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1.
Funct Integr Genomics ; 23(3): 253, 2023 Jul 25.
Artículo en Inglés | MEDLINE | ID: mdl-37488420

RESUMEN

Highland barley (HB) is an important cereal crop distributed in the plateau region. Bioactive peptides (BAPs) derived from cereal proteins have shown biological functions. However, the knowledge of highland barley peptide (HBP) is limited. This study aims to explore the immunomodulatory activity of HBP and the relationship between immunomodulatory activity and related gene expression through RNA-seq. Firstly, HBP is isolated from protease hydrolysates of HB protein, yielding 12.04% of crude HB protein. The molecular weight of HBP is about 1702 Da analyzed by gel filtration chromatography, and HBP has a specific amino acid sequence as Gln-Pro-Gln-Gln-Pro-Phe-Pro-Gln (QPQPFPQ) analyzed by LC-MS. Besides, HBP contains 42.20% hydrophobic amino acids and 10.86% basic amino acids. Next, the immunomodulatory activity of HBP in vitro shows that HBP enhances the phagocytosis of RAW264.7 macrophages, promotes nitric oxide (NO) production and the mRNA expression of pro-inflammatory genes including tumor necrosis factor α (TNF-α), interleukin 1ß (IL-1ß), and inducible nitric oxide synthase (iNOS), and decreases the mRNA expression of anti-inflammatory gene, transforming growth factor ß1 (TGF-ß1). RNA-seq analysis reveals TNF and nuclear factor kappa B (NF-κB) pathways are upregulated, and RT-qPCR is performed to verify RNA-seq analysis. In conclusion, HBP activates RAW264.7 macrophages via TNF/NF-κB signaling pathway. HBP, as a significant immunomodulatory peptide, might be a promising resource for future functional foods.


Asunto(s)
Hordeum , FN-kappa B , RNA-Seq , Transducción de Señal , Péptidos , Macrófagos , ARN Mensajero
2.
Ecotoxicol Environ Saf ; 249: 114459, 2023 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-38321678

RESUMEN

The investigation of the toxic effects of cadmium (Cd) on rice field invertebrates has attracted accumulating attention. Spider grants a novel insight into the impacts of Cd stress on invertebrates, but the effects of Cd-induced toxicity and molecular response mechanism of related metabolites in spider's egg sacs remain elusive. This investigation found that Cd stress distinctively decreased vitellogenin (Vg) content and hatched spiderlings numbers in the egg sac of Pardosa pseudoannulata. In addition, Cd stress exerted oxidative stress in the egg sac, manifested as the increase of superoxide dismutase and malondialdehyde levels. Further results showed that Cd exposure could affect egg sacs' energy metabolism, including protein and lipid contents. Metabolome analysis generated 73 up-regulated and 63 down-regulated differentially expressed metabolites (DEMs), mainly affecting phenylalanine metabolism, alpha-linolenic acid metabolism, pentose phosphate pathway, and biosynthesis of amino acids. Specifically, pathway analysis showed that Cd exposure down-regulated several key factors, including tyrosine, L-phenylalanine, O-phospho-L-serine, and L-cystathionine, and inhibited the metabolism of amino acids in the egg sacs. The subsequent correlation analysis found that three metabolite indicators, 9-Oxo-ODE, PG (17:0/18:2), and PE (17:0/20:5), were the dominant contributors to the egg sec's properties (i.e., Vg content and gained spiderlings). Collectively, this study hopes to provide valuable data for the protection of rice field spiders and offer novel perspectives for Cd pollution assessment and management.


Asunto(s)
Animales Ponzoñosos , Oryza , Arañas , Animales , Transcriptoma , Cadmio/toxicidad , Metaboloma , Aminoácidos
3.
Molecules ; 28(7)2023 Apr 06.
Artículo en Inglés | MEDLINE | ID: mdl-37050035

RESUMEN

The aim of this study was to evaluate the application potential of a recombinant fungal immunomodulatory protein from Ganoderma lucidum (rFIP-glu). First, a recombinant plasmid pPIC9K::FIP-glu-His was transferred into Pichia pastoris for the production of protein. The protein was then to assess its free radical scavenging abilities and the effect on the viability of both human immortalized keratinocytes (HaCaT cells) and mouse B16-F10 melanoma cells (B16 cells) in vitro, followed by the effect on the melanin synthesis of B16 cells. The results of SDS-PAGE and western blot showed that rFIP-glu was successfully expressed. Furtherly, a bioactivity assay in vitro indicated that the scavenging rate of 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals reached 84.5% at 6.0 mg/mL (p ≤ 0.0001) of rFIP-glu, showing strong antioxidant activity. Subsequently, a safety evaluation demonstrated that rFIP-glu promoted the proliferation of HaCaT cells, with the cell viability reaching 124.3% at 48 µg/mL (p ≤ 0.01), regarding the cell viability of B16 cells after exposure to rFIP-glu (48 µg/mL) significantly inhibited, to 80.7% (p ≤ 0.01). Besides, rFIP-glu inhibited the melanin synthesis of B16 cells in a dose-dependent manner from 100-1000 µg/mL, and rFIP-glu at 500 µg/mL (p ≤ 0.01) exhibited the highest intracellular melanin amount reduction of 16.8%. Furthermore, a mechanism analysis showed that rFIP-glu inhibited tyrosinase (TYR) activity by up-regulating the expression of the microphthalmia-associated transcription factor (MITF) and down-regulating the gene expression of TYR and tyrosinase-related protein-1 (TYRP-1), thus inhibiting melanin synthesis. The data implied that rFIP-glu had significant antioxidant activity and whitening potency. It should be used as raw materials for cosmeceutical applications.


Asunto(s)
Ganoderma , Melanoma Experimental , Reishi , Animales , Ratones , Humanos , Ganoderma/metabolismo , Melaninas/metabolismo , Antioxidantes/metabolismo , Proteínas Recombinantes/metabolismo , Reishi/metabolismo , Monofenol Monooxigenasa/genética , Monofenol Monooxigenasa/metabolismo , Melanoma Experimental/tratamiento farmacológico , Línea Celular Tumoral
4.
Molecules ; 27(19)2022 Oct 04.
Artículo en Inglés | MEDLINE | ID: mdl-36235111

RESUMEN

Chinese Cordyceps is a valuable source of natural products with various therapeutic effects. It is rich in various active components, of which adenosine, cordycepin and polysaccharides have been confirmed with significant immunomodulatory and antitumor functions. However, the underlying antitumor mechanism remains poorly understood. In this review, we summarized and analyzed the chemical characteristics of the main components and their pharmacological effects and mechanism on immunomodulatory and antitumor functions. The analysis revealed that Chinese Cordyceps promotes immune cells' antitumor function by via upregulating immune responses and downregulating immunosuppression in the tumor microenvironment and resetting the immune cells' phenotype. Moreover, Chinese Cordyceps can inhibit the growth and metastasis of tumor cells by death (including apoptosis and autophagy) induction, cell-cycle arrest, and angiogenesis inhibition. Recent evidence has revealed that the signal pathways of mitogen-activated protein kinases (MAPKs), nuclear factor kappaB (NF-κB), cysteine-aspartic proteases (caspases) and serine/threonine kinase Akt were involved in the antitumor mechanisms. In conclusion, Chinese Cordyceps, one type of magic mushroom, can be potentially developed as immunomodulator and anticancer therapeutic agents.


Asunto(s)
Antineoplásicos , Productos Biológicos , Cordyceps , Adenosina/metabolismo , Antineoplásicos/metabolismo , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Productos Biológicos/metabolismo , Productos Biológicos/farmacología , Productos Biológicos/uso terapéutico , Caspasas/metabolismo , China , Cordyceps/metabolismo , Cisteína/metabolismo , Factores Inmunológicos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Serina/metabolismo
5.
Prep Biochem Biotechnol ; 50(4): 357-364, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-31846385

RESUMEN

In order to obtain a better fermentation parameter for the production of recombinant Ganoderma lucidum immunomodulatory protein (rFIP-glu), an engineered Pichia pastoris GS115 was investigated on the fermentation time, temperature, methanol concentration and initial pH of media, while immunomodulatory activities of the rFIP-glu was confirmed. L9(33) orthogonal experiment were firstly employed to optimize various fermentation parameters in the shake-flask level. The optimized fermentation parameters were subsequently verified in a 5 L fermenter. Biological activities including cell viability and tumor necrosis factor-alpha (TNF-α) mRNA of the rFIP-glu were evaluated on murine macrophage RAW264.7 cells. The results showed that the yield of rFIP-glu was up to 368.71 µg/ml in the shake-flask, and 613.47 µg/ml in the 5 L fermenter, when the Pichia pastoris was incubated in basic media with the methanol concentration 1.0% and initial pH 6.5, and with constant shaking at 280 rpm for 4 days at 26 °C. In vitro assays of biological activity indicated that rFIP-glu had significant toxicity against RAW264.7 cells, and possessed the ability to induce TNF-α mRNA expression in macrophage RAW264.7 cells. In conclusion, engineered P. pastoris showed a good fermentation property under the optimum fermentation parameters. It could be a candidate industrial strain for further study.


Asunto(s)
Reactores Biológicos , Proteínas Fúngicas/biosíntesis , Pichia/metabolismo , Proteínas Recombinantes/biosíntesis , Animales , Fermentación , Proteínas Fúngicas/toxicidad , Ratones , Células RAW 264.7 , Proteínas Recombinantes/toxicidad , Reishi/química , Factor de Necrosis Tumoral alfa/metabolismo
6.
J Cell Physiol ; 234(12): 23337-23348, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31148200

RESUMEN

Ganoderma lucidum immunomodulatory protein (FIP-glu) is an active ingredient with potential immunoregulatory functions. The study was conducted to explore the immunomodulatory activities of recombinant FIP-glu (rFIP-glu) and its possible mechanism in macrophage RAW264.7 cells. In vitro assays of biological activity indicated that rFIP-glu significantly activated RAW264.7 cells and possessed proinflammatory and anti-inflammatory abilities. RNA sequencing analysis and Western blot analysis showed that macrophage activation involved PI3K/Akt and MAPK pathways. Furthermore, real-time quantitative polymerase chain reaction indicated that the PI3K inhibitor LY294002 blocked the messenger RNA (mRNA) levels of MCP-1 (CCL-2), the MEK1/2 inhibitor U0126 reduced the mRNA levels of TNF-α and MCP-1 (CCL-2), and the JNK1/2/3 inhibitor SP600125 prevented the upregulation of inducible nitric oxide synthase mRNA in rFIP-glu-induced cells. rFIP-glu did not mediate these inflammatory effects through a general pathway but rather through a different pathway for a different inflammatory mediator. These data imply that rFIP-glu possessed immunomodulatory activity in macrophages, which was mediated through PI3K/Akt and MAPK pathways.


Asunto(s)
Proteínas Fúngicas/farmacología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Macrófagos/efectos de los fármacos , Fosfatidilinositol 3-Quinasas/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/efectos de los fármacos , Reishi , Animales , Inmunomodulación/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/fisiología , Activación de Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Células RAW 264.7
7.
Appl Microbiol Biotechnol ; 103(23-24): 9239-9250, 2019 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-31659419

RESUMEN

Ganoderma have been regarded as a traditional source of natural bioactive compounds for centuries and have recently been exploited for potential components in the cosmetics industry. Besides Ganoderma polysaccharides and triterpenes, multiple proteins have been found in Ganoderma. With the in-depth study of these proteins, various pharmacological functions of Ganoderma have become important in the discovery and development of new products. In the review, we summarized and discussed the kinds and characteristics of Ganoderma proteins, especially on fungal immunomodulatory proteins (FIPs) which can be potentially developed into cosmeceuticals or nutricosmetics and are a suitable target for production using established biotechnological methods. Furthermore, we discuss their pharmacological activities of the proteins with a focus on their pharmacological functions related to cosmetics, such as antioxidant activity, inhibition of melanin, antibacterial activity, and regulation of inflammatory mediators. Numerous other questions also are addressed before the proteins can be widely accepted and used as cosmetic additives.


Asunto(s)
Cosméticos/análisis , Proteínas Fúngicas/química , Ganoderma/química , Animales , Humanos , Ratones , Polisacáridos/metabolismo , Triterpenos/metabolismo
8.
BMC Biotechnol ; 17(1): 19, 2017 02 23.
Artículo en Inglés | MEDLINE | ID: mdl-28231778

RESUMEN

BACKGROUND: Manganese peroxidase (MnP) of white rot basidiomycetes, an extracellular heme enzyme, is part of a peroxidase superfamily that is capable of degrading the different phenolic compounds. Ganoderma, a white rot basidiomycete widely distributed worldwide, could secrete lignin-modifying enzymes (LME), including laccase (Lac), lignin peroxidases (LiP) and MnP. RESULTS: After the selection of a G. lucidum strain from five Ganoderma strains, the 1092 bp full-length cDNA of the MnP gene, designated as G. lucidum MnP (GluMnP1), was cloned from the selected strain. We subsequently constructed an eukaryotic expression vector, pAO815:: GlMnP, and transferred it into Pichia pastoris SMD116. Recombinant GluMnP1 (rGluMnP1) was with a yield of 126 mg/L and a molecular weight of approximately 37.72 kDa and a specific enzyme activity of 524.61 U/L. The rGluMnP1 could be capable of the decolorization of four types of dyes and the degradation of phenol. Phenol and its principal degradation products including hydroquinone, pyrocatechol, resorcinol, benzoquinone, were detected successfully in the experiments. CONCLUSIONS: The rGluMnP1 could be effectively expressed in Pichia pastoris and with a higher oxidation activity. We infer that, in the initial stages of the reaction, the catechol-mediated cycle should be the principal route of enzymatic degradation of phenol and its oxidation products. This study highlights the potential industrial applications associated with the production of MnP by genetic engineering methods, and the application of industrial wastewater treatment.


Asunto(s)
Colorantes/química , Peroxidasas/química , Fenol/química , Pichia/enzimología , Reishi/enzimología , Contaminantes Químicos del Agua/química , Biodegradación Ambiental , Clonación Molecular/métodos , Colorantes/aislamiento & purificación , Activación Enzimática , Peroxidasas/genética , Peroxidasas/metabolismo , Fenol/aislamiento & purificación , Pichia/genética , Ingeniería de Proteínas/métodos , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Reishi/clasificación , Reishi/genética , Especificidad de la Especie , Contaminantes Químicos del Agua/aislamiento & purificación , Purificación del Agua/métodos
9.
BMC Microbiol ; 16(1): 196, 2016 08 26.
Artículo en Inglés | MEDLINE | ID: mdl-27565900

RESUMEN

BACKGROUND: Ophiocordyceps sinensis (DongChong XiaCao (DCXC) in Chinese), a fungal parasite of caterpillars, is a traditional Chinese medicine. Bioactive components isolated from natural DCXC possess a wide range of pharmacological actions. Many efforts have been directed towards isolating the fungi based on culture-dependent methods for investigation of fungal diversity in order to determine the anamorph of natural DCXC and find new medicinal fungi resources, and the results have been varied. RESULTS: In the present study, a total of 44,588 bacterial and 51,584 fungal sequences corresponding to 11,694 and 9297 putative operational taxonomic units (OTU) were respectively identified by a Roche/454-based, high throughput sequence analysis of 16S rRNA genes and ITS regions. The main bacterial groups were Proteobacteria, Acidobacteria, Bacteroidetes, Actinobacteria and Firmicutes, while the Ascomycota, Basidiomycota and Zygomycota were the main fungal phyla. Proteobacteria presented 68.4, 49.5, 38.9 and 35.6 % of all bacteria in the sclerotia, stromata, external mycelial cortices and soil, respectively. As the main fungi phyla, Ascomycota presented 21.0, 45.6 26.4 and 59.3 % in the sclerotia, stromata, external mycelial cortices and soil, respectively. Bacterial and fungal communities were more diverse in the environmental sample than in the natural DCXC sample. Microbial communities were obviously distinct in each sample. Several novel unclassifiable bacterial (10.41 %) and fungal (37.92 %) species were also detected. CONCLUSIONS: This study revealed an abundant endogenetic fungal and bacterial resources and a variety of genetic information in natural DCXC by high-throughput 454 sequencing technology. Microorganism that had been discovered in natural DCXC will provide sources for screening the new bioactive metabolites and its biotechnological application.


Asunto(s)
Bacterias/genética , Hongos/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Microbiota , Microbiología del Suelo , Secuencia de Bases , Biodiversidad , ADN Bacteriano/genética , ADN de Hongos/genética , Ecosistema , Filogenia , ARN Ribosómico 16S/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Suelo , Tibet
10.
Can J Microbiol ; 61(2): 104-11, 2015 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-25578897

RESUMEN

Ophiocordyceps sinensis is a fungus that parasitizes caterpillars, and more than 30 species of filamentous fungi have been isolated from its fruiting body. However, its microbiological diversity remains unclear. Based on the clone library and quantitative PCR techniques, the bacterial flora and mycobiota of 3 different samples (larva, stromata/sclerotia, and surface soil) from natural O. sinensis specimens were investigated using primer sets that targeted the 16S rRNA gene and internal transcribed spacer region of ribosomal DNA. The results showed that the abundance of bacterial and fungal communities in the soil attached to the surface of O. sinensis was (6.4 ± 1.4) × 10(6) and (6.0 ± 0.3) × 10(7) copies/g dry matter, respectively, which was the highest compared with that in the larva and stromal samples. The main groups of bacteria in the O. sinensis samples were Proteobacteria and Actinobacteria, while Ascomycota was the most dominant fungal group in the 3 samples. At the genus level, Geomyces, Phoma, and Trichocladium were the dominant genera in the larval sample, while Geomyces and Cladosporium were the dominant genera in the stromal sample. In conclusion, a great number of bacterial and fungal species were present in naturally occurring O. sinensis specimens, and there was a high diversity of bacterial and fungal communities. These findings contribute to the understanding of the bacterial and fungal community structure of this valuable medicinal fungus and lay the foundation for the future discovery of new medicinal microorganism resources.


Asunto(s)
Actinobacteria/clasificación , Ascomicetos/clasificación , Hypocreales , Lepidópteros/microbiología , Proteobacteria/clasificación , Actinobacteria/aislamiento & purificación , Animales , Ascomicetos/aislamiento & purificación , ADN , Cartilla de ADN , ADN Bacteriano/genética , ADN de Hongos/genética , ADN Intergénico/genética , ADN Ribosómico/genética , Biblioteca de Genes , Proteobacteria/aislamiento & purificación , ARN Ribosómico 16S/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Suelo , Microbiología del Suelo , Tibet
11.
Crit Rev Biotechnol ; 34(3): 233-43, 2014 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-23875938

RESUMEN

Ophiocordyceps sinensis (syn. Cordyceps sinensis), a traditional Chinese medicine called DongChongXiaCao (DCXC) in Chinese, is well known and has been used in Asia countries since the fifteenth century, and it contains some valuable medicinal component defined by modern pharmacological science. DCXC only appears at high altitudes on the Qinghai-Tibetan Plateau. Consequently, it is difficult to find and harvest. Because of its rarity and medicinal value, DCXC has always been one of the most expensive medicines known. As the price of DCXC has risen in recent years, thousands of migrants have entered into the various grasslands to search for them in season, which makes ecological environments of the grassland more fragile. In order to relieve the environmental pressures and protect this valuable resource, the artificial cultivation of DCXC involving two aspects of the genus Hepialus and the fungi of the host larvae should be employed and applied at the first available time point. In this article, the reproduction of moth larvae of the genus Hepialus is first described, which includes their ecological characteristics and the methods of artificial feeding. Second, the generation and isolation method of the fungi from DCXC are subsequently summarized, and then the mechanism of fungal spores to attack the moth larvae are restated. Finally, the basic model of artificial cultivation of DCXC is introduced; meanwhile, the potential application of modern biotechnology to the artificial cultivation is analyzed in prospect. This review article will not only expand people's knowledge regarding the artificial cultivation of DCXC, but also hopefully provide an informative reference for the development of this valuable resource and the environmental protection of alpine meadows.


Asunto(s)
Cordyceps/fisiología , Microbiología Industrial/métodos , Mariposas Nocturnas/parasitología , Animales , China , Interacciones Huésped-Parásitos , Larva/parasitología , Medicina Tradicional China , Investigación
12.
Mol Biol Rep ; 41(7): 4381-7, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24584577

RESUMEN

Fugal immunomodulatory protein from Flammulina velutipes (FIP-fve) belongs to FIPs family, which has precious pharmaceutical value. To understand the regulatory mechanism of FIP-fve expression, we have cloned a 900 bp genomic DNA fragment from the transcriptional start site of the FIP-fve gene using genomic walker technology. Sequence analysis showed the presence of several eukaryotic transcription factor binding motifs in the 900 bp of upstream region of the FIP-fve gene, which contains one putative TATA-boxes, four possible CAAT-boxes, one ABRE, one ARE, three CGTCA-motifs, two TGA-elements and four Skn-1 motifs. The eukaryotic expression vector pfveP:: GUS-GFP was transferred into tobacco via an agrobacterium-mediated leaf disc transformation. The results showed that the FIP-fve promoter could induce the reporter gene GUS or GFP expression in different tissues of tobaccos. This study would lay a foundation for expression regulation of FIP-fve and development of genetic-modified plant products.


Asunto(s)
Flammulina/genética , Cuerpos Fructíferos de los Hongos/genética , Proteínas Fúngicas/genética , Factores Inmunológicos/genética , Nicotiana/genética , Regiones Promotoras Genéticas , Agrobacterium tumefaciens/genética , Sitios de Unión , Clonación Molecular , Flammulina/química , Flammulina/metabolismo , Cuerpos Fructíferos de los Hongos/química , Cuerpos Fructíferos de los Hongos/metabolismo , Proteínas Fúngicas/metabolismo , Técnicas de Transferencia de Gen , Genes Reporteros , Vectores Genéticos , Factores Inmunológicos/metabolismo , Datos de Secuencia Molecular , Motivos de Nucleótidos , Unión Proteica , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Nicotiana/metabolismo
13.
Appl Microbiol Biotechnol ; 98(13): 5967-75, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24682474

RESUMEN

Fungal immunomodulatory protein (FIP), extracted from higher basidiomycetes, is a kind of small molecule protein with extensive biological functions, including anti-tumor and anti-allergy, stimulating immune cells to produce a variety of cytokines, etc. Compared with FIP-glu, FIP-SN15, a novel gene shuffled from the genes of Ganoderma sinensis and Ganoderma lucidum FIP, was used as the object in this study. Based on the construction of prokaryotic expression vectors, both pET30a-FIP-glu and pET30a-FIP-SN15 were expressed in Escherichia coli. Then the recombinant proteins are respectively analyzed by Western blot, Q-TOF MS, and bioinformatics techniques. Finally, effects of reFIPs on cell cycle and apoptosis of human glioblastoma cell line U-251 MG were studied by fluorescence activated cell sorting (FACS). The results showed that the recombinant proteins FIP-SN15 and FIP-glu could be successfully expressed in E. coli, the yield of which was 35.95 and 36.67 mg/L, respectively. The recombinant protein FIP-SN15 consisted of 111 amino acids, and four peptides were identified by Q-TOF MS with a coverage of 91.9 %. The secondary and tertiary structure of FIP-SN15 were also predicted by bioinformatics method which suggest that reFIP-SN15 was a new member of FIPs family. FACS analysis showed that 10 µg/mL FIP-SN15 and FIP-glu could induce U-251 MG cells apoptosis, the apoptotic rates were increased by 6.03 and 22.01 %, respectively. The results of reFIPs on U-251 MG cell cycle indicated that reFIPs could inhibit cell cycle progression by retardation of G1/S transition. The efforts in this assay would lay the foundation for further development of reFIPs products and research on the anti-tumor mechanisms of FIP-SN15.


Asunto(s)
Antineoplásicos/aislamiento & purificación , Antineoplásicos/farmacología , Apoptosis , Ciclo Celular/efectos de los fármacos , Línea Celular , Clonación Molecular , Barajamiento de ADN , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/aislamiento & purificación , Proteínas Fúngicas/farmacología , Ganoderma/genética , Expresión Génica , Humanos , Neuroglía/efectos de los fármacos , Neuroglía/fisiología , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/farmacología
14.
Crit Rev Microbiol ; 39(4): 416-26, 2013 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-22992227

RESUMEN

White-rot fungal species belonging to Ganoderma have long been used as medicinal mushrooms in many Asian countries. In recent years, however, attention is not just being paid to their pharmacological properties, but to their other potentially valuable features as well, including their secretion of enzymes which decompose lignin. The current literature regarding lignin-modifying enzymes from the genus Ganoderma, their potential uses, and the components, structures and processes of lignocellulose degradation are discussed. The ligninolytic enzymes from the genus Ganoderma, as well as the number of additional enzymes that participate in lignin degradation, are summarized; further, the potential applications of these enzymes are analyzed and probed in this article. This review will provide insight on the valuable applications of Ganoderma spp. and will serve as a useful reference on the use of lignocellulose degradation as a means of environmental protection.


Asunto(s)
Ganoderma/enzimología , Hidroliasas/metabolismo , Lignina/metabolismo , Biotecnología/métodos
15.
Appl Microbiol Biotechnol ; 97(6): 2455-65, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22615051

RESUMEN

Fungal immunomodulatory proteins (FIPs) found in a wide variety of mushrooms hold significant therapeutic potential. Despite much research, the structural determinants for their immunomodulatory functions remain unknown. In this study, a DNA shuffling technique was used to create two shuffled FIP protein libraries: an intrageneric group containing products of shuffling between FIP-glu (FIP gene isolated from Ganoderma lucidum) and FIP-gsi (FIP gene isolated from Ganoderma sinense) genes and an intergeneric group containing the products of shuffling between FIP-glu, FIP-fve (FIP gene isolated from Flammulina velutipes), and FIP-vvo (FIP gene isolated from Volvariella volvacea) genes. The gene shuffling generated 426 and 412 recombinant clones, respectively. Using colony blot analysis, we selected clones that expressed relatively high levels of shuffled gene products recognized by specific polyclonal antibodies. We analyzed the DNA sequences of the selected shuffled genes, and testing of their protein products revealed that they maintained functional abilities to agglutinate blood cells and induce cytokine production by splenocytes from Kunming mice in vitro. Meanwhile, the relationships between protein structure and the hemagglutination activity and between the changed nucleotide sites and expression levels were explored by bioinformatic analysis. These combined analyses identified the nucleotide changes involved in regulating the expression levels and hemagglutination activities of the FIPs. Therefore, we were able to generate recombinant FIPs with improved biological activities and expression levels by using DNA shuffling, a powerful tool for the generation of novel therapeutic proteins and for their structural and functional studies.


Asunto(s)
Barajamiento de ADN/métodos , Evolución Molecular Dirigida/métodos , Proteínas Fúngicas/genética , Proteínas Fúngicas/inmunología , Factores Inmunológicos/genética , Factores Inmunológicos/inmunología , Secuencia de Aminoácidos , Animales , Citocinas/metabolismo , Flammulina/genética , Flammulina/inmunología , Ganoderma/genética , Ganoderma/inmunología , Hemaglutinación , Leucocitos Mononucleares/inmunología , Ratones , Proteínas Recombinantes/genética , Proteínas Recombinantes/inmunología , Análisis de Secuencia de ADN , Volvariella/genética , Volvariella/inmunología
16.
Environ Pollut ; 317: 120763, 2023 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-36503821

RESUMEN

High temperature and drought are abiotic stresses restricting many arthropods' survival and growth. Wolf spiders are poikilothermic arthropods that are vital in managing insects and pests. Nonetheless, investigating changes in spiders under temperature and drought stress are limited, especially at the molecular and gene expression levels. The study found that the combined effects of high temperature and drought stress significantly reduced survival rates and raised superoxide dismutase and malondialdehyde levels in the wolf spider Pardosa pseudoannulata. An integrated transcriptome and metabolome analysis showed that differentially expressed genes and metabolites were highly enriched in pathways involved in the proteolysis and oxidation-reduction process. The gene expression profiles displayed that heat shock protein (HSP) families (i.e., small heat shock protein, HSP70, HSP90, and HSP beta protein) were up-regulated under temperature and/or drought stresses. Additionally, a conjoint analysis revealed that under the combined stress, several important enzymes, including maltase-glucoamylase, glycerol-6-phosphate transporter, alanine-glyoxylate transaminase, and prostaglandin-H2 D-isomerase, were altered, affecting the metabolism of starch, sucrose, amino acids, and arachidonic acid. The protein interaction network further confirmed that under the combined stress, metabolic processes, peptide metabolic processes, and ATP generation from ADP were up-regulated, indicating that spiders could accelerate the metabolism of carbohydrates and proteins to combat stress and maintain homeostasis. Overall, this work showed that exposure to a combination of pressures might cause distinct defensive reactions in spiders and offered novel perspectives to research the molecular underpinnings of spider adaptation to a changing climate.


Asunto(s)
Arañas , Transcriptoma , Animales , Temperatura , Arañas/genética , Sequías , Metaboloma , Estrés Fisiológico
17.
Food Sci Biotechnol ; 32(3): 265-282, 2023 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-36619215

RESUMEN

Proteins do not only serve as nutrients to fulfill the demand for food, but also are used as a source of bioactive proteins/polypeptides for regulating physical functions and promoting physical health. Female breast cancer has the highest incidence in the world and is a serious threat to women's health. Bioactive proteins/polypeptides exert strong anti-tumor effects and exhibit inhibition of multiple breast cancer cells. This review discussed the suppressing effects of bioactive proteins/polypeptides on breast cancer in vitro and in vivo, and their mechanisms of migration and invasion inhibition, apoptosis induction, and cell cycle arrest. This may contribute to providing a basis for the development of bioactive proteins/polypeptides for the treatment of breast cancer.

18.
Mol Biol Rep ; 39(12): 10303-11, 2012 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-23053936

RESUMEN

A Cu/Zn-superoxide dismutase (SOD) gene was characterized from Cordycepes militaris by gene cloning, heterogeneous expression and function analysis. This 154-aa SOD (CmSOD) was deduced from a 465-bp gene cloned, showing 72-95 % sequence identity to Cu/Zn-SODs from other fungi. The deduced amino acid sequence of the cDNA is highly similar to Beauveria bassiana (95 %), Isaria tenuipes (94 %) and Claviceps purpurea (88 %), respectively. The SOD gene of C. militaris spin 589 bp and consisted of two introns and three exons. The CmSOD coding region sequence was inserted into plasmid pQE-30 in order to construct prokaryotic expression vector, then transformed into Escherichia coli M15 cells for expression, and a mass of rCmSOD was obtained by IPTG induction. The enzyme activity of the purified rCmSOD was approximately 714.48 U/mg after the assay. The study provided a way for in-depth research on the expression and regulation of the CmSOD, and the molecular mechanism of anti-oxidative effect in C. militaris.


Asunto(s)
Cordyceps/enzimología , Proteínas Fúngicas/metabolismo , Superóxido Dismutasa/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , Escherichia coli , Exones , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/aislamiento & purificación , Expresión Génica , Insectos/microbiología , Intrones , Modelos Moleculares , Datos de Secuencia Molecular , Filogenia , Estructura Terciaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Homología Estructural de Proteína , Superóxido Dismutasa/química , Superóxido Dismutasa/genética , Superóxido Dismutasa/aislamiento & purificación
19.
Appl Microbiol Biotechnol ; 93(3): 941-63, 2012 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-22170106

RESUMEN

A white-rot basidiomycete Ganoderma spp. has long been used as a medicinal mushroom in Asia, and it has an array of pharmacological properties for immunomodulatory activity. There have been many reports about the bioactive components and their pharmacological properties. In order to analyze the current status of Ganoderma products, the detailed process of cultivation of Ganoderma spp. and development of their products are restated in this review article. These include the breeding, cultivating, extracting bioactive component, and processing Ganoderma products, etc. This article will expand people's common knowledge on Ganoderma, and provide a beneficial reference for research and industrial production.


Asunto(s)
Factores Biológicos/metabolismo , Biotecnología/métodos , Ganoderma/crecimiento & desarrollo , Agaricales/química , Agaricales/crecimiento & desarrollo , Agaricales/metabolismo , Asia , Factores Biológicos/aislamiento & purificación , Factores Biológicos/uso terapéutico , Ganoderma/química , Ganoderma/metabolismo , Humanos , Medicina Tradicional China , Control de Calidad
20.
Artículo en Inglés | MEDLINE | ID: mdl-35981662

RESUMEN

Cadmium (Cd) is a kind of toxic heavy metal widely distributed in the environment, posing life-threatening challenges to organisms. The paddy field spider is a natural enemy of pests and an essential component of rice biodiversity. Nonetheless, the effects of Cd stress on the postembryonic development of spiders and its detailed mechanism remain to be investigated. In the present study, we found that Cd stress posed adverse impacts on the growth indicators (e.g., carapace length, development duration, and survival rate) and increased the levels of three antioxidants (i.e., superoxide dismutase, glutathione S-transferase, and glutathione peroxidase) in the spiderlings of Pirata subpiraticus. An in-depth transcriptome analysis was employed in the study, and the results displayed that differentially expressed genes (DEGs) involved in postembryonic morphogenesis, development involved in symbiotic interaction, postembryonic development, and growth were distinctively altered under Cd stress. Further enrichment analysis showed that Cd exposure could activate the apoptosis pathway in the spider via the up-regulation of several key factors, including caspase-10, α-tubulin, actin, etc. In addition, we demonstrated that the increased level of glutathione-related enzymes in spiderlings was caused by the activation of glutathione metabolic pathway. The altered hedgehog signaling pathway might affect cell proliferation, tissue patterning, and development of spiderlings. Further protein interaction network displayed that Cd stress could affect multiple biological processes in spiderlings, particularly cellular response to stimulus and system development. To sum up, this study can provide multi-level perspectives to understand the toxicity of Cd on the growth and development of spiders.


Asunto(s)
Cadmio , Arañas , Animales , Cadmio/toxicidad , Perfilación de la Expresión Génica , Glutatión , Proteínas Hedgehog/genética , Proteínas Hedgehog/farmacología , Arañas/genética , Transcriptoma
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