 22.8 %) and C16 : 0 (13 %). The major polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phospholipid and some unidentified lipids. The phylogenetic analysis and physiological and biochemical data showed that strain M5T should be classified as representing a novel species in the genus Pseudomonas, for which the name Pseudomonas profundi sp. nov. is proposed. The type strain is M5T (=CCTCC AB 2017186T=KCTC 62119T=CICC 24308T).


Assuntos
Filogenia , Pseudomonas/classificação , Água do Mar/microbiologia , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/química , Oceano Pacífico , Fosfolipídeos/química , Pseudomonas/genética , Pseudomonas/isolamento & purificação , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Ubiquinona/química
11.
Phytopathology ; 108(1): 60-69, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28876208

RESUMO

Small RNA (sRNA) is a class of noncoding RNA that can silence the expression of target genes. In rice, the majority of characterized sRNAs are within the range of 21 to 24 nucleotides (nt) long, whose biogenesis and function are associated with a specific sets of components, such as Dicer-like (OsDCLs) and Argonaute proteins (OsAGOs). Rice sRNAs longer than 24 nt are occasionally reported, with biogenesis and functional mechanism uninvestigated, especially in a context of defense responses against pathogen infection. By using deep sequencing, we identified a group of rice long small interfering RNAs (lsiRNAs) that are within the range of 25 to 40 nt in length. Our results show that some rice lsiRNAs are differentially expressed upon infection of Rhizoctonia solani, the causal agent of the rice sheath blight disease. Bioinformatic analysis and experimental validation indicate that some rice lsiRNAs can target defense-related genes. We further demonstrate that rice lsiRNAs are neither derived from RNA degradation nor originated as secondary small interfering RNAs (siRNAs). Moreover, lsiRNAs require OsDCL4 for biogenesis and OsAGO18 for function. Therefore, our study indicates that rice lsiRNAs are a unique class of endogenous sRNAs produced in rice, which may participate in response against pathogens.


Assuntos
Oryza/genética , Doenças das Plantas/imunologia , Imunidade Vegetal , RNA Interferente Pequeno/genética , Rhizoctonia/fisiologia , Biblioteca Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Oryza/imunologia , Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controle , RNA de Plantas/genética , Análise de Sequência de DNA , Nicotiana/imunologia , Nicotiana/microbiologia
12.
Mar Drugs ; 16(12)2018 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-30486362

RESUMO

The bacterium Alteromonas sp. ML52, isolated from deep-sea water, was found to synthesize an intracellular cold-adapted ß-galactosidase. A novel ß-galactosidase gene from strain ML52, encoding 1058 amino acids residues, was cloned and expressed in Escherichia coli. The enzyme belongs to glycoside hydrolase family 2 and is active as a homotetrameric protein. The recombinant enzyme had maximum activity at 35 °C and pH 8 with a low thermal stability over 30 °C. The enzyme also exhibited a Km of 0.14 mM, a Vmax of 464.7 U/mg and a kcat of 3688.1 S-1 at 35 °C with 2-nitrophenyl-ß-d-galactopyranoside as a substrate. Hydrolysis of lactose assay, performed using milk, indicated that over 90% lactose in milk was hydrolyzed after incubation for 5 h at 25 °C or 24 h at 4 °C and 10 °C, respectively. These properties suggest that recombinant Alteromonas sp. ML52 ß-galactosidase is a potential biocatalyst for the lactose-reduced dairy industry.


Assuntos
Alteromonas/metabolismo , Organismos Aquáticos/metabolismo , Biocatálise , Temperatura Baixa , beta-Galactosidase/metabolismo , Alteromonas/genética , Animais , Organismos Aquáticos/genética , Clonagem Molecular , Indústria de Laticínios/métodos , Ensaios Enzimáticos/métodos , Estabilidade Enzimática , Galactose/metabolismo , Concentração de Íons de Hidrogênio , Lactose/metabolismo , Leite/química , Multimerização Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Especificidade por Substrato , beta-Galactosidase/química , beta-Galactosidase/genética , beta-Galactosidase/isolamento & purificação
13.
Protein Expr Purif ; 140: 8-15, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28757468

RESUMO

Cyclodextrin glycosyltransferase (CGTase) is an enzyme able to convert starch and other substrates into cyclodextrins (CDs). A marine strain Y112 producing α-CGTase was identified as Bacillus agaradhaerens Y112 by physiological and biochemical characterization, and 16S rDNA analysis. The gene coding for α-CGTase was cloned, sequenced and expressed in Escherichia coli BL21 (DE3) cells. Recombinant α-CGTase was purified in one-step chromatographic separation and its purity evaluated by SDS-PAGE, showing the presence of one band with a molecular mass of about 92 kDa. Additionally, enzymatic capability was analyzed by measuring the starch conversion, and resulted in about 45% of CDs obtained after 6 h of cyclodextrin reaction. Of these CDs, mainly α-CD was produced (70% of the total CDs yield), suggesting the potential of this CGTase for industrial applications.


