925 resultados para Ca2 -independent enzymatic activity
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Estudos demostram que a hiperalimentação no período pós-natal causa obesidade, alterações cardiometabólicas e resistência à insulina em longo prazo. O objetivo do estudo foi investigar as consequências da hiperalimentação na lactação nos corações de camundongos filhotes e adultos ao longo do desenvolvimento. Para induzir a hiperalimentação na lactação, o tamanho da ninhada foi reduzida a 3 filhotes machos no terceiro dia, grupo hiperalimentado (GH). O grupo controle (GC) permaneceu com 9 filhotes da lactação ao desmame. Avaliamos a massa corporal, gordura epididimária e retroperitoneal, morfologia hepática e cardíaca, ultraestrutura dos cardiomiócitos, peso do PVE/CT, glicemia de jejum, triglicerídeos, colesterol total, insulina plasmática e HOMA-IR. Analisamos o consumo de oxigênio das fibras cardíacas através da respirometria de alta resolução, atividade enzimática da PDH, CS e LDH no coração e glicogênio hepático. Biologia molecular, através das proteínas: IRβ, IRS1, pIRS1, PTP1B, PI3K, Akt, pAkt, GLUT1, GLUT4, AMPKα, pAMPKα, HKII, CPT1, UCP2, FABPm, CD36, PGC-1α, PPARα, 4HNE, complexos da CTE (I, II, III, IV e V), α-tubulina, GP91 e VADC. Diferenças entre os grupos analisadas por Two-Way ANOVA, com significância p<0,05. O GH apresentou aumento da massa corporal, gordura epididimária, retroperitoneal e colesterol total em todas as idades; glicemia de jejum, insulina, índice de HOMA-IR e triglicerídeos aos 21 e 90 dias. Aumento do índice de Lee aos 60 e 90 dias. GH apresentou diminuição: do IRβ e GLUT4 aos 21 e 60 dias; aumento do IRβ aos 90 dias; aumento do IRS1, PTP1B, aos 21 e 90 dias e da AKT, pAMPK/AMPK e GLUT1 aos 21 dias; diminuição da pIRS1/IRS1, PI3K, pAKT/AKT aos 21 e 90 dias; diminuição da HKII aos 21 dias e aumento aos 60 e 90 dias; aumento da PDH aos 90 dias; aumento da LDH aos 21 dias e redução aos 60 dias; aumento da CS aos 21 dias e diminuição aos 60 e 90 dias; aumento da oxidação de carboidratos aos 21 dias e redução aos 90 dias; diminuição na oxidação de ácidos graxos aos 60 e 90 dias. Adicionalmente, aumento do desacoplamento mitocondrial entre a fosforilação oxidativa e a síntese de ATP aos 60 e 90 dias. Diminuição da CPT1 e aumento da UCP2 aos 21 e 90 dias. Diminuição da PGC-1α aos 60 e 90 dias; da FABPm e CD36 em todas idades. Aumento da 4HNE aos 21 e diminuição aos 90 dias. Diminuição na expressão do mRNA para CPT1 aos 21, 60 dias. Diminuição na expressão do mRNA para PPARα e aumento na expressão do mRNA para UCP2 aos 21 dias; diminuição na expressão do mRNA para UCP2 ao 60 dias. Alterações morfológicas cardíacas e hepáticas, assim como na ultraestrutura dos cardiomiócitos, em todas as idades, maior conteúdo de glicogênio hepático aos 21 e 90 dias. Concluímos que a hiperalimentação na lactação levou à obesidade, com aumento da oxidação de glicose, alterações no metabolismo energético associadas à diminuição da sensibilidade à insulina, redução da capacidade oxidativa mitocondrial, levando ao desacoplamento e alteração da morfologia e ultraestrutura dos cardiomiócitos do desmame até a idade adulta.
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Traditionally tubers of cyperus (Cyperus rotundus) and its extracts have been used for alluring fish during harvesting in India. An experiment was conducted to evaluate its feeding stimulatory activity and effect on the growth of a commercially important freshwater fish, Cirrhinus mrigala. Three isonitrogenous and isocaloric formulated diets viz. plant ingredient based control and control supplemented with cyperus tuber (CS) at 1% and 5% levels were fed to the fingerlings of mrigal, C. mrigala (2.68+0.20 g) for a period of 45 days. The growth performance and the activity of metabolic enzymes, aspartate aminotransferase (AST) and alanine aminotransferase (ALT), in liver, gill and muscle tissues of mrigal were studied during every 15 days interval. Highest relative growth (72.28%) was obtained in the mrigal fed with the diet containing 5% cyperus (5% CS), while the relative growths were 66.18% and 43.40% for the fish fed with the 1% CS diet and control respectively. The activities of AST and ALT were significantly higher (p<0.01) in both 1% and 5% CS diets as compared to the control in all the tissues studied. Higher aminotransferase activities were observed in the tissues of 5% CS group than in those of 1% CS group throughout the experimental period. The observed higher enzymatic activity was concomitant with the higher growth rate in fish. The results suggested that cyperus tuber supplementation increased feed palatability and growth.
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The effect of sodium lactate is compared with sucrose + sorbitol + sodium tri-poly phosphate as cryoprotectant on gel forming ability & protein denaturation of croaker surimi during frozen storage at -20±2°C for 90 days was evaluated. The quality of Croaker surimi with 6% (w/v) sodium lactate was examined in terms of biochemical parameters of muscle protein, thaw drip, gel strength and calcium ATPase activity :.omparing with those of surimi added with sucrose/sorbitol & without additive as control. Both the cryoprotectants minimized the negative effects of frozen storage on physico-chemical traits of myofibrillar proteins which was evident from the biochemical and sensory parameters. The residual Ca2+ ATPase activity and gel strength of surimi with sodium lactate were higher than those of control throughout 90 days of storage. Ca2+ A TPase activity and gel strength found a high positive correlation. From the results, it was found that sodium lactate was equally effective in preservation of croaker muscle protein native structure during frozen storage as the sucrose/ sorbitol and also less sweet without any risk of maillard browning.
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Study on the biomarkers types to assess health status of marine ecosystems in environmental biomonitoring has an important value. Accordingly, accumulation of polycyclic aromatic hydrocarbons(PAHs) in sediment, water and tissues (liver and gill) of mudskipper(i.e. Boleophthalmus dussumieri) and some physiological responses like lysosomal membrane change performed on haemocytes, stability of red blood cell membrane and the Glutathione-S Transferase (GST) activity in the liver were measured in mudskipper. Samples were obtained from five sites along north western coast of the Persian Gulf (Khuzestan coast). Red blood cell membrane changes after different concentration of PAHs at different time was also studied to evaluate impact of PAHs compound on cell membrane. PAHs concentration was measured by HPLC method. The activity of GST enzyme was analysed by spectrophotometric method. Lysosomal membrane change was measured by NRR time method and stability of red blood cell membrane was evaluated by EOF test. Total PAH concentrations in the coastal sea water, the sediments, the liver and the gill tissues ranged between 0.80-18.34 μg/l, 113.50-3384.34 ng g-1 (dry weight), 3.99-46.64 ng g-1 dw and 3.11-17.76 ng g-1 dw, respectively. Highest PAHs pollution was found at Jafari while the lowest was detected at Bahrakan sampling sites. The lowest enzymatic activity was identified at Bahrakan (7.19 ± 1.541 nmol/mg protein/min), while the highest was recorded at Jafari (46.96 ± 7.877 nmol/mg protein/min). Comparative analysis of GST activity in the liver of mudskippers showed significant difference (p < 0.05) between the locations of Jafari and Bahrakan, and with other sites. Moreover, no significant difference was detected between the locations of Arvand, Zangi and Samayeli (p < 0.05). The mean RT was below 90 minutes in all sampling sites. Values of mean RT of the dye ranged from 34 (for the blood samples of mudskipper collected from Jafari site) to 78 minutes (for the blood samples of mudskipper collected from Bahrakan site). Spatial evaluation revealed the longest RT in fish from Bahrakan as compared with those from other sites. Preliminary results showed a significant difference (p < 0.05) among sampling sites except between Arvand and Zangi (p > 0.05). Osmotic fragility curves indicated that erythrocytes collected from mudskippers at Jafari were the most 009 fragile followed by Zangi> Arvand> Samayeli> and Bahrakan. The mean erythrocyte fragility was significantly higher at Jafari site (p < 0.05) when compared to other sites. Significant differences were found between the various sites (p < 0.05).The result indicated no significant differences between the control and treatments of mudskipper RBC exposed to field concentrations of PAHs (P>0.05). The results further indicated significant differences (P<0.05) between the control and treatments of mudskipper RBC exposed to acute. Potency Divisor concentrations. It is clear from the present result that chronic. Potency Divisor concentrations protect red cells against osmotic hemolysis. This study, however, showed that PAH concentrations in this region are not higher than the available standards. The findings showed that Lysosomal membrane destabilization, liver GST activities and fragility of red cell membrane are highly sensitive in the mudskipper, B. dussumieri. Thus, mudskipper perceived to be good sentinel organisms for PAH pollution monitoring. Sediment PAH concentrations were strongly correlated with biomarkers, indicating that PAH type pollutants were biologically available to fish. One of the possible risk assessment implications of this study is that biomarkers can be applied not only to characterize biological effects of pollution exposures, but also to determine the bioavailability of pollution in aquatic systems. The results also indicated that PAHs compound possess anti haemolytic property.
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为了探讨当今世界使用量最大的除草剂——草甘膦的土壤环境效应,本文采用室内模拟方法,较为系统地研究了我国4类土壤:褐土、黄绵土、风沙土和红壤,共11个土样中4种主要酶类(脲酶、转化酶、磷酸酶以及脱氢酶)活性与草甘膦间的关系,计算并得到了能够表征土壤轻度污染的生态剂量值ED10。结果表明:非缓冲液法较好地反映了土壤酶的实际情况;草甘膦总体上激活土壤脲酶、转化酶和脱氢酶活性,最大增幅分别为190%、1372%和42%;抑制磷酸酶活性,最大幅度为35%;磷酸酶与草甘膦间为完全抑制作用机理;激活脱氢酶活性揭示出草甘膦导致了土壤中微生物活性增强,从侧面反映出草甘膦是一种毒性较低的农药。计算获得4类供试土壤褐土、黄绵土、风沙土和红壤ED10值分别为168.3、438.5、35.1和141.4mg·kg-1;在一定程度上用土壤酶活性比生物来表征土壤污染程度更敏感。土壤性质对草甘膦的毒性有重要影响。
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Maize ribosome-inactivating protein (RIP) is a plant toxin that inactivates eukaryotic ribosomes by depurinating a specific adenine residue at the a-sarcin/ricin loop of 28S rRNA. Maize RIP is first produced as a proenzyme with a 25-amino acid internal inactivation region on the protein surface. During germination, proteolytic removal of this internal inactivation region generates the active heterodimeric maize RIP with full N-glycosidase activity. This naturally occurring switch-on mechanism provides an opportunity for targeting the cytotoxin to pathogen-infected cells. Here, we report the addition of HIV-1 protease recognition sequences to the internal inactivation region and the activation of the maize RIP variants by HIV-1 protease in vitro and in HIV-infected cells. Among the variants generated, two were cleaved efficiently by HIV-1 protease. The HIV-1 protease-activated variants showed enhanced N-glycosidase activity in vivo as compared to their un-activated counterparts. They also possessed potent inhibitory effect on p24 antigen production in human T cells infected by two HIV-1 strains. This switch-on strategy for activating the enzymatic activity of maize RIP in target cells provides a platform for combating pathogens with a specific protease.
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本文以黄土高原丘陵区陕北延安燕沟流域为例,研究了退耕地土壤有机碳、全氮和酶活性对植被恢复过程的响应。结果表明,随植被恢复年限的增加,植被盖度、多度和物种数均呈现先增加后减少然后又增加的趋势。同当年对照农地相比,随着植被恢复年限的增加,表层(0~20 cm)土壤有机碳、全氮、蔗糖酶、脲酶和碱性磷酸酶活性均明显增加,过氧化氢酶活性随植被恢复年限的增加变化不明显。同对照农地相比,表层土壤有机碳、全氮、蔗糖酶、脲酶和碱性磷酸酶活性分别增加了35%~184.6%、5.7%~157.4%、89.6%~566%、32.9%~331.2%和0.18%~184.8%。表层土壤有机碳和脲酶活性对植被恢复的响应是在植被恢复初期(0~8年),随植被恢复年限的增加而增加,植被恢复8~29年期间,随植被恢复年限的增加而减少,而植被恢复29 a以后,又随植被恢复年限的增加而增加。而表层土壤全氮、蔗糖酶和碱性磷酸酶活性对植被恢复的响应是在植被恢复初期(0~16年),随植被恢复年限的增加而增加,植被恢复16~29年期间,随植被恢复年限的增加而减少,植被恢复29 a以后,又随植被恢复年限的增加而增加。表层土壤蔗糖酶、脲酶和碱性磷酸酶活性与有机碳...
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研究了黄土塬区塬面不同套种作物的幼树苹果园土壤微生物的分布和酶活性。结果表明,果园土壤中微生物的分布在果树行与套种行中是不均匀的,细菌和真菌多为果树行多于套种行;土壤的过氧化氢酶,蔗糖酶,脲酶以及中性磷酸酶活性在套种小麦的果园中以套作行高于果树行,而套种烤烟则使脲酶和中性磷酸酶活性有所降低。套种豆类有利于土壤肥力的均衡发展。
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The conformational changes of bovine serum albumin (BSA) in the albumin:gold nanoparticle bioconjugates were investigated in detail by various spectroscopic techniques including UV-vis absorption, fluorescence, circular dichroism, and Fourier transform infrared spectroscopies. Our studies suggested that albumin in the bioconjugates that was prepared by the common adsorption method underwent substantial conformational changes at both secondary and tertiary structure levels. BSA was found to adopt a more flexible conformational state on the boundary surface of gold nanoparticles as a result of the conformational changes in the bioconjugates. The conformational changes at pH 3.8, 7.0, and 9.0, which corresponded to different isomeric forms of albumin, were investigated, respectively, to probe the pH effect on the conformational changes of BSA in the bioconjugates. The results showed that the pH of the medium influenced the changes greatly and that fluorescence and circular dichroism studies further indicated that the changes were larger at higher pH.
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A more stably dispersing of multi-wall carbon nanotube composite (noted as PDDA-MWNT), which was obtained by wrapping the MWNT with poly (diallydimethylammonium) chloride (PDDA), was used for the immobilization of glucose oxidase (GOD) and its bioelectrochemical studies. The morphologies and structures of the PDDA-MWNT composite were characterized by environment-canning electron microscopy (ESEM) and X-ray photoelectron spectroscopy (XPS). Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry were used to feature the GOD adsorbed onto the electrode modified by PDDA-MWNT composite. The immobilized GOD at the PDDA-MWNT films exhibited a pair of well-defined nearly reversible redox peaks and a fast heterogeneous electron transfer rate with the rate constant (k(s)) of 2.76 s(-1). In addition, GOD immobilized in this way retained its bioelectrocatalytic activity for the oxidation of glucose. The method of immobilizing GOD without any additional cross-linking agents presented here is easy and facile, which provides a model for other redox enzymes and proteins.
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The effect of Tb3+ ion on the Activity Index (RZ=A403/A275) of HRP was investigated by a combination of ultraviolet-visible, FT-infrared and X-ray photoelectron spectroscopic techniques. It was found that Tb3+ ion could bond to the O and/or N group of the amides in the polypeptide chains of HRP, leading to a decrease in the enzymatic activity index of HRP.
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Conversion of thyroxine (T-4) to 3,5,3'-triiodothyronine is an essential first step in controlling thyroid hormone action. Type I deiodinase (DI) can catalyze the conversion to produce the bulk of serum 3,5,3'-triiodothyronine. Acting as a mimic of DI, a selenium-containing catalytic antibody (Se-4C5) prepared by converting the serine residues of monoclonal antibody 4C5 raised against T4 into selenocysteines, can catalyze the deiodination of T4 with dithiothreitol (DTT) as cosubstrate. The mimic enzyme Se-4C5 exhibited a much greater deiodinase activity than model compound ebselen and another selenium-containing antibody Se-Hp4 against GSH. The coupling of selenocysteine with the combining pocket of antibody 4C5 endowed Se-4C5 with enzymatic activity. To probe the catalytic mechanism of the catalytic antibody, detailed kinetic studies were carried out in this paper. Investigations into the deiodinative reaction revealed the relationship between the initial velocity and substrate concentration. The characteristic parallel Dalziel plots demonstrated that Se-4C5-catalyzed reaction mechanism was ping-pong one, involving at least one covalent enzyme intermediate. The kinetic properties of the catalytic antibody were similar to those of DI, with K-m values for T-4 and DTT of approximately 0.8 muM and 1.8 muM, respectively, and a V-m value of 270 pmol per mg of protein per min. The activity could be sensitively inhibited by 6-propyl-2-thiouracil (PTU) with a K-i value of similar to 120 muM at 2.0 muM T-4 concentration. The PTU inhibition was progressively alleviated with the increasing concentration of added DTT, revealing that PTU was a competitive inhibitor for DTT.
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The thymidylate synthase (TS), an important target for many anticancer drugs, has been cloned from different species. But the cDNA property and function of TS in zebrafish are not well documented. In order to use zebrafish as an animal model for screening novel anticancer agents, we isolated TS cDNA from zebrafish and compared its sequence with those from other species. The open reading frame (ORF) of zebrafish TS cDNA sequence was 954 nucleotides, encoding a 318-amino acid protein with a calculated molecular mass of 36.15 kDa. The deduced amino acid sequence of zebrafish TS was similar to those from other organisms, including rat, mouse and humans. The zebrafish TS protein was expressed in Escherichia coli and purified to homogeneity. The purified zebrafish TS showed maximal activity at 28 degrees C with similar K-m value to human TS. Western immunoblot assay confirmed that TS was expressed in all the developmental stages of zebrafish with a high level of expression at the 1-4 cell stages. To study the function of TS in zebrafish embryo development, a short hairpin RNA (shRNA) expression vector, pSilencer 4.1-CMV/TS, was constructed which targeted the protein-coding region of zebrafish TS mRNA. Significant change in the development of tail and epiboly was found in zebrafish embryos microinjected pSilencer4.1-CMV/TS siRNA expression vector.
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The growth, mortality and digestive enzymes (trypsin, amylase and lipase) in miiuy croaker Miichthys miiuy larvae and juveniles (2-53 dph) were investigated at four photoperiods: 24L:OD), 18L:6D, 12L:12D and OL:24D. Larvae could not feed at OL:24D and did not survive up to 7 dph. In the 24L:OD, 18L:6D, 12L:12D groups, photoperiod had not significant effects on the growth of the rniiuy croaker younger than 20 dph. However, their total length and specific growth rate (SGR) were significantly larger at 18L:6D and 24L:OD than 12L:12D after 20 dph. Photoperiod also affected the mortality of the first feeding larvae (5 dph). being apparently higher in 5 dph larvae at OL:24D (60%) than at other photopenods (20-27%), but no significant differences in mortality were found among other photoperiods. High mortality of the miiuy croaker in 12L:12D, 18L:6D and 24 L:OD groups mainly occurred from 5 (20-27%) to 11 dph (11-16%) and tended to decrease gradually from 15 dph onwards. Digestive enzymes activities in the rniiuy croaker larvae and juveniles had a similar change trend with age at all photoperiods. They underwent drastic changes with age. The specific activity of lipase was significantly higher at 18L:6D and 24L:0D than 12L:12D, but there were no significant differences in specific activities of either trypsin or amylase among photoperiods. With regard to the total length, SGR, survival and digestive enzyme activities, our findings suggested that the optimal light regime for the culture of miiuy croaker during the early life stage was 18L:6D. (C) 2008 Elsevier B.V. All rights reserved.
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Oxalate decarboxylases (OXDCs) (E.C. 4.1.1.2) are enzymes catalyzing the conversion of oxalate to formate and CO2. The OXDCs found in fungi and bacteria belong to a functionally diverse protein superfamily known as the cupins. Fungi-originated OXDCs are secretory enzymes. However, most bacterial OXDCs are localized in the cytosol, and may be involved in energy metabolism. In Agrobacterium tumefaciens C58, a locus for a putative oxalate decarboxylase is present. In the study reported here, an enzyme was overexpressed in Escherichia coli and showed oxalate decarboxylase activity. Computational analysis revealed the A. tumefaciens C58 OXDC contains a signal peptide mediating translocation of the enzyme into the periplasm that was supported by expression of signal-peptideless and full-length versions of the enzyme in A. tumefaciens C58. Further site-directed mutagenesis experiment demonstrated that the A. tumefaciens C58 OXDC is most likely translocated by a twin-arginine translocation (TAT) system.