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1.
J Pharm Biomed Anal ; 236: 115686, 2023 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-37690189

RESUMO

This study investigated the seasonal variation (over seven months) of phenolic acid (PHA) components in different parts of Centaurea sp. The primary objective was to determine the pattern of variation, while the secondary objective was to identify which month or growth stage provides a greater total PHA content or percentage of bioactive components. Different patterns of seasonal variations were highlighted for the different PHA components and their classes (hydroxybenzoic and -cinnamic acids) in different parts of the plant. The leaves exclusively provided the highest PHA contents, with maximum values reached in April (1368.06 µg/g). The major hydroxybenzoic acid derivatives (HBAs) identified in the leaves were vanillic acid (VaA) "154.18-374.06 µg/g" and protocatechuic acid (PA) "9.37-595.61 µg/g", while the major hydroxycinnamic acid derivatives (HCAs) were p-coumaric acid (p-CoA) "109.35-261.77 µg/g", m-coumaric acid (m-CoA) "10.22-70.57 µg/g", and ferulic acid (FeA) "35.54-109.13 µg/g". The maximum percentage of PA was obtained in April "595.61 µg/g", while the maximum p-CoA content was obtained in January "261.77 µg/g". Therefore, the leaves can be recommended as the optimal source of PHAs. If there is a specific interest in certain PHA components, we recommend collection in either January or April. Multivariate statistical analysis (PCA & AHC) showed the existence of two main clusters. The first cluster comprised the leaves, distinguished by the highest VaA, PA, and p-CoA contents. The second cluster comprised roots and the root bark samples. This study provides information on the development of PHAs in different parts of Centaurea sp. and explores potential applications. It will be of considerable interest for determining the optimal harvesting time of shrub species used for their medicinal properties and bio-active phenolic contents.

2.
J Sci Food Agric ; 102(2): 813-822, 2022 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-34223652

RESUMO

BACKGROUND: 'Karaerik' is a novel table grape (Vitis vinifera L.) native to Turkey and widely cultivated in areas bordering the city of Erzincan. Because of the demonstrated beneficial effects on human health of the grape phenolic composition, the aim of this work was to conduct a detailed profiling of non-anthocyanin phenolic fractions from different grape tissues of the 'Karaerik' table grape. Both qualitative and quantitative characterization of phenolic compounds were achieved using high-performance liquid chromatography-diode array detection-electrospray ionization-tandem mass spectrometry. Total phenolic content and oxygen radical absorbance capacity were also determined to evaluate the antioxidant properties of this table grape. RESULTS: A high number of non-anthocyanin phenolic compounds was identified in 'Karaerik' table grape skins and seeds, including 11 flavonols, six hydroxycinnamic acid derivatives, two stilbenes, several monomeric and dimeric flavan-3-ols and proanthocyanidins. Quercetin-type derivatives dominated the flavonol profile of grape skins, followed by myricetin type. Tartaric acid esters of three acids (caffeic, coumaric and ferulic acids) were the main hydroxycinnamic acid derivatives in this cultivar. Qualitative and quantitative differences were observed in flavan-3-ol composition among the grape tissues. Proanthocyanidins were the most abundant class of phenolic compounds in 'Karaerik' grapes, being mainly located in seeds. Higher antioxidant capacity values were determined in grape seeds, in correlation with the total phenolic content. CONCLUSION: These results provide useful information for a better understanding of phenolic antioxidants from the 'Karaerik' table grape and will contribute to promoting the varietal identity and health-related properties of this fruit. © 2021 Society of Chemical Industry.


Assuntos
Antioxidantes/química , Fenóis/química , Extratos Vegetais/química , Vitis/química , Cromatografia Líquida de Alta Pressão , Frutas/química , Turquia
3.
Andrologia ; 53(3): e13985, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33474739