Assuntos
Sequência de Aminoácidos/genética , Bacillus/enzimologia , Glucosiltransferases/isolamento & purificação , Clonagem Molecular , Ciclodextrinas/química , Regulação Enzimológica da Expressão Gênica , Glucosiltransferases/biossíntese , Glucosiltransferases/genética , Amido/química , Especificidade por Substrato
14.
J Sep Sci ; 40(9): 1960-1965, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28304138

RESUMO

The metalloproteinase MP belongs to the serralysin family, which is involved in important functions such as nutrient acquisition and infection pathogenesis. Serralysin proteases in highly purified form are commonly used at the industrial level with several purposes. In this study, we set up an efficient and rapid purification protocol for MP using a p-aminobenzamidine-modified affinity chromatography. The affinity medium was synthesized by using p-aminobenzamidine as affinity ligand immobilized via cyanuric chloride spacer to Sepharose 6B sorbent carrier. According to the adsorption analysis, the dissociation constant Kd and theoretical maximum adsorption Qmax of this medium were 24.2 µg/mL and 24.1 mg/g wet sorbent, respectively. The purity of MP was assessed by a high-performance liquid chromatography on a TSK3000SW column and sodium dodecyl sulfate polyacrylamide gel electrophoresis, revealing values of 98.7 and ∼98%, respectively. The specific activity of purified MP was 95.6 U/mg, which is similar to values obtained through traditional purification protocols. In conclusion, our protocol could be easily employed for the rapid isolation of MP with high purity, and could be implemented for other serralysin family proteases.


Assuntos
Benzamidinas , Metaloendopeptidases/isolamento & purificação , Cromatografia de Afinidade , Eletroforese em Gel de Poliacrilamida
15.
Cell Physiol Biochem ; 39(2): 467-80, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27383216

RESUMO

BACKGROUND/AIMS: To date, several positron emission tomography/computed tomography (PET/CT) radiotracers including fluorine-18 fluorodeoxyglucose (18F-FDG), carbon-11 labeled choline (11C-choline), 18-F fluorocholine (18F-FCH) and carbon-11 acetate (11C-acetate) have already been assessed in the application of prostate cancer (PCa) diagnosis to some extent, the diagnostic efficiency of these radiotracers still remain controversial. As a result of this, we carried out this meta-analysis for the purpose of comparing the diagnostic accuracy among four PET/CT radiotracers. METHODS: A systematical literature search for articles was performed until July 3, 2015. We implemented all analysis using the statistical software of STATA12 and quality assessment was performed using QUADAS-2. RESULTS: A total of 56 studies containing 3,586 patients were included in this meta-analysis. Parameter estimates of the overall analysis are as follows: sensitivity, 0.80 (95% CI: 0.74-0.85); specificity, 0.84 (95% CI: 0.77-0.89) and area under roc curve-AUC of SROC, 0.89 (95% CI: 0.86-0.91), indicating a relatively high level of accuracy in diagnosis of PCa. When different radiotracers of PET/CT were compared, 18F-FCH-PET/CT was ranked as the most favorable with the highest value of AUC (AUC = 0.94; 95% CI: 0.92-0.96) whereas 18F-FDG was the least favorable (AUC = 0.73, 95% CI: 0.69-0.77). CONCLUSION: This study suggested that PET/CT imaging plays an invaluable role in the diagnosis of PCa and 18F-FCH-PET/CT was considered as a superior diagnostic tool over other radiotracers. More attention should be paid to the diagnostic efficiency of the four radiotracers particularly for PCa patients with different clinical stages.


Assuntos
Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Neoplasias da Próstata/diagnóstico por imagem , Neoplasias da Próstata/metabolismo , Compostos Radiofarmacêuticos/farmacocinética , Acetatos/farmacocinética , Radioisótopos de Carbono/farmacocinética , Colina/análogos & derivados , Colina/farmacocinética , Radioisótopos de Flúor/farmacocinética , Fluordesoxiglucose F18/farmacocinética , Humanos , Masculino , Curva ROC
16.
Appl Microbiol Biotechnol ; 100(4): 1691-1700, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26476652

RESUMO

Tyramine has been paid more attention in recent years as a significant metabolite of tyrosine and catecholamine drug and an intermediate of medicinal material and some drugs. In this study, an effective, green, and three-step biocatalytic synthesis method for production of tyramine starting from serine in keratin acid hydrolysis wastewater was developed and investigated. Serine deaminase from Escherichia coli was first combined with tyrosine phenol-lyase from Citrobacter koseri, to convert L-serine to L-tyrosine. L-Tyrosine can then be decarboxylated to tyramine by tyrosinede carboxylase from Lactobacillus brevis. All these enzymes originated from recombinant whole cells. Serine deaminaseand tyrosine phenol-lyase could efficiently convert L-serine in wastewater to L-tyrosine at pH 8.0, 37 °C, and Triton X-100 of 0.04% when tyrosine phenol-lyase and its corresponding substrates were sequentially added. Tyrosine conversion rate reached 98 % by L-tyrosine decarboxylase. In scale-up study, the conversion yield of L-serine in wastewater to tyrosine was up to 89 %. L-Tyrosine was decarboxylated to tyramine with a high yield 94 %. Tyramine hydrochloride was obtained with a total yield 84 %. This study has provided an efficient way of recycling keratin acid hydrolysis wastewater to produce tyramine.


Assuntos
Enzimas/metabolismo , Serina/metabolismo , Tiramina/metabolismo , Ácidos , Citrobacter koseri/enzimologia , Citrobacter koseri/genética , Enzimas/genética , Escherichia coli/enzimologia , Escherichia coli/genética , Concentração de Íons de Hidrogênio , Hidrólise , Queratinas/metabolismo , Levilactobacillus brevis/enzimologia , Levilactobacillus brevis/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Temperatura , Águas Residuárias
17.
J Sep Sci ; 39(11): 2050-6, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27058973

RESUMO

In this study, an efficient affinity purification protocol for an alkaline metalloprotease from marine bacterium was developed using immobilized metal affinity chromatography. After screening and optimization of the affinity ligands and spacer arm lengths, Cu-iminmodiacetic acid was chosen as the optimal affinity ligand, which was coupled to Sepharose 6B via a 14-atom spacer arm. The absorption analysis of this medium revealed a desorption constant Kd of 21.5 µg/mL and a theoretical maximum absorption Qmax of 24.9 mg/g. Thanks to this affinity medium, the enzyme could be purified by only one affinity purification step with a purity of approximately 95% pure when analyzed by high-performance liquid chromatography and reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis. The recovery of the protease activity reached 74.6%, which is much higher than the value obtained by traditional protocols (8.9%). These results contribute to the industrial purifications and contribute a significant reference for the purification of other metalloproteases.


Assuntos
Cromatografia de Afinidade , Flavobacterium/enzimologia , Metaloproteases/isolamento & purificação , Metaloproteases/metabolismo
18.
Proc Natl Acad Sci U S A ; 110(22): 9171-6, 2013 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-23686579

RESUMO

Owing to their sessile nature, plants have evolved sophisticated genetic and epigenetic regulatory systems to respond quickly and reversibly to daily and seasonal temperature changes. However, our knowledge of how plants sense and respond to warming ambient temperatures is rather limited. Here we show that an increase in growth temperature from 22 °C to 30 °C effectively inhibited transgene-induced posttranscriptional gene silencing (PTGS) in Arabidopsis. Interestingly, warmth-induced PTGS release exhibited transgenerational epigenetic inheritance. We discovered that the warmth-induced PTGS release occurred during a critical step that leads to the formation of double-stranded RNA (dsRNA) for producing small interfering RNAs (siRNAs). Deep sequencing of small RNAs and RNA blot analysis indicated that the 22-30 °C increase resulted in a significant reduction in the abundance of many trans-acting siRNAs that require dsRNA for biogenesis. We discovered that the temperature increase reduced the protein abundance of SUPPRESSOR OF GENE SILENCING 3, as a consequence, attenuating the formation of stable dsRNAs required for siRNA biogenesis. Importantly, SUPPRESSOR OF GENE SILENCING 3 overexpression released the warmth-triggered inhibition of siRNA biogenesis and reduced the transgenerational epigenetic memory. Thus, our study reveals a previously undescribed association between warming temperatures, an epigenetic system, and siRNA biogenesis.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Epigênese Genética/fisiologia , Regulação da Expressão Gênica de Plantas/fisiologia , Interferência de RNA/fisiologia , RNA Interferente Pequeno/biossíntese , Temperatura , Sequência de Bases , Sequenciamento de Nucleotídeos em Larga Escala , Dados de Sequência Molecular , Plantas Geneticamente Modificadas , Proteínas Quinases/metabolismo , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de DNA
19.
Exp Eye Res ; 122: 1-8, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24607444

RESUMO

MiR-21 acts as a ubiquitous oncogene in major classes of human cancers and is a potential target for therapeutic intervention. However, the relative expression of miR-21 in retinoblastoma is poorly understood. Here we detected miR-21 expression in HXO-RB44 cell line human normal retinal tissues and retinoblastoma (Rb) tissue specimens, and studied its function using an 8-mer tiny seed-targeting anti-miR-21 (t-anti-miR-21). RT-PCR revealed that miR-21 was highly overexpressed in HXO-RB44 cells and Rb tissue specimens compared with normal human retinal tissues. The localization and transfection efficiency of t-anti-miR-21 and the cell cycle distribution were detected by confocal microscopy and flow cytometry. In addition, we found that t-anti-miR-21 led to a significant inhibition of retinoblastoma cell proliferation, migration and colony formation in vitro, with a similar effect to anti-miR-21. Anti-miR-21 down-regulated the miR-21 level, whereas both 8-mer t-anti-miR-21 and 15-mer m-anti-miR-21 had no impact on miR-21 expression levels. Finally, the phosphorylation signaling pathway, down-regulated by t-anti-miR-21, was integrated by KEGG assay, which elucidated the potential mechanisms of inhibition of miR-21 in retinoblastoma. Taken together, knockdown of miR-21 in the HXO-RB44 cell is capable of inhibiting cancer progression in retinoblastoma. Seed-targeting t-anti-miR-21 was a novel strategy for mir-21-based therapeutics and drug discovery.


Assuntos
MicroRNAs/genética , Terapia de Alvo Molecular , Oligonucleotídeos Antissenso/genética , Neoplasias da Retina/prevenção & controle , Retinoblastoma/prevenção & controle , Transdução de Sinais/fisiologia , Apoptose , Proteínas de Ciclo Celular/metabolismo , Movimento Celular , Proliferação de Células , Progressão da Doença , Regulação para Baixo , Citometria de Fluxo , Regulação Neoplásica da Expressão Gênica/fisiologia , Técnicas de Silenciamento de Genes , Humanos , MicroRNAs/metabolismo , Microscopia Confocal , Proteínas de Neoplasias/metabolismo , Fosforilação , Análise Serial de Proteínas , Reação em Cadeia da Polimerase em Tempo Real , Retina/metabolismo , Neoplasias da Retina/genética , Neoplasias da Retina/metabolismo , Neoplasias da Retina/patologia , Retinoblastoma/genética , Retinoblastoma/metabolismo , Retinoblastoma/patologia , Transfecção , Células Tumorais Cultivadas
20.
J Cardiothorac Surg ; 19(1): 35, 2024 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-38297385

RESUMO

BACKGROUND: With the implementation of lung cancer screening programs, an increasing number of pulmonary nodules have been detected.Video-assisted thoracoscopic surgery (VATS) could provide adequate tissue specimens for pathological analysis, and has few postoperative complications.However, locating the nodules intraoperatively by palpation can be difficult for thoracic surgeons. The preoperative pulmonary nodule localization technique is a very effective method.We compared the safety and effectiveness of two methods for the preoperative localization of pulmonary ground glass nodules. METHODS: From October 2020 to April 2021, 133 patients who underwent CT-guided single pulmonary nodule localization were retrospectively reviewed. All patients underwent video-assisted thoracoscopic surgery (VATS) after successful localization. Statistical analysis was used to evaluate the localization accuracy, safety, information related to surgery and postoperative pathology information. The aim of this study was to evaluate the clinical effects of the two localization needles. RESULTS: The mean maximal transverse nodule diameters in the four-hook needle and hook wire groups were 8.97 ± 3.85 mm and 9.00 ± 3.19 mm, respectively (P = 0.967). The localization times in the four-hook needle and hook wire groups were 20.58 ± 2.65 min and 21.43 ± 3.06 min, respectively (P = 0.09). The dislodgement rate was significantly higher in the hook wire group than in the four-hook needle group (7.46% vs. 0, P = 0.024). The mean patient pain scores based on the visual analog scale in the four-hook needle and hook wire groups were 2.87 ± 0.67 and 6.10 ± 2.39, respectively (P = 0.000). All ground glass nodules (GGNs) were successfully resected by VATS. CONCLUSIONS: Preoperative pulmonary nodule localization with both a four-hook needle and hook wire is safe, convenient and effective.


Assuntos
Neoplasias Pulmonares , Nódulos Pulmonares Múltiplos , Nódulo Pulmonar Solitário , Humanos , Neoplasias Pulmonares/diagnóstico por imagem , Neoplasias Pulmonares/cirurgia , Neoplasias Pulmonares/patologia , Estudos Retrospectivos , Detecção Precoce de Câncer , Nódulos Pulmonares Múltiplos/diagnóstico por imagem , Nódulos Pulmonares Múltiplos/cirurgia , Nódulos Pulmonares Múltiplos/patologia , Nódulo Pulmonar Solitário/diagnóstico por imagem , Nódulo Pulmonar Solitário/cirurgia , Nódulo Pulmonar Solitário/patologia , Cirurgia Torácica Vídeoassistida/métodos
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