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1.
Extracts of the adult worms of both Schistosoma mansoni and Schistosoma haematobium can metabolise some typical P450 substrates but to differing degrees. S. mansoni worm extracts displayed a approximately 12-fold higher specific activity for an aminopyrine substrate than rat liver microsomes. At 4 mM substrate concentration the demethylation reaction with N-nitrosodimethylamine (NDMA) (5 nmol HCHO/mg protein/min) was only half that of rat liver microsomes, whereas in extracts of S. haematobium, no detectable activity was found towards NDMA. Using ethylmorphine as substrate the demethylation activity of S. mansoni extracts (1.82 nmol HCHO/mg protein/min) was 5.5-fold lower than that of rat liver microsomes. Benzphetamine demethylase activity was also readily detectable in S. mansoni worm extracts at 6.79 nmol HCHO/mg protein/min compared with 10.20 nmol HCHO/mg protein/min in the case of rat liver microsomes. When aniline was used as substrate, surprisingly, no activity was found in worm extracts of either S. mansoni or S. haematobium, whereas rat liver microsomes showed high activity towards this amine. The anti-P450 2E1 and 2B1/2 cross-reacted with both worm homogenates and gave a specific band corresponding to a protein of molecular weight of approximately 50.0 kDa. A study with anti-P450 IVA antibody revealed that while this protein was strongly expressed in S. haematobium worm extracts, no immunoreactivity was observed with extracts of S. mansoni. Immunoblotting analyses with anti-P450 IIIA and P450 1A1 did not detect immunoreactive protein in either S. mansoni or S. haematobium. 相似文献
2.
Rubrerythrin from the hyperthermophilic archaeon Pyrococcus furiosus is a rubredoxin-dependent, iron-containing peroxidase
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Rubrerythrin was purified by multistep chromatography under anaerobic, reducing conditions from the hyperthermophilic archaeon Pyrococcus furiosus. It is a homodimer with a molecular mass of 39.2 kDa and contains 2.9 +/- 0.2 iron atoms per subunit. The purified protein had peroxidase activity at 85 degrees C using hydrogen peroxide with reduced P. furiosus rubredoxin as the electron donor. The specific activity was 36 micromol of rubredoxin oxidized/min/mg with apparent K(m) values of 35 and 70 microM for hydrogen peroxide and rubredoxin, respectively. When rubrerythrin was combined with rubredoxin and P. furiosus NADH:rubredoxin oxidoreductase, the complete system used NADH as the electron donor to reduce hydrogen peroxide with a specific activity of 7.0 micromol of H(2)O(2) reduced/min/mg of rubrerythrin at 85 degrees C. Strangely, as-purified (reduced) rubrerythrin precipitated when oxidized by either hydrogen peroxide, air, or ferricyanide. The gene (PF1283) encoding rubrerythrin was expressed in Escherichia coli grown in medium with various metal contents. The purified recombinant proteins each contained approximately three metal atoms/subunit, ranging from 0.4 Fe plus 2.2 Zn to 1.9 Fe plus 1.2 Zn, where the metal content of the protein depended on the metal content of the E. coli growth medium. The peroxidase activities of the recombinant forms were proportional to the iron content. P. furiosus rubrerythrin is the first to be characterized from a hyperthermophile or from an archaeon, and the results are the first demonstration that this protein functions in an NADH-dependent, hydrogen peroxide:rubredoxin oxidoreductase system. Rubrerythrin is proposed to play a role in the recently defined anaerobic detoxification pathway for reactive oxygen species. 相似文献
3.
Purification and properties of a glutathione peroxidase from Southern bluefin tuna (Thunnus maccoyii) liver 总被引:1,自引:0,他引:1
Thompson JL Thomas PM Schuller KA 《Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology》2006,144(1):86-93
A glutathione peroxidase (GPX) protein was purified approximately 1000-fold from Southern bluefin tuna (Thunnus maccoyii) liver to a final specific activity of 256 micromol NADPH oxidised min(-1) mg(-1) protein. Gel filtration chromatography and denaturing protein gel electrophoresis of the purified preparation indicated that the protein has a native molecular mass of 85 kDa and is most likely a homotetramer with subunits of approximately 24 kDa. The Km values of the purified enzyme for hydrogen peroxide, cumene hydroperoxide, t-butyl hydroperoxide and glutathione were 12, 90, 90 and 5900 microM, respectively. The Km values for cumene hydroperoxide and t-butyl hydroperoxide were approximately 8-fold greater than the Km value for hydrogen peroxide. Thus, the SBT liver GPX has a considerably greater affinity for hydrogen peroxide than for the other two substrates. The pH optimum of the purified enzyme was pH 8.0. Immunoblotting experiments with polyclonal antibodies, raised against a recombinant human GPX, provided further evidence that the purified SBT enzyme is a genuine GPX. 相似文献
4.
A gene (APE2278) encoding the peroxiredoxin (Prx) homologous protein of yeast and human was identified in the genome data base of the aerobic hyperthermophilic archaeon Aeropyrum pernix. We cloned the gene and produced the encoded protein in Escherichia coli cells. The isolated recombinant protein showed peroxidase activity in vitro and used the thioredoxin system of A. pernix as an electron donor. These results indicate that the recombinant protein is in fact thioredoxin peroxidase (ApTPx) of A. pernix. Immunoblot analysis revealed that the expression of ApTPx was induced as a cellular adaptation in response to the addition of exogenous H2O2 and may exert an antioxidant activity in vivo. An analysis of the ApTPx oligomers by high pressure liquid chromatography and electron microscopic studies showed that ApTPx exhibited the hexadecameric protein forming 2-fold toroid-shaped structure with outer and inner diameters of 14 and 6 nm, respectively. These results indicated that ApTPx is a novel hexadecameric protein composed of two identical octamers. Although oligomerization of individual subunits does not take place through an intersubunit-disulfide linkage involving Cys50 and Cys213, Cys50 is essential for the formation of the hexadecamer. Mutagenesis studies suggest that the sulfhydryl group of Cys50 is the site of oxidation by peroxide and that oxidized Cys50 reacts with the sulfhydryl group of Cys213 of another subunit to form an intermolecular disulfide bond. The resulting disulfide can then be reduced by thioredoxin. In support of this hypothesis, ApTPx mutants lacking either Cys50 or Cys213 showed no TPx activity, whereas the mutant lacking Cys207 had a TPx activity. This is the first report on the biochemical and structural features of a novel hexadecameric thioredoxin peroxidase from the archaea. 相似文献
5.
日本血吸虫新抗原基因Sj—Ts4的克隆、表达及免疫保护性 总被引:4,自引:0,他引:4
为探讨旋毛虫感染小鼠后抗血吸虫感染的分子机制 ,并为血吸虫病疫苗的研究提供新的抗原分子 ,用旋毛虫感染鼠血清筛选日本血吸虫成虫cDNA文库。经过 3轮筛选 ,获得 9个阳性克隆 ,其中新基因Sj Ts4有一完整的编码框 ,编码2 10个氨基酸。该蛋白质的理论分子量为 2 3kD ,等电点为 7.72。将Sj Ts4亚克隆于原核表达载体 pGEX 5X 3,以GST融合蛋白的形式在E .coliDH5α中表达。Western印迹分析显示 ,重组Sj Ts4蛋白 (rSj Ts4)能被慢性感染鼠血清识别。rSj Ts4或与弗氏完全佐剂混合后免疫小鼠 ,均可以诱导产生特异性的IgG抗体 ,抗体效价可高达 1∶2 5 6 0 0。两免疫组的减虫率分别为31.36 %和 36 .80 % ,与对照组相比都具有统计学意义 相似文献
6.
E M Cornford A M Fitzpatrick T L Quirk C P Diep E M Landaw 《The Journal of parasitology》1988,74(1):116-128
Tegumental hexose transporters have been kinetically characterized in mated and separated male and female Schistosoma mansoni 8-12 wk postinfection. Significant gender-specific differences in Km and Vmax were observed. In mated males, the estimated constants (mean +/- SE) were: Km = 0.63 +/- 0.31 mM, Vmax = 0.93 +/- 0.44 nmol/mg worm water/min, and the Kd = 0.25 +/- 0.09 microliter/mg worm water/min. In mated females the kinetics were: Km = 0.99 +/- 0.40 mM, Vmax = 1.22 +/- 0.42 nmol/mg worm water/min, and Kd = 0.60 +/- 0.14 microliter/mg worm water/min. The influx of 2-deoxy-D-glucose and 3-O-methylglucose has been similarly characterized; these analogs share the same glucose transporter in male and female schistosomes. 2-Deoxy-D-glucose has a higher affinity, and 3-O-methylglucose a lower affinity, than does glucose. Because mated male schistosomes supply glucose to female partners, similarities between the free glucose concentration of the male and the affinity of the transporter determined for mated female schistosomes suggest that male-to-female transfer may be a potentially rate-limiting step in glucose utilization by the female. Permeability x surface are (PS) products and Vmax/Km ratios were significantly elevated in mated schistosomes, suggesting that the transporter is primarily localized to the dorsal surface of the male. Gender- and mating-specific analyses of PS products indicate that tegumental permeability to glucose is significantly increased in mated schistosomes, and compares very favorably to that of the host liver. 相似文献
7.
Dmitri M. Hushpulian Andrew A. Poloznikov Pavel A. Savitski Alexandra M. Rozhkova Tatyana A. Chubar Victoria A. Fechina L. Mark Lagrimini Vladimir I. Tishkov Irina G. Gazaryan 《Biocatalysis and Biotransformation》2007,25(2):163-170
The wild-type anionic tobacco peroxidase and its Glu141Phe mutant have been expressed in Escherichia coli, and reactivated to yield active enzymes. A Glu141Phe substitution was made with the tobacco anionic peroxidase (TOP) to mimic neutral plant peroxidases, such as horseradish peroxidase (HRP). Both recombinant forms of tobacco peroxidase show extremely high activity in luminol oxidation with hydrogen peroxide, and thus, preserve the unique property of the native tobacco peroxidase, a superior chemiluminescent reagent. The chemiluminescent signal intensity for both recombinant forms of TOP is orders of magnitude higher than that for wild-type recombinant HRP. The substitution slightly increases TOP activity and stability in the reaction course, but has almost no effect on the optimal parameters of the reaction (pH, luminol and hydrogen peroxide concentrations) and calibration plot. Comparison of substrate specificity profiles for recombinant TOP and HRP demonstrates that Glu141 has no principal effect on the enzyme activity. It is not the presence of the negative charge at the haem edge, but the high redox potential of TOP Compounds I and II that provides high activity towards aromatic amines and aminophenols, and luminol in particular. 相似文献
8.
Recently, novel hybrid thiol peroxidase (TPx) proteins fused with a glutaredoxin (Grx) were found from some pathogenic bacteria, cyanobacteria, and anaerobic sulfur-oxidizing phototroph. The phylogenic tree analysis that was constructed from the aligned sequences showed two major branches. Haemophilus influenzae TPx.Grx was grouped in one branch as a 1-Cys subfamily of the thiol-specific antioxident protein/AhpC family. Most TPx.Grx proteins, including Vibrio cholerae TPx.Grx, were grouped in the 2-Cys subfamily. To explain the existence of two subgroups in novel hybrid TPx proteins, we have compared the kinetics given by V. cholerae TPx.Grx, H. influenzae TPx.Grx, their separated TPx domains, and a set of mutants devoid of the redox-active cysteines. The kinetic study described here demonstrates clearly that V. cholerae TPx.Grx is a 2-Cys TPx subfamily. For the first time, we also demonstrate the lipid peroxidase activity of V. cholerae TPx.Grx fusion and suggest the in vivo function of 2-Cys TPx.Grx fusion serving as a lipid peroxidase. 相似文献
9.
Peroxiredoxins are antioxidant enzymes whose peroxidase activity depends on a redox-sensitive cysteine residue at the active center. In this study we investigated properties of the active center cysteine of bovine 1-Cys peroxiredoxin using a recombinant protein (BRPrx). The only cysteine residue of the BRPrx molecule was oxidized rapidly by an equimolar peroxide or peroxynitrite to the cysteine sulfenic acid. Approximate rates of oxidation of BRPrx by different peroxides were estimated using selenium glutathione peroxidase as a competitor. Oxidation of the active center cysteine of BRPrx by H2O2 proceeded only several times slowly than that of the selenocysteine of glutathione peroxidase. The rate of oxidation varied depending on peroxides tested, with H2O2 being about 7 and 80 times faster than tert-butyl hydroperoxide and cumene hydroperoxide, respectively. Peroxynitrite oxidized BRPrx slower than H2O2 but faster than tert-butyl hydroperoxide. Further oxidation of the cysteine sulfenic acid of BRPrx to higher oxidation states proceeded slowly. Oxidized BRPrx was reduced by dithiothreitol, dihydrolipoic acid, and hydrogen sulfide, and demonstrated peroxidase activity (about 30 nmol/mg/min) with these reductants as electron donors. beta-Mercaptoethanol formed a mixed disulfide and did not support peroxidase activity. Oxidized BRPrx did not react with glutathione, cysteine, homocysteine, N-acetyl-cysteine, and mercaptosuccinic acid. 相似文献
10.
To investigate the antioxidative capacities of oligodendrocytes, rat brain cultures enriched for oligodendroglial cells were prepared and characterized. These cultures contained predominantly oligodendroglial cells as determined by immunocytochemical staining for the markers galactocerebroside and myelin basic protein. If oligodendroglial cultures were exposed to exogenous hydrogen peroxide (100 micro m), the peroxide disappeared from the incubation medium following first order kinetics with a half-time of approximately 18 min. Normalization of the disposal rate to the protein content of the cultures by calculation of the specific hydrogen peroxide detoxification rate constant revealed that the cells in oligodendroglial cultures have a 60% to 120% higher specific capacity to dispose of hydrogen peroxide than cultures enriched for astroglial cells, microglial cells or neurones. Oligodendroglial cultures contained specific activities of 133.5 +/- 30.4 nmol x min(-1) x mg protein(-1) and 27.5 +/- 5.4 nmol x min(-1) x mg protein(-1) of glutathione peroxidase and glutathione reductase, respectively. The specific rate constant of catalase in these cultures was 1.61 +/- 0.54 min(-1) x mg protein(-1). Comparison with data obtained by identical methods for cultures of astroglial cells, microglial cells and neurones revealed that all three of the enzymes which are involved in hydrogen peroxide disposal were present in oligodendroglial cultures in the highest specific activities. These results demonstrate that oligodendroglial cells in culture have a prominent machinery for the disposal of hydrogen peroxide, which is likely to support the protection of these cells in brain against peroxides when produced by these or by surrounding brain cells. 相似文献
11.
Members of the PAR-1/MARK serine/threonine protein kinase (STK) subfamily are important regulators of the cytoskeleton, and their characterization can provide insights into a number of critical processes relating to the development and survival of an organism. We previously investigated the mRNA expression for and organization of a gene (hcstk) representing HcSTK, an STK from the parasitic nematode Haemonchus contortus. In the present study, a recombinant form of HcSTK was expressed and characterized. Affinity-purified anti-HcSTK antibodies reacted with native HcSTK in protein homogenates extracted from third-stage larvae (L3) of H. contortus and were also used to immunolocalize the protein around the nuclei of ovarian and intestinal tissues of adult H. contortus. The enzyme activity of the recombinant HcSTK protein was also demonstrated. The findings show that recombinant HcSTK is a functional protein kinase, with activity directed to KXGS motifs, consistent with other members of the PAR-1/MARK STK subfamily. 相似文献
12.
J Rodríguez-Pérez J R Rodríguez-Medina M A García-Blanco G V Hillyer 《Experimental parasitology》1992,74(4):400-407
Immunization of mice with an antigenic polypeptide from Fasciola hepatica adult worms and having an apparent molecular mass of 12,000 Da (Fh12) has been shown to reduce the worm burden from challenge infection with Schistosoma mansoni by more than 50%. Moreover, mice infected with S. mansoni develop antibodies to Fh12 after 5-6 weeks of infection, indicating that this Fasciola-derived antigen is a cross-reactive, cross-protective protein. A lambda gt11 F. hepatica cDNA library was constructed from poly(A)+ RNA extracted from adult worms. A cDNA encoding a cross-reactive polypeptide (Fh15) was cloned by screening the F. hepatica lambda gt11 library with a monospecific, polyclonal rabbit antiserum against pure, native Fh12. The cDNA was sequenced and the predicted amino acid sequence revealed an open reading frame encoding a 132-amino-acid protein with a predicted molecular weight of 14,700 Da. This protein has significant homology to a 14-kDa S. mansoni fatty acid-binding protein. Comparison of the protective-inducing activity of recombinant Fh15 with that of purified Fh12 against schistosomes and Fasciola is warranted. 相似文献
13.
Timofeevski SL Nie G Reading NS Aust SD 《Biochemical and biophysical research communications》1999,256(3):500-504
Manganese peroxidase and lignin peroxidase are ligninolytic heme-containing enzymes secreted by the white-rot fungus Phanerochaete chrysosporium. Despite structural similarity, these peroxidases oxidize different substrates. Veratryl alcohol is a typical substrate for lignin peroxidase, while manganese peroxidase oxidizes chelated Mn2+. By a single mutation, S168W, we have added veratryl alcohol oxidase activity to recombinant manganese peroxidase expressed in Escherichia coli. The kcat for veratryl alcohol oxidation was 11 s-1, Km for veratryl alcohol approximately 0.49 mM, and Km for hydrogen peroxide approximately 25 microM at pH 2.3. The Km for veratryl alcohol was higher and Km for hydrogen peroxide was lower for this manganese peroxidase mutant compared to two recombinant lignin peroxidase isoenzymes. The mutant retained full manganese peroxidase activity and the kcat was approximately 2.6 x 10(2) s-1 at pH 4.3. Consistent with relative activities with respect to these substrates, Mn2+ strongly inhibited veratryl alcohol oxidation. The single productive mutation in manganese peroxidase suggested that this surface tryptophan residue (W171) in lignin peroxidase is involved in catalysis. 相似文献
14.
Missirlis F Rahlfs S Dimopoulos N Bauer H Becker K Hilliker A Phillips JP Jäckle H 《Biological chemistry》2003,384(3):463-472
Cellular defense systems against reactive oxygen species (ROS) include thioredoxin reductase (TrxR) and glutathione reductase (GR). They generate sulfhydryl-reducing systems which are coupled to antioxidant enzymes, the thioredoxin and glutathione peroxidases (TPx and GPx). The fruit fly Drosophila lacks a functional GR, suggesting that the thioredoxin system is the major source for recycling glutathione. Whole genome in silico analysis identified two non-selenium containing putative GPx genes. We examined the biochemical characteristics of one of these gene products and found that it lacks GPx activity and functions as a TPx. Transgene-dependent overexpression of the newly identified Glutathione peroxidase homolog with thioredoxin peroxidase activity (Gtpx-1) gene increases resistance to experimentally induced oxidative stress, but does not compensate for the loss of catalase, an enzyme which, like GTPx-1, functions to eliminate hydrogen peroxide. The results suggest that GTPx-1 is part of the Drosophila Trx antioxidant defense system but acts in a genetically distinct pathway or in a different cellular compartment than catalase. 相似文献
15.
Hushpulian DM Savitski PA Rojkova AM Chubar TA Fechina VA Sakharov IY Lagrimini LM Tishkov VI Gazaryan IG 《Biochemistry. Biokhimii?a》2003,68(11):1189-1194
Coding DNA of the tobacco anionic peroxidase gene was cloned in pET40b vector. The problem of 11 arginine codons, rare in procaryotes, in the tobacco peroxidase gene was solved using E. coli BL21(DE3) Codon Plus strain. The expression level of the tobacco apo-peroxidase in the above strain was approximately 40% of the total E. coli protein. The tobacco peroxidase refolding was optimized based on the earlier developed protocol for horseradish peroxidase. The reactivation yield of recombinant tobacco enzyme was about 7% with the specific activity of 1100-1200 U/mg towards 2,2;-azino-bis(3-ethylbenzothiazoline-6