RESUMO

The study aimed to investigate the effects of dexmedetomidine against ischaemia-reperfusion injury occurring after priapism in a model of induced-priapism in rats. A total of 18 male rats were randomised into three groups. Group 1 was the control group. A priapism model was performed rats in Group 2 and then ischaemia-reperfusion injury was evaluated. Group 3 had similar procedures to the rats in Group 2. Rats in Group 3 additionally had 100 µg/kg dexmedetomidine administered intraperitoneally immediately after reperfusion. Blood and tissue samples were analysed. Biochemical analysis of blood samples revealed a decrease in the levels of the pro-inflammatory cytokines including interleukin-1 beta (IL-1 Beta), interleukin-6 (IL-6), and tumour necrosis factor-alpha (TNF-alpha) in Group 3 compared to Group 2 (p:.04, p:.009 and p:.009, respectively). Similarly, the highest malondialdehyde (MDA) level was in Group 2 (p:.002). The levels of antioxidant enzymes superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were significantly higher in Group 3 than that of Group 2 (p:.037 and p:.045, respectively). Direct microscopic examinations revealed positive changes in desquamation, oedema, inflammation and vasocongestion scores in Group 3 compared to Group 2 (p:.007, p:.008, p:.007 and p:.006, respectively). Dexmedetomidine has a protective effect against ischaemia-reperfusion injury in penile tissue.


Assuntos
Dexmedetomidina , Priapismo , Traumatismo por Reperfusão , Animais , Dexmedetomidina/farmacologia , Dexmedetomidina/uso terapêutico , Humanos , Masculino , Malondialdeído , Priapismo/etiologia , Ratos , Traumatismo por Reperfusão/prevenção & controle , Superóxido Dismutase
4.
Methods Enzymol ; 599: 355-386, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29746246

RESUMO

Nitrogenase is a complex, bacterial enzyme that catalyzes the ATP-dependent reduction of dinitrogen (N2) to ammonia (NH3). In its most prevalent form, it consists of two proteins, the catalytic molybdenum-iron protein (MoFeP) and its specific reductase, the iron protein (FeP). A defining feature of nitrogenase is that electron and proton transfer processes linked to substrate reduction are synchronized by conformational changes driven by ATP-dependent FeP-MoFeP interactions. Yet, despite extensive crystallographic, spectroscopic, and biochemical information on nitrogenase, the structural basis of the ATP-dependent synchronization mechanism is not understood in detail. In this chapter, we summarize some of our efforts toward obtaining such an understanding. Experimental investigations of the structure-function relationships in nitrogenase are challenged by the fact that it cannot be readily expressed heterologously in nondiazotrophic bacteria, and the purification protocols for nitrogenase are only known for a small number of diazotrophic organisms. Here, we present methods for purifying and characterizing nitrogenase from a new model organism, Gluconacetobacter diazotrophicus. We also describe procedures for observing redox-dependent conformational changes in G. diazotrophicus nitrogenase by X-ray crystallography and electron paramagnetic resonance spectroscopy, which have provided new insights into the redox-dependent conformational gating processes in nitrogenase.


Assuntos
Gluconacetobacter/enzimologia , Nitrogenase/química , Cristalização/métodos , Cristalografia por Raios X/métodos , Espectroscopia de Ressonância de Spin Eletrônica/métodos , Transporte de Elétrons , Ensaios Enzimáticos/métodos , Gluconacetobacter/química , Gluconacetobacter/metabolismo , Modelos Moleculares , Molibdoferredoxina/química , Molibdoferredoxina/isolamento & purificação , Molibdoferredoxina/metabolismo , Nitrogenase/isolamento & purificação , Nitrogenase/metabolismo , Oxirredução , Conformação Proteica
5.
J Prev Med Hyg ; 58(2): E105-E113, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28900350

RESUMO

INTRODUCTION: Being overweight and obesity are a growing problem and are often related to a lack of physical activity among younger people. This study aims to describe the prevalence of weight disorders in Belgian schoolchildren. Secondly, this study examines the association between physical activity, weight disorders and the interest in food and sciences. METHODS: We examined 525 children aged between 8 and 18 years old, who attended the Brussels Food Fair or the Belgian Science Day in 2013. They completed a standardized questionnaire about lifestyle and physical activity. Their weight, height, blood pressure and waist circumference were measured. The physical condition of participants was estimated using the Ruffier test. RESULTS: The average age of all participants was 11.2 years (95% CI: 8.7-13.7), the prevalence of being overweight and obesity was 16.3% and 5.4%, respectively. For all participants in the representative group the affiliation to a sports club was associated with a normal weight (P < 0.05). According to this study, the kind of transportation to school (foot/bike or car/bus) had no effect on their body mass index (BMI). Neither was there a significant relationship between the physical activity of the children and the result of their Ruffier test. CONCLUSIONS: The prevalence of being overweight and obesity in children aged 8 to 18 years is alarming. Membership to a sports club was linked significantly to a normal weight and a lower prevalence of being overweight and obesity.


Assuntos
Atitude Frente a Saúde , Ciências da Nutrição Infantil , Exercício Físico , Alimentos , Sobrepeso/epidemiologia , Obesidade Infantil/epidemiologia , Adolescente , Antropometria , Bélgica/epidemiologia , Criança , Feminino , Humanos , Estilo de Vida , Masculino , Prevalência , Inquéritos e Questionários
6.
Dalton Trans ; 44(31): 13867-80, 2015 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-26146028

RESUMO

A powdered La2CoMnO6 double perovskite was synthesized by the solid-state reaction method, and its crystal structure was investigated by (mode-crystallography) Rietveld analysis using X-ray and neutron powder diffraction data. La2CoMnO6 material is a monoclinic perovskite at room temperature, adopting the space group P21/n (a(-)a(-)b(+)), , c ≈ 2ap and Z = 2. The P21/n phase can be described effectively by three distortion modes (GM4(+), X3(+), X5(+)) of the Fm3[combining macron]m (a(0)a(0)a(0)) parent phase. The comparative study of the material and those in the Ln2CoMnO6 and Ln2NiMnO6 families has shown a general trend in nearly all the materials, has served to select a common direction in the sub-space spanned by X5(+), tri-linearly coupled to the order parameters of the cubic to monoclinic first order phase transition. This direction has been used to parametrize the refinements and to perform reliable refinements in the high-temperature intermediate distorted trigonal phase, R3[combining macron] (a(-)a(-)a(-)), for which only one effectively acting irrep has been deduced: GM5(+), physically a tilt of the oxygen sharing octahedra of Co and Mn. Its temperature evolution up to the prototype cubic phase has been fitted in the framework of the Landau Theory of Phase Transitions, showing a behavior typical of a tricritical point. The low-temperature neutron powder diffraction data have served to solve the magnetic structure: three indistinguishable ferromagnetic models with the space groups P21/n and P2/n' are proposed.

7.
Int J Food Sci Nutr ; 62(4): 328-35, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21166481

RESUMO

The fatty acid and amino acid compositions of 11 mushroom species commonly consumed were collected from the East Black Sea region of Turkey and analyzed. All species were characterized by a high content of linoleic acid (C18:2n-6) and glutamic acid. The highest content of linoleic acid (78.0%) and glutamic acid (29.4 µg/mg dry weight [d.w.]) was found in Agaricus arvensis and the lowest in Cantharellus tubaeformis, 19.8% and 10.9 µg/mg d.w., respectively. The average content of amino acids for all species was 148 µg/mg d.w. Overall, these results demonstrate that the 11 different kinds of wild edible mushrooms gathered from the region represent substantial sources of fatty acids and amino acids that are essential in the diet of humans. Quality of the mushroom protein compares favorably with the FAO/WHO Standard. The present study demonstrates that macrofungi from the East Black Sea region (Turkey) are a good source of many nutrients essential to human well-being.


Assuntos
Agaricales/química , Aminoácidos/análise , Gorduras na Dieta/análise , Proteínas Alimentares/análise , Ácidos Graxos/análise , Agaricales/classificação , Mar Negro , Humanos , Valor Nutritivo , Especificidade da Espécie , Turquia
8.
Acta Crystallogr B ; 66(Pt 2): 109-16, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20305344

RESUMO

Single crystals of Sr(2)CaWO(6) have been prepared by sintering at high temperature. Powder samples were compressed into rods and heated up to 1953 K. This seems a promising new route for further studies of the structure and physical properties of double perovskites. The structural model of Sr(2)CaWO(6) includes a quantitative description of the twinning shown by the diffraction pattern that should be present in almost any single-crystal specimen for this type of compound.

9.
Plant Foods Hum Nutr ; 64(4): 286-92, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19763833

RESUMO

Although the fruit of the carob tree (Ceratonia siliqua L. Fabaceae) is nutritious and widely available in Turkey, especially in West and South Anatolia, much remains to be learned about its nutrient composition. The main goal of our study was to determine if there are differences in the content of certain nutrients in commercially-prepared carob flour (CPCP) and domestic or home-prepared carob powder (HPCP). Sucrose was the main sugar in CPCP and HPCP. Total protein was 40% lower in CPCP than HPCP due mainly to decreases in the content of several essential amino acids. However, except for lysine in CPCP, HPCP and CPCP compared favourably to a WHO protein standard. There were large differences in terms of their content of the two essential fatty acids, linoleic and alpha-linolenic acid, and the linoleic acid/alpha-linolenic acid ratio was 3.6 for CPCP, and 6.1 for HPCP. Manganese and iron were 2.5-fold higher in HPCP than CPCP. This study demonstrates that carob flour prepared in either the household or industrially is a good source of many, but not all essential nutrients, and that commercial processing of carob fruit into flour seems to affect its content of several important nutrients.


Assuntos
Fabaceae/química , Análise de Alimentos , Manipulação de Alimentos , Valor Nutritivo , Preparações de Plantas/química , Ácidos Graxos Essenciais/análise , Indústria Alimentícia , Frutas/química , Humanos , Ferro/análise , Lisina/análise , Manganês/análise , Proteínas/análise , Valores de Referência , Sacarose/análise , Turquia
10.
J Agric Food Chem ; 57(12): 5244-9, 2009 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-19480388

RESUMO

High-performance liquid chromatography (HPLC) combined with diode array (DAD) and electrospray ionization mass spectrometric (ESI-MS) detections were used to characterize anthocyanins in the berries of Vaccinium arctostaphylos L. The dark purple-black berries were collected from five Caucasian blueberry populations in northeastern Turkey. The HPLC-DAD profile consisted of 19 anthocyanin peaks, but HPLC-ESI-MS revealed fragment ion patterns of 26 anthocyanins. Delphinidin, cyanidin, petunidin, peonidin, and malvidin were all glycosylated with four different monosaccharide moieties (galactose, glucose, arabinose, and xylose) with the first two also conjugated with rhamnose. Furthermore, anthocyanidin disaccharides, tentatively identified as anthocyanidin sambubiosides, were characteristic for these berries. The mean content of the total anthocyanins was 1420 mg/100 g dry weight. The most predominant anthocyanidins were delphinidin (41%), petunidin (19%), and malvidin (19%). Glucose was the most typical (61%) sugar moiety. This study revealed that wild Caucasian blueberries contain an abundance of bioactive anthocyanins and thus are ideal for various functional food purposes.


Assuntos
Antocianinas/química , Mirtilos Azuis (Planta)/química , Humanos , Extratos Vegetais/química , Espectrometria de Massas por Ionização por Electrospray , Turquia
11.
Acta Crystallogr Sect E Struct Rep Online ; 64(Pt 8): o1425, 2008 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-19079747

RESUMO

The title compound, C(13)H(16)F(6)NO(5)PS, is of inter-est with respect to inhibition of serine hydro-lases. Its structure contains a 1.8797 (13) ŠP-C bond and two inter-molecular N-H⋯O=P hydrogen bonds, resulting in centrosymmetric dimers. An intra-molecular N-H⋯O=P hydrogen bond is also present.

12.
Rheumatol Int ; 28(7): 677-83, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18092169

RESUMO

The aim of this study is to assess the reliability and validity of the Western Ontario and McMaster University Osteoarthritis Index (WOMAC) in Moroccan patients with knee osteoarthritis. The WOMAC was translated and back translated to and from dialectal Arabic, pre-tested and reviewed by a committee following the Guillemin criteria. The Moroccan version of the WOMAC was administered twice during a 24-48 h interval to 71 Moroccan patients with symptomatic knee osteoarthritis, fulfilling the revised criteria of the American College of Rheumatology. The test-retest reliability was assessed using intra-class correlation coefficient, and the Bland and Altman method. Internal consistency was assessed by Cronbach's alpha coefficient. Construct validity was tested by correlating the WOMAC subscales with visual analogic scale (VAS) of pain, VAS of handicap, maximum distance walked and clinical characteristics. The Moroccan version of the WOMAC showed good reliability, with ICC values of the three dimensions: pain, stiffness and physical function being 0.80, 0.77 and 0.89, respectively. Bland and Altman analysis showed that means of differences did not differ significantly from 0 and that no systematic trend was observed. Internal consistency with Cronbach's alpha for pain was found to be 0.76, and its equivalents for stiffness and physical function subscales were evaluated at 0.76, 0.90, respectively. Construct validity showed statistically significant correlation with all WOMAC subscales and VAS of pain (rho=0.38, 0.42, 0.63 respectively, P<0.01). Correlation between VAS handicap (rho=0.38 P<0.001) and maximum distance walked (rho=-0.40, P<0.01) was observed with physical function subscale. There was no correlation between age, duration of disease, BMI and severity of pain and physical function in knee OA. The Moroccan version of the WOMAC is a comprehensible, reliable, and valid instrument to measure outcome in patients with knee OA.


Assuntos
Osteoartrite do Joelho/diagnóstico , Índice de Gravidade de Doença , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Articulação do Joelho/fisiopatologia , Masculino , Pessoa de Meia-Idade , Osteoartrite do Joelho/fisiopatologia , Medição da Dor
13.
J Forensic Sci ; 51(6): 1372-5, 2006 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-17199623

RESUMO

Bitemark determination in forensic odontology is commonly performed by comparing the morphology of the dentition of the suspect with life-sized photographs of injury on the victim's skin using transparent overlays or computers. The purpose of this study is to investigate the suitability of two new different methods for identification of bitemarks by digital analysis. A sample of 50 volunteers was asked to make experimental bitemarks on the arms of each other. Stone study casts were prepared from upper and lower dental arches of each volunteer. The bitemarks and the study casts were photographed; the photos were entered into the computer and Adobe Photoshop software program was applied to analyze the results. Two methods (2D polyline and Painting) of identification were used. In the 2D polyline method, fixed points were chosen on the tips of the canines and a straight line was drawn between the two fixed points in the arch (intercanine line). Straight lines passing between the incisal edges of the incisors were drawn vertically on the intercanine line; the lines and angles created were calculated. In the painting method, identification was based on canine-to-canine distance, tooth width and the thickness, and rotational value of each tooth. The results showed that both methods were applicable. However, the 2D polyline method was more convenient to use and gave prompt computer-read results, whereas the painting method depended on the visual reading of the operator.


Assuntos
Mordeduras Humanas/patologia , Odontologia Legal/métodos , Processamento de Imagem Assistida por Computador , Adulto , Humanos , Modelos Dentários , Fotografia Dentária , Software
14.
J Agric Food Chem ; 53(21): 8116-22, 2005 Oct 19.
Artigo em Inglês | MEDLINE | ID: mdl-16218652

RESUMO

The aim of this study was the qualitative and quantitative determination of free, ester, glycoside, and ester-bound phenolic acids in the blueberry (Vaccinium arctostaphylos L.) fruit. A method for the determination of the profile of phenolic acids of four different phenolic fractions in the fruit was developed using high-performance liquid chromatography-mass spectrometry (HPLC-MS). Thirteen compounds (gallic, protocatechuic, p-hydroxybenzoic, m-hydroxybenzoic, gentisic, chlorogenic, p-coumaric, caffeic, ferulic, syringic, sinapic, salicylic, and trans-cinnamic acids) were identified and quantified in the berry. These experimental results showed that the predominant phenolic acid in the fruit of V. arctostaphylos is caffeic acid in free and insoluble ester-bound forms and p-coumaric acid in soluble ester and glycoside forms. Seven phenolic acids were identified as hydroxybenzoic acid derivatives (HBAs) and four as hydroxycinnamic acid derivatives (HCAs). Total content of HBAs and HCAs in the four phenolic fractions constituted 30.1 and 69.9% of the free, 27.9 and 72.1% of the ester, 24.7 and 75.3% of the glycoside, and 51.7 and 48.3% of the ester-bound forms, respectively. Total phenolics as the sum of individual phenolic acids identified is 698.5 ng/g of fresh weight (fw) for the free, 3399.2 ng/g of fw for the ester, 3522.1 ng/g of fw for the glycoside, and 3671.6 ng/g of fw for the ester-bound phenolic fractions. The present results were compared with reported levels of phenolic acids in the fruits of different Vaccinium species. These data suggest that the fruit can be considered as a potentially good dietary source of phenolic acids.


Assuntos
Frutas/química , Hidroxibenzoatos/isolamento & purificação , Vaccinium/química , Cromatografia Líquida de Alta Pressão , Hidroxibenzoatos/análise , Espectrometria de Massas
15.
Plant Physiol ; 127(4): 1595-606, 2001 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-11743104

RESUMO

To understand primary cell wall assembly in Arabidopsis, we have focused on identifying and characterizing enzymes involved in xyloglucan biosynthesis. Nine genes (AtFUT2-10) were identified that share between 47% and 62% amino acid similarity with the xyloglucan-specific fucosyltransferase AtFUT1. Reverse transcriptase-PCR analysis indicates that all these genes are expressed. Bioinformatic analysis predicts that these family members are fucosyltransferases, and we first hypothesized that some may also be involved in xyloglucan biosynthesis. AtFUT3, AtFUT4, and AtFUT5 were expressed in tobacco (Nicotiana tabacum L. cv BY2) suspension culture cells, and the resulting proteins did not transfer fucose (Fuc) from GDP-Fuc to tamarind xyloglucan. AtFUT3, AtFUT4, and AtFUT5 were overexpressed in Arabidopsis plants. Leaves of plants overexpressing AtFUT4 or AtFUT5 contained more Fuc than wild-type plants. Stems of plants overexpressing AtFUT4 or AtFUT5 contained more xylose, less arabinose, and less galactose than wild-type plants. We suggest that the AtFUT family is likely to include fucosyltransferases important for the synthesis of wall carbohydrates. A targeted analysis of isolated cell wall matrix components from plants altered in expression of these proteins will help determine their specificity and biological function.


Assuntos
Arabidopsis/genética , Fucosiltransferases/genética , Sequência de Aminoácidos , Arabidopsis/enzimologia , Parede Celular/enzimologia , Parede Celular/metabolismo , Células Cultivadas , Fucosiltransferases/metabolismo , Perfilação da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Dados de Sequência Molecular , Família Multigênica , Fenótipo , Filogenia , Alinhamento de Sequência , Galactosídeo 2-alfa-L-Fucosiltransferase
16.
Biochem J ; 347 Pt 3: 857-64, 2000 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-10769192

RESUMO

Polypeptide assemblies cross-linked by S-S bonds (molecular mass>200 kDa) and single polypeptides folded with internal S-S cross-links (<41 kDa) have been detected by SDS/PAGE in particulate membranes and soluble extracts of developing cotyledons of nasturtium (Tropaeolum majus L.). When first prepared from fruit homogenates, these polypeptides were found to bind reversibly to UDP-Gal (labelled with [(14)C]Gal or [(3)H]uridine), and to co-precipitate specifically with added xyloglucan from solutions made with 67% ethanol. Initially, the bound UDP-[(14)C]Gal could be replaced (bumped) by adding excess UDP, or exchanged (chased) with UDP-Gal, -Glc, -Man or -Xyl. However, this capacity for turnover was lost during incubation in reaction media, or during SDS/PAGE under reducing conditions, even as the glycone moiety was conserved by autoglycosylation to form a stable 41 kDa polypeptide. Polyclonal antibodies raised to a similar product purified from Arabidopsis bound to all the labelled nasturtium polypeptides in immunoblotting tests. The antibodies also inhibited the binding of nasturtium polypeptides to UDP-Gal, the uptake of UDP-[(14)C]Gal into intact nasturtium membrane vesicles and the incorporation of [(14)C]Gal into nascent xyloglucan within these vesicles. This is the first direct evidence that these polypeptides facilitate the channelling of UDP-activated sugars from the cytoplasm through Golgi vesicle membranes to lumenal sites, where they can be used as substrates for glycosyltransferases to synthesize products such as xyloglucan.


Assuntos
Proteínas de Arabidopsis , Brassicaceae/química , Frutas/química , Glucanos , Glicoproteínas/metabolismo , Peptídeos/metabolismo , Proteínas de Plantas/metabolismo , Açúcares de Uridina Difosfato/metabolismo , Xilanos , Anticorpos/imunologia , Anticorpos/farmacologia , Arabidopsis/imunologia , Transporte Biológico/efeitos dos fármacos , Brassicaceae/citologia , Cetomacrogol/farmacologia , Precipitação Química , Reações Cruzadas , Dissulfetos/metabolismo , Frutas/citologia , Galactose/metabolismo , Glicoproteínas/química , Glicoproteínas/imunologia , Glicosilação/efeitos dos fármacos , Peso Molecular , Peptídeos/química , Peptídeos/imunologia , Proteínas de Plantas/química , Proteínas de Plantas/imunologia , Polissacarídeos/biossíntese , Polissacarídeos/metabolismo , Ligação Proteica/efeitos dos fármacos , Especificidade por Substrato , Difosfato de Uridina/metabolismo , Uridina Difosfato Galactose/metabolismo , Vacúolos/química , Vacúolos/efeitos dos fármacos , Vacúolos/metabolismo
17.
J Biol Chem ; 275(20): 15082-9, 2000 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-10747946

RESUMO

Pea microsomes contain an alpha-fucosyltransferase that incorporates fucose from GDP-fucose into xyloglucan, adding it preferentially to the 2-O-position of the galactosyl residue closest to the reducing end of the repeating subunit. This enzyme was solubilized with detergent and purified by affinity chromatography on GDP-hexanolamine-agarose followed by gel filtration. By utilizing peptide sequences obtained from the purified enzyme, a cDNA clone was isolated that encodes a 565-amino acid protein with a predicted molecular mass of 64 kDa and shows 62.3% identity to its Arabidopsis homolog. The purified transferase migrates at approximately 63 kDa by SDS-polyacrylamide gel electrophoresis but elutes from the gel filtration column as an active protein of higher molecular weight ( approximately 250 kDa), indicating that the active form is an oligomer. The enzyme is specific for xyloglucan and is inhibited by xyloglucan oligosaccharides and by the by-product GDP. The enzyme has a neutral pH optimum and does not require divalent ions. Kinetic analysis indicates that GDP-fucose and xyloglucan associate with the enzyme in a random order. N-Ethylmaleimide, a cysteine-specific modifying reagent, had little effect on activity, although several other amino acid-modifying reagents strongly inhibited activity.


Assuntos
Fucosiltransferases/metabolismo , Glucanos , Pisum sativum/enzimologia , Polissacarídeos/biossíntese , Xilanos , Sequência de Aminoácidos , Arabidopsis/enzimologia , Cromatografia de Afinidade , Cromatografia em Gel , Clonagem Molecular , Fucosiltransferases/genética , Fucosiltransferases/isolamento & purificação , Cinética , Dados de Sequência Molecular , Peso Molecular , Pisum sativum/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Galactosídeo 2-alfa-L-Fucosiltransferase
18.
FEBS Lett ; 442(2-3): 123-8, 1999 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-9928986

RESUMO

When maize calluses are grown in the presence of the RGD peptide, important morphological changes are observed indicating the presence of a likely RGD-binding receptor. Polyclonal antibodies generated against the human beta1 integrin subunit, the platelet integrin alphaIIbeta3 (P23) and antibodies specific for either the beta3 platelet chain or the alphaIIb polypeptide cross-react with glycoproteins in Western blot analyses. Immunoprecipitation assays indicate that this maize integrin-like protein shares structural similarities with the animal alphaIIbeta3 complex. We also show that AcAt2, a polyclonal antibody raised against Arabidopsis proteins purified on an RGD column, interacts with a maize protein.


Assuntos
Integrinas/análise , Oligopeptídeos/farmacologia , Zea mays/química , Zea mays/crescimento & desenvolvimento , Animais , Arabidopsis , Células CHO , Cromatografia de Afinidade , Cricetinae , Reações Cruzadas , Imunofluorescência , Glicoproteínas/química , Glicoproteínas/imunologia , Humanos , Soros Imunes , Integrina beta1/imunologia , Integrinas/imunologia , Integrinas/isolamento & purificação , Integrinas/metabolismo , Peso Molecular , Oligopeptídeos/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/imunologia , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/química , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/imunologia , Testes de Precipitina , Zea mays/citologia , Zea mays/efeitos dos fármacos
19.
Plant Physiol ; 118(3): 885-94, 1998 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-9808733

RESUMO

Young, developing fruits of nasturtium (Tropaeolum majus L.) accumulate large deposits of nonfucosylated xyloglucan (XG) in periplasmic spaces of cotyledon cells. This "storage" XG can be fucosylated by a nasturtium transferase in vitro, but this does not happen in vivo, even as a transitory signal for secretion. The only XG that is clearly fucosylated in these fruits is the structural fraction (approximately 1% total) that is bound to cellulose in growing primary walls. The two fucosylated subunits that are formed in vitro are identical to those found in structural XG in vivo. The yield of XG-fucosyltransferase activity from membrane fractions is highest per unit fresh weight in the youngest fruits, especially in dissected cotyledons, but declines when storage XG is forming. A block appears to develop in the secretory machinery of young cotyledon cells between sites that galactosylate and those that fucosylate nascent XG. After extensive galactosylation, XG traffic is diverted to the periplasm without fucosylation. The primary walls buried beneath accretions of storage XG eventually swell and lose cohesion, probably because they continue to extend without incorporating components such as fucosylated XG that are needed to maintain wall integrity.

20.
Eur J Biochem ; 253(3): 552-9, 1998 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-9654050

RESUMO

Using a polyclonal antibody (P23) generated against the human platelet integrin aIIb beta3 and a FITC-conjugate secondary antibody, fluorescence is observed at the surface of protoplasts isolated from Arabidopsis thaliana and Rubus fruticosus. Arabidopsis thaliana cells grown in suspension culture containing P23 and glycylarginylglycylaspartylserine (GRGDS), a synthetic peptide containing the RGD sequence found in many extracellular matrix adhesive proteins demonstrated aberrant cell wall/plasma membrane interactions and organization. When glycoproteins from these plants, purified on a concanavalin A Sepharose 4B, were subjected to SDS/PAGE and Western blotting, under reduced and non-reduced conditions, immunoblots probed with P23 revealed bands in both species. A shift in electrophoretic mobility is observed to different apparent molecular mass when no reducing agent is present. When purified by immunoaffinity chromatography on anti-aIIb beta3 Sepharose or Sepharose linked to the synthetic peptide D-Arg-Gly-Asp-Trp, the major antigenic components detected migrate at 30 kDa and 60 kDa in the first experiment and 60 kDa in the second one. Only the 60-kDa component is immunodetected with antibodies specific for either the beta3 platelet chain or the aIIb polypeptide, suggesting the presence of two polypeptides co-migrating. To address more precisely the structure of this complex in plants, competition assays were performed. A significant inhibition is observed with CS3 a monoclonal antibody that interacts with the complexed form aIIb beta3 but not the dissociated subunits. Further structural similarities with the animal aIIb beta3 complex is demonstrated with Western blotting detection after plant glycoproteins immunoprecipitation with CS3 in absence or presence of 5 mM EDTA to dissociate the complex. We also present data on the characterization of a polyclonal antibody, named AcAt2, raised against Arabidopsis glycocoproteins purified by affinity chromatography on a D-RGDW column and eluted with the same peptide, that specifically interacts with the animal aIIb beta3 receptor.


Assuntos
Arabidopsis/metabolismo , Glicoproteínas/biossíntese , Proteínas de Plantas/biossíntese , Proteínas de Plantas/química , Plantas/metabolismo , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/química , Sequência de Aminoácidos , Animais , Arabidopsis/citologia , Células Cultivadas , Cromatografia de Afinidade , Epitopos/análise , Epitopos/química , Glicoproteínas/química , Glicoproteínas/isolamento & purificação , Humanos , Immunoblotting , Integrinas/química , Modelos Moleculares , Peso Molecular , Oligopeptídeos/metabolismo , Células Vegetais , Proteínas de Plantas/isolamento & purificação , Conformação Proteica , Protoplastos/metabolismo , Homologia de Sequência de Aminoácidos
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