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1.
J Agric Food Chem ; 67(37): 10423-10431, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31487168

RESUMO

Plants often produce antifungal peptides and proteins in response to infection. Also wheat, which is the main ingredient of bread dough, contains such components. Here, we show that while some industrial strains of the baker's yeast Saccharomyces cerevisiae can efficiently ferment dough, some other strains show much lower fermentation capacities because they are sensitive to a specific wheat protein. We purified and identified what turned out to be a thaumatin-like protein through a combination of activity-guided fractionation, cation exchange chromatography, reversed-phase HPLC, and LC-MS/MS. Recombinant expression of the corresponding gene and testing the activity confirmed the inhibitory activity of the protein.


Assuntos
Proteínas de Plantas/química , Proteínas de Plantas/farmacologia , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/crescimento & desenvolvimento , Triticum/química , Cromatografia Líquida , Fermentação , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/metabolismo , Saccharomyces cerevisiae/metabolismo , Espectrometria de Massas em Tandem , Triticum/genética , Triticum/metabolismo , Triticum/microbiologia
2.
J Agric Food Chem ; 67(37): 10296-10305, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31464437

RESUMO

Grass pea is an orphan legume that is grown in many places in the world. It is a high-protein, drought-tolerant legume that is capable of surviving extreme environmental challenges and can be a sole food source during famine. However, grass pea produces the neurotoxin ß-N-oxalyl-L-α,ß-diaminopropionic acid (ß-ODAP), which can cause a neurological disease. This crop is promising as a food source for both animals and humans if ß-ODAP levels and other antinutritional factors such as protease inhibitors are lowered or removed. To understand more about these proteins, a proteomic analysis of grass pea was conducted using three different extraction methods to determine which was more efficient at isolating antinutritional factors. Seed proteins extracted with Tris-buffered saline (TBS), 30% ethanol, and 50% isopropanol were identified by mass spectrometry, resulting in the documentation of the most abundant proteins for each extraction method. Mass spectrometry spectral data and BLAST2GO analysis led to the identification of 1376 proteins from all extraction methods. The molecular function of the extracted proteins revealed distinctly different protein functional profiles. The majority of the TBS-extracted proteins were annotated with nutrient reservoir activity, while the isopropanol extraction yielded the highest percentage of endopeptidase proteinase inhibitors. Our results demonstrate that the 50% isopropanol extraction method was the most efficient at isolating antinutritional factors including protease inhibitors.


Assuntos
Fracionamento Químico/métodos , Fabaceae/química , Extratos Vegetais/isolamento & purificação , Inibidores de Proteases/isolamento & purificação , Sementes/química , Endopeptidases/química , Fabaceae/genética , Fabaceae/metabolismo , Espectrometria de Massas , Extratos Vegetais/química , Extratos Vegetais/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/metabolismo , Inibidores de Proteases/química , Inibidores de Proteases/metabolismo , Proteômica , Sementes/genética , Sementes/metabolismo
3.
J Food Sci ; 84(8): 2357-2363, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31364176

RESUMO

Peanut allergic consumers rely on food package labels to avoid foods containing peanut. The inadvertent presence of peanut in foods due to cross-contact can be fatal if ingestion of such food leads to an allergic reaction. Analytical methods are available to detect undeclared peanut in foods. However, depending on the type of food matrix and food processing parameters, method performance can be adversely affected due to reduction in the extraction efficiency of peanut proteins. Temperature and probe sonication were used as a preincubation treatment for peanut flour slurries to assess their effect on the total peanut protein solubility from raw, light-roasted, and dark-roasted peanut flours. The effect of these treatments on the immunoreactivity of peanut allergens (Ara h 1, Ara h 2, Ara h 3, and Ara h 6) was determined by an indirect enzyme-linked immunosorbent assay using antibodies raised against these individual peanut proteins. Preincubation at 50 °C did not significantly improve the peanut protein solubility, whereas an increase in protein solubility was observed when light- and dark-roasted peanut flour slurries were preincubated at 90 °C or sonicated. The immunoreactivity of peanut allergens varied depending on the degree of peanut flour roasting and type of preincubation treatment. Overall, the immunoreactivity of peanut allergens from most peanut flour slurries was unaffected when preincubated at 50 °C for up to 60 min or sonicated with a probe for up to 5 min, whereas preincubation at 90 °C resulted in a time-dependent reduction in immunoreactivity of peanut allergens. Sonication treatment may improve peanut protein extraction without markedly affecting their immunoreactivity. PRACTICAL APPLICATION: Extraction of peanut proteins is vital for developed analytical methods to estimate peanut allergens in foods. The manuscript describes the effect of two different temperatures (50 and 90 °C) and probe-type sonication on peanut protein solubility. The findings suggest sonication can improve peanut protein solubility without markedly affecting their immunoreactivity.


Assuntos
Arachis/imunologia , Antígenos de Plantas/análise , Antígenos de Plantas/imunologia , Antígenos de Plantas/isolamento & purificação , Arachis/química , Ensaio de Imunoadsorção Enzimática , Farinha/análise , Manipulação de Alimentos , Humanos , Proteínas de Plantas/análise , Proteínas de Plantas/imunologia , Proteínas de Plantas/isolamento & purificação , Temperatura Ambiente
4.
Rev Bras Parasitol Vet ; 28(3): 339-345, 2019 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-31291435

RESUMO

Gastrointestinal nematode infection is an important cause of high economic losses in livestock production. Nematode control based on a synthetic chemical approach is considered unsustainable due to the increasing incidence of anthelmintic resistance. Control alternatives such as the use of natural products are therefore becoming relevant from an environmental and economic point of view. Proteins are macromolecules with various properties that can be obtained from a wide range of organisms, including plants and fungi. Proteins belonging to different classes have shown great potential for the control of nematodes. The action of proteins can occur at specific stages of the nematode life cycle, depending on the composition of the external layers of the nematode body and the active site of the protein. Advances in biotechnology have resulted in the emergence of numerous protein and peptide therapeutics; however, few have been discussed with a focus on the control of animal nematodes. Here, we discuss the use of exogenous proteins and peptides in the control of gastrointestinal.


Assuntos
Antinematódeos/isolamento & purificação , Proteínas Fúngicas/isolamento & purificação , Gastroenteropatias/veterinária , Infecções por Nematoides/veterinária , Peptídeos/isolamento & purificação , Proteínas de Plantas/isolamento & purificação , Animais , Antinematódeos/administração & dosagem , Biotecnologia , Quitinases/administração & dosagem , Quitinases/isolamento & purificação , Proteínas Fúngicas/administração & dosagem , Gastroenteropatias/parasitologia , Infecções por Nematoides/tratamento farmacológico , Peptídeo Hidrolases/administração & dosagem , Peptídeo Hidrolases/isolamento & purificação , Peptídeos/administração & dosagem , Proteínas de Plantas/administração & dosagem
5.
J Agric Food Chem ; 67(29): 8119-8129, 2019 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-31265283

RESUMO

Grass pea (Lathyrus sativus L.) is an important legume commonly grown in arid and semi-arid regions. This protein-rich legume performs well even under harsh environmental conditions and is considered to be a strategic famine food in developing countries. Unfortunately, its potential usage is greatly limited as a result of the presence of antinutritional factors, including the neuroexcitatory amino acid ß-N-oxalyl-l-α,ß-diaminopropionic acid (ß-ODAP) and protease inhibitors. ß-ODAP is responsible for a neurodegenerative syndrome that results in the paralysis of lower limbs, while protease inhibitors affect protein digestibility, resulting in reduced growth. Concerted research efforts have led to development of grass pea cultivars with reduced ß-ODAP content. In contrast, very little information is available on the protease inhibitors of L. sativus. In this study, we have conducted biochemical characterization of 51 L. sativus accessions originating from different geographical regions. Sodium dodecyl sulfate polyacrylamide gel electrophoresis analyses of seed globulins and prolamins revealed striking similarity in their protein profile, although geographic-specific variations in profiles was also evident. Measurement of Bowman-Birk chymotrypsin inhibitor (BBi) and Kunitz trypsin inhibitor (KTi) activities in accessions revealed striking differences among them. Amino acid sequence alignment of grass pea BBi and KTi revealed significant homology to protease inhibitors from several legumes. Real-time polymerase chain reaction analysis demonstrated high-level expression of BBi and KTi in dry seeds and weak expression in other organs. Our study demonstrates substantial variation in BBi and KTi among grass pea accessions that could be exploited in breeding programs for the development of grass pea lines that are devoid of these antinutritional factors.


Assuntos
Lathyrus/química , Proteínas de Plantas/química , Inibidor da Tripsina de Soja de Bowman-Birk/química , Sequência de Aminoácidos , Geografia , Lathyrus/genética , Lathyrus/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/metabolismo , Alinhamento de Sequência , Inibidor da Tripsina de Soja de Bowman-Birk/genética , Inibidor da Tripsina de Soja de Bowman-Birk/isolamento & purificação , Inibidor da Tripsina de Soja de Bowman-Birk/metabolismo
6.
Food Chem ; 300: 125162, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31325745

RESUMO

Moringa oleifera is a worldwide cultivated edible and medicinal plant. Its seeds are rich in oil, proteins, and glucosinolates. A practical method was developed to simultaneously extract and separate the three groups of substances from M. oleifera seeds. Smashed seed material was loaded into columns with petroleum ether: ethanol 8:2 (PE-ethanol) and eluted sequentially with 4.8-fold PE-ethanol to extract oil, and 10.8-fold water to extract proteins and glucosinolates. More than 95% of oil, proteins, and glucosinolates were extracted. The extracts were separated automatically into ether (oil) phase and ethanol aqueous phase. The latter was further separated into proteins and glucosinolates by 70% ethanol precipitation. The main glucosinolate was identified by LC-MS as GLC (4-α-rhamnopyranosyloxy-benzyl glucosinolate). After purification, 22.3 g refined oil, 33.0 g proteins, and 5.5 g purified GLC from 100 g M. oleifera seeds were obtained. This study provides a simple and high-efficient method to utilize M. oleifera seeds.


Assuntos
Cromatografia Líquida/métodos , Glucosinolatos/isolamento & purificação , Espectrometria de Massas/métodos , Moringa oleifera/química , Óleos Vegetais/isolamento & purificação , Proteínas de Plantas/isolamento & purificação , Extratos Vegetais/química , Sementes/química
7.
Carbohydr Polym ; 220: 247-255, 2019 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-31196547

RESUMO

A water-soluble polysaccharide (type II arabinogalactan-protein) extracted from the gum exudate of the native New Zealand puka tree (Meryta sinclairii), was characterised for its molecular, rheological and physicochemical properties. In 0.1 M NaCl, the weight average molecular weight (Mw) of puka gum is 5.9 × 106 Da with an RMS radius of 56 nm and z-average hydrodynamic radius of 79 nm. The intrinsic viscosity of the polysaccharide is 57 ml/g with a coil overlap concentration 15% w/w. Together, the shape factor, p, of 0.70 (exponent of RMS radius vs. hydrodynamic radius), Smidsrød-Haug's stiffness parameter B of 0.031 and Mark-Houwink exponent α of 0.375 indicate that the polysaccharide adopts a spherical conformation in solution, similar to gum arabic. The pKa is 1.8. The polysaccharide exhibits a Newtonian to shear-thinning behaviour from 0.2 to 25% w/w. Viscosity of the polysaccharide (1 s-1) decreases with decreasing concentration, increasing temperature, ionic strength, and at acidic pH.


Assuntos
Araliaceae/metabolismo , Mucoproteínas/química , Gomas Vegetais/química , Polissacarídeos/química , Árvores/metabolismo , Peso Molecular , Mucoproteínas/isolamento & purificação , Nova Zelândia , Gomas Vegetais/isolamento & purificação , Proteínas de Plantas/química , Proteínas de Plantas/isolamento & purificação , Polissacarídeos/isolamento & purificação , Reologia , Solubilidade , Viscosidade
8.
Life Sci ; 231: 116535, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31175857

RESUMO

Latex proteins from P. pudica (LPPp) have anti-inflammatory activity. In the present study, LPPp was evaluated to protect animals against inflammatory ulcerative colitis (UC). UC was induced by intracolonic instillation of a 6% acetic acid solution and the animals received LPPp (10, 20 or 40 mg/kg) by intraperitoneal route 1 h before and 17 h after acetic acid injection. Eighteen hours after instillation of acetic acid, the mice were euthanized and the colons were excised to determine the wet weight, macroscopic and microscopic lesion scores, myeloperoxidase (MPO) activity, IL1-ß levels, glutathione (GSH) and malondialdehyde (MDA) concentration and superoxide dismutase (SOD) activity. The results revealed that LPPp treatment (40 mg/kg) had a protective effect on acetic acid-induced colitis by reducing the wet weight, macroscopic and microscopic scores of intestinal lesions and colonic MPO activity. Additionally, LPPp inhibited tissue oxidative stress, since decreases in GSH consumption, MDA concentration and SOD activity were observed. The treatment with LPPp reduced the levels of cytokine IL-1ß, contributing to the reduction of colon inflammation. Biochemical investigation showed that LPPp comprises a mixture of proteins containing proteinases, chitinases and proteinase inhibitors. These data suggest that LPPp has a protective effect against intestinal damage through mechanisms that involve the inhibition of inflammatory cell infiltration, cytokine release and oxidative stress.


Assuntos
Apocynaceae/química , Colite/tratamento farmacológico , Látex/farmacologia , Proteínas de Plantas/farmacologia , Ácido Acético , Animais , Apocynaceae/metabolismo , Colite/induzido quimicamente , Colite/metabolismo , Colo/efeitos dos fármacos , Citocinas/metabolismo , Glutationa/metabolismo , Inflamação/tratamento farmacológico , Interleucina-1beta/metabolismo , Intestinos/patologia , Látex/isolamento & purificação , Masculino , Camundongos , Estresse Oxidativo/efeitos dos fármacos , Proteínas de Plantas/isolamento & purificação , Substâncias Protetoras/farmacologia , Fator de Necrose Tumoral alfa/metabolismo
9.
Food Chem ; 294: 557-564, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31126499

RESUMO

Superoxide dismutase (SOD) is an important enzyme with antioxidant function. The activity of purified SOD from chestnut rose was increased by 22.23-38.02% after high pressure processing (HPP, 100-500 MPa/0-20 min/30-50 °C). The properties of SOD induced by high pressure were studied by means of electrophoresis, dynamic light scattering, enthalpy and surface hydrophobicity, circular dichroism and fluorescence spectrometry. Results showed that high pressure did not change the electrophoretic properties, particle size distribution, digestive system stability and antioxidant capacity of SOD. However, the enthalpy and surface hydrophobicity of SOD was increased. The increase of SOD activity was related to the increase of SOD surface hydrophobicity. Moreover, the α-helix fraction of SOD was decreased by 8.7% and the intrinsic fluorescence intensity of SOD was increased by 5.7% when exposed to high pressure. This study suggested that HPP can be used as a novel way of increasing the activity of SOD in chestnut rose.


Assuntos
Proteínas de Plantas/metabolismo , Rosa/metabolismo , Superóxido Dismutase/metabolismo , Antioxidantes/metabolismo , Dicroísmo Circular , Difusão Dinâmica da Luz , Interações Hidrofóbicas e Hidrofílicas , Proteínas de Plantas/química , Proteínas de Plantas/isolamento & purificação , Pressão , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Superóxido Dismutase/química , Superóxido Dismutase/isolamento & purificação , Propriedades de Superfície
10.
Molecules ; 24(9)2019 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-31058866

RESUMO

Background: Wet methods of 1-3, 1-4 -ß-D-glucan isolation from cereals differ mainly in the type of grain fraction used as raw material, the solid-liquid ratio of ß-glucan in raw material vs. solvent used, and the type of aqueous solvent modification (alkali, neutral or acidic). All these factors impact the characterization of the residues finally found in extracts. Oat bran is a rich source of globulin fraction which can be transferred into the extracts, especially when a high pH is employed. Methods: A multi-stage (enzymatic and acidic) purification procedure was performed to remove the residues, especially starch and protein, from ß-glucan isolates from oat of different molar mass. Pancreatin, thermostable α-amylase, amyloglucosidase, and papain were used for consecutive residue removal. Three levels of low pH = 4.5, 3.5 and 3.0 were also tested for effective protein precipitation. Results: The starch hydrolysis and liquefaction significantly facilitate the proteinaceous matter removal although papain usage showed an intensive unfavorable impact on ß-glucan molar mass. Soluble protein content was significantly decreased after pancreatin and α-amylase treatment, while the significant reduction of amine nitrogen was noted after complete starch hydrolysis and a second acidification step. Conclusions: A complex procedure employing different enzymes is needed to successfully reduce the possibly bioactive residues in isolated oat ß-glucan fractions.


Assuntos
Enzimas/metabolismo , Globulinas/isolamento & purificação , Amido/isolamento & purificação , beta-Glucanas/química , Avena , Precipitação Fracionada , Globulinas/química , Concentração de Íons de Hidrogênio , Hidrólise , Proteínas de Plantas/química , Proteínas de Plantas/isolamento & purificação , Solventes/química , Amido/química , Viscosidade , beta-Glucanas/isolamento & purificação
11.
Molecules ; 24(9)2019 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-31052194

RESUMO

This work demonstrated a method combining reversed-phase high-performance liquid chromatography (RP-HPLC) with chemometrics analysis to identify the authenticity of Ranae Oviductus. The fingerprint chromatograms of the Ranae Oviductus protein were established through an Agilent Zorbax 300SB-C8 column and diode array detection at 215 nm, using 0.085% TFA (v/v) in acetonitrile (A) and 0.1% TFA in ultrapure water (B) as mobile phase. The similarity was in the range of 0.779-0.980. The fingerprint chromatogram of Ranae Oviductus showed a significant difference with counterfeit products. Hierarchical clustering analysis (HCA) and principal component analysis (PCA) successfully identified Ranae Oviductus from the samples. These results indicated that the method established in this work was reliable.


Assuntos
Cromatografia Líquida de Alta Pressão , Cromatografia de Fase Reversa , Medicamentos de Ervas Chinesas/análise , Medicamentos de Ervas Chinesas/química , Materia Medica/química , Mapeamento de Peptídeos , Proteínas de Plantas/química , Análise por Conglomerados , Materia Medica/classificação , Mapeamento de Peptídeos/métodos , Proteínas de Plantas/análise , Proteínas de Plantas/isolamento & purificação , Reprodutibilidade dos Testes
12.
J Agric Food Chem ; 67(23): 6625-6632, 2019 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-31117491

RESUMO

Fava bean protein isolate (FBPI) was hydrolyzed by Alcalase with different degrees of hydrolysis (DHs), and the role of hydrolysates in oil-in-water (O/W) emulsion stability was investigated. Four emulsions, DH0, DH4, DH9, and DH15, were prepared by 1% (w/v) FBPI hydrolysates with different DHs (0% as the control and 4, 9, and 15%) and 5% (w/v) purified rapeseed oil. The emulsions were monitored for physical and oxidative stability at 37 °C for 7 days. DH4 and DH0 exhibited better physical stability than DH9 and DH15, indicated by droplet size, morphology, and Turbiscan stability index. More importantly, FBPI hydrolysates with DH of 4% most effectively inhibited lipid oxidation (i.e., formation of conjugated dienes and hexanal) while maintaining protein oxidative stability compared to the native and extensively hydrolyzed FBPI. Higher DHs (9 and 15%) induced unduly decreased surface hydrophobicity and increased surface load, which might negatively affect the emulsifying activity. FBPI hydrolysates with DH of 4% had suitable molecular weight for better interfacial layer stability, increased surface net charge for more repulsive electrostatic force, and increased hydrophobicity for better adsorption at the interface and, therefore, may serve as potential natural emulsifiers to maintain both physical and oxidative stability of O/W emulsions.


Assuntos
Proteínas de Plantas/química , Óleo de Brassica napus/química , Sementes/química , Vicia faba/química , Biocatálise , Emulsões/química , Hidrólise , Interações Hidrofóbicas e Hidrofílicas , Oxirredução , Proteínas de Plantas/isolamento & purificação , Estabilidade Proteica , Subtilisinas/química
13.
Molecules ; 24(10)2019 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-31109069

RESUMO

Stable-isotope dimethyl labeling is a highly reactive and cost-effective derivatization procedure that could be utilized in proteomics analysis. In this study, a liquid chromatography- tandem mass spectrometry in multiple reaction monitoring mode (LC-MS-MRM) platform for the quantification of kiwi allergens was first developed using this strategy. Three signature peptides for target allergens Act d 1, Act d 5, and Act d 11 were determined and were derivatized with normal and deuterated formaldehyde as external calibrants and internal standards, respectively. The results showed that sample preparation with the phenol method provided comprehensive protein populations. Recoveries at four different levels ranging from 72.5-109.3% were achieved for the H-labeled signature peptides of Act d 1 (SPA1-H) and Act d 5 (SPA5-H) with precision ranging from 1.86-9.92%. The limit of quantification (LOQ) was set at 8 pg mL-1 for SPA1-H and at 8 ng mL-1 for SPA5-H. The developed procedure was utilized to analyze seven kinds of hand-made kiwi foods containing 0.0175-0.0515 mg g-1 of Act d 1 and 0.0252-0.0556 mg g-1 of Act d 5. This study extended the applicability of stable-isotope dimethyl labeling to the economical and precise determination of food allergens and peptides.


Assuntos
Alérgenos/análise , Cromatografia Líquida , Análise de Alimentos , Alimentos/efeitos adversos , Marcação por Isótopo , Espectrometria de Massas em Tandem , Actinidia/efeitos adversos , Alérgenos/imunologia , Sequência de Aminoácidos , Análise de Alimentos/métodos , Frutas/efeitos adversos , Limite de Detecção , Proteínas de Plantas/análise , Proteínas de Plantas/imunologia , Proteínas de Plantas/isolamento & purificação , Padrões de Referência , Reprodutibilidade dos Testes
14.
Molecules ; 24(10)2019 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-31109085

RESUMO

Apart from non-enzymatic browning, polyphenol oxidase (PPO) also plays a role in the browning reaction of orange (Citrus sinensis Osbeck) juice, and needs to be inactivated during the processing. In this study, the protein with high PPO activity was purified from orange (Citrus sinensis Osbeck) and inactivated by ultrasonic processing. Fluorescence spectroscopy, circular dichroism (CD) and Dynamic light scattering (DLS) were used to investigate the ultrasonic effect on PPO activity and structural changes on purified PPO. DLS analysis illustrated that ultrasonic processing leads to initial dissociation and final aggregation of the protein. Fluorescence spectroscopy analysis showed the decrease in fluorescence intensity leading to the exposure of Trp residues to the polar environment, thereby causing the disruption of the tertiary structure after ultrasonic processing. Loss of α-helix conformation leading to the reorganization of secondary structure was triggered after the ultrasonic processing, according to CD analysis. Ultrasonic processing could induce aggregation and modification in the tertiary and secondary structure of a protein containing high PPO activity in orange (Citrus sinensis Osbeck), thereby causing inactivation of the enzyme.


Assuntos
Catecol Oxidase/química , Citrus sinensis/química , Proteínas de Plantas/química , Conformação Proteica/efeitos da radiação , Ondas Ultrassônicas , Catecol Oxidase/isolamento & purificação , Catecol Oxidase/metabolismo , Dicroísmo Circular , Citrus sinensis/enzimologia , Ativação Enzimática , Reação de Maillard , Peso Molecular , Tamanho da Partícula , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/metabolismo , Análise Espectral
15.
Food Funct ; 10(5): 2894-2905, 2019 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-31070610

RESUMO

Maca protein isolate (MPI) was extracted from maca root, and its physicochemical and functional properties, and the secondary structure and immunomodulatory activity of its major protein component, MMP, were investigated. The MPI lacked essential amino acids compared with soybean protein isolate (SPI) and casein, but was rich in cysteine and proline. The MPI had rich free sulfhydryl (20.6 µmol g-1), and its surface hydrophobicity (H0, 812.4), oil absorption capacity (7.4 g g-1), foaming capacity (100%) and emulsifying activity (58.2 m2 g-1) were higher than that of SPI. However, the thermal stability (Td, 87.4 °C), foaming stability (75%) and emulsifying stability (26.3 min) of the MPI were weaker than that of the SPI. MMP was a pentamer with a molecular weight of 22 kDa and rich in ß-sheets. MMP could significantly enhance the phagocytic capacity and promote the NO, TNF-α and IL-6 secretion of RAW 264.7 cells, involving toll-like receptor 4 and complement receptor 3 mainly.


Assuntos
Fatores Imunológicos/química , Lepidium/química , Proteínas de Plantas/química , Animais , Interações Hidrofóbicas e Hidrofílicas , Fatores Imunológicos/farmacologia , Interleucina-6/genética , Interleucina-6/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Peso Molecular , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/farmacologia , Estrutura Secundária de Proteína , Células RAW 264.7 , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia
16.
J Agric Food Chem ; 67(24): 6748-6756, 2019 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-31136167

RESUMO

A 11 kDa antifungal protein FEAP was purified from buckwheat ( Fagopyrum esculentum) seed extract with a procedure involving (NH4)2SO4 precipitation and chromatography on SP-Sepharose, Affi-gel blue gel, Mono S, and Superdex peptide. Its N-terminal sequence was AQXGAQGGGAT, resembling those of buckwheat peptides Fα-AMP1 and Fα-AMP2. FEAP exhibited thermostability (20-100 °C) and acid resistance (pH 1-5). Its antifungal activity was retained in the presence of 10-150 mmol/L of K+, Mn2+, or Fe3+ ions, 10-50 mmol/L of Ca2+ or Mg2+ ions, and 50% methanol, 50% ethanol, 50% isopropanol, or 50% chloroform. Its half-maximal inhibitory concentrations toward spore germination and mycelial growth in Botrytis cinerea were 79.9 and 236.7 µg/mL, respectively. Its antifungal activity was superior to the fungicide cymoxanil mancozeb (248.1 µg/mL). FEAP prevented B. cinerea from infecting excised leaves, intact leaves, and isolated fruits of cherry tomato. Its mechanism involved induction of an increase in cell membrane permeability and a decrease in mitochondrial membrane potential.


Assuntos
Botrytis/fisiologia , Fagopyrum/química , Fungicidas Industriais/farmacologia , Lycopersicon esculentum/microbiologia , Doenças das Plantas/prevenção & controle , Proteínas de Plantas/farmacologia , Motivos de Aminoácidos , Botrytis/efeitos dos fármacos , Frutas/microbiologia , Fungicidas Industriais/química , Fungicidas Industriais/isolamento & purificação , Doenças das Plantas/microbiologia , Proteínas de Plantas/química , Proteínas de Plantas/isolamento & purificação
17.
J Agric Food Chem ; 67(23): 6633-6641, 2019 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-31099573

RESUMO

Ferritin is a multisubunit protein with a hollow interior interface and modifiable surfaces. In this study, the manothermosonication (MTS) technology was applied to apo-red bean seed ferritin (apoRBF) to produce the MTS-treated apoRBF (MTFS). MTS treatment (200 kPa, 50 °C, and 40 s) maintained the spherical morphology of apoRBF (12 nm), but reduced the content of α-helix structure and increased the content of random coil structure, and correspondingly decreased the ferritin stability. The MTS treatment also affected the ferritin's iron storage function by decreasing its iron oxidative deposition activity and increasing the iron release activity. Importantly, the disassembly and reassembly properties of the MTFS induced by pH changes were retained, which facilitated its usage in encapsulation of tea polyphenol-epigallocatechin gallate (EGCG) into the ferritin by a relatively benign pH conversion routine (pH 3.0/6.8). In addition, the water solubility of the MTFS was increased, leading to the improved encapsulation efficiency of the EGCG molecules. This study will facilitate the ferritin modification and functionalization by MTS to design a protein variant to be used as new scaffold for iron and bioactive compounds.


Assuntos
Apoferritinas/química , Apoproteínas/química , Portadores de Fármacos/química , Fabaceae/química , Ferro/química , Proteínas de Plantas/química , Sonicação/métodos , Apoferritinas/isolamento & purificação , Apoproteínas/isolamento & purificação , Catequina/análogos & derivados , Catequina/química , Portadores de Fármacos/isolamento & purificação , Concentração de Íons de Hidrogênio , Oxirredução , Proteínas de Plantas/isolamento & purificação , Estabilidade Proteica , Solubilidade , Sonicação/instrumentação
18.
Nat Commun ; 10(1): 2222, 2019 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-31110237

RESUMO

Substrates associate and products dissociate from enzyme catalytic sites rapidly, which hampers investigations of their trajectories. The high-resolution structure of the native Hordeum exo-hydrolase HvExoI isolated from seedlings reveals that non-covalently trapped glucose forms a stable enzyme-product complex. Here, we report that the alkyl ß-D-glucoside and methyl 6-thio-ß-gentiobioside substrate analogues perfused in crystalline HvExoI bind across the catalytic site after they displace glucose, while methyl 2-thio-ß-sophoroside attaches nearby. Structural analyses and multi-scale molecular modelling of nanoscale reactant movements in HvExoI reveal that upon productive binding of incoming substrates, the glucose product modifies its binding patterns and evokes the formation of a transient lateral cavity, which serves as a conduit for glucose departure to allow for the next catalytic round. This path enables substrate-product assisted processive catalysis through multiple hydrolytic events without HvExoI losing contact with oligo- or polymeric substrates. We anticipate that such enzyme plasticity could be prevalent among exo-hydrolases.


Assuntos
Domínio Catalítico , Glucosidases/metabolismo , Modelos Moleculares , Proteínas de Plantas/metabolismo , Biocatálise , Cristalografia por Raios X , Ensaios Enzimáticos/métodos , Glucosidases/química , Glucosidases/isolamento & purificação , Glicosídeos/metabolismo , Hordeum/metabolismo , Simulação de Dinâmica Molecular , Ressonância Magnética Nuclear Biomolecular , Proteínas de Plantas/química , Proteínas de Plantas/isolamento & purificação , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Plântula/metabolismo , Especificidade por Substrato
19.
Comput Biol Chem ; 80: 195-205, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30978571

RESUMO

The GRAS gene family is a class of plant-specific transcription factors which play pivotal roles in the regulation of plant growth and development. At present, the GRAS gene family has been completely identified in Arabidopsis thaliana, however, there are no systematic research reports in potato. In the present study, we obtained an overview of the GRAS gene family including gene structure, gene expression, chromosome mapping and phylogenetic analysis, and 52 StGRASs were identified in the potato by bioinformatics analysis, which could be divided into eight subfamilies based on phylogeny. More than 90% of genes do not contain introns and the StGRAS family major function is protein binding according to gene ontology analysis (GO).The tissue specific expression analysis showed that StGRAS3, StGRAS35 and StGRAS50 gene had the higher expression in roots, stems and leaves compared with other StGRAS, StGRAS9 and StGRAS28 genes were responded to plant hormones IAA, ABA and GA3 treatment. The result could provide a basis for further studying the function of GRAS genes and GRAS-mediated signal transduction pathways in potato.


Assuntos
Genes de Plantas , Proteínas de Plantas/genética , Solanum tuberosum/genética , Fatores de Transcrição/genética , Arabidopsis/genética , Mapeamento Cromossômico , Cromossomos de Plantas , Perfilação da Expressão Gênica , Lycopersicon esculentum/genética , Família Multigênica , Oryza/genética , Filogenia , Proteínas de Plantas/isolamento & purificação , Fatores de Transcrição/isolamento & purificação
20.
Food Microbiol ; 82: 504-514, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31027812

RESUMO

As a result of the rapidly growing human population, reducing post-harvest crop losses of cereals due to microbial pests has major importance. Plant defensins have the potential to fulfil these demands, being highly specific and efficient antimicrobial agents. Hence, this study aimed to extract and characterise a peptide from cowpea seeds and investigate its antifungal performance. After extraction and partial purification, N-terminal sequencing was used to identify the primary peptide in the extract as cowpea-thionin II. Antifungal activity in vitro was found against Fusarium culmorum (MIC = 50 µg/mL), but Aspergillus niger and Penecillium expansum showed an MIC > 500 µg/mL. The extract was resistant against heat treatment (100 °C, 15 min) but lost its antifungal activity in presence of cations (Na+, K+, Ca2+ and Mg2+, respectively). Membrane permeabilization of fungal hyphae was evident at 25 µg/mL, while induction of oxidative stress only had minor contribution to the antifungal performance. The extract did not induce haemolysis at all concentrations tested (up to 200 µg/mL). Finally, it was successfully used to protect stored wheat grains from fungal spoilage (determined via ergosterol content) when applied at 100 µg/mL. In conclusion, the defensin Cp-thionin II showed the potential for future application as food bio-preservative.


Assuntos
Antifúngicos/farmacologia , Conservantes de Alimentos/farmacologia , Fungos/efeitos dos fármacos , Proteínas de Plantas/farmacologia , Tioninas/farmacologia , Vigna/química , Antifúngicos/química , Antifúngicos/isolamento & purificação , Cátions , Permeabilidade da Membrana Celular/efeitos dos fármacos , Defensinas/química , Defensinas/isolamento & purificação , Defensinas/farmacologia , Grão Comestível/microbiologia , Ergosterol/análise , Ergosterol/metabolismo , Microbiologia de Alimentos , Conservantes de Alimentos/química , Conservantes de Alimentos/isolamento & purificação , Fungos/metabolismo , Fungos/fisiologia , Temperatura Alta , Hifas/efeitos dos fármacos , Hifas/metabolismo , Hifas/fisiologia , Testes de Sensibilidade Microbiana , Proteínas de Plantas/química , Proteínas de Plantas/isolamento & purificação , Estabilidade Proteica , Sementes/química , Tioninas/química , Tioninas/isolamento & purificação
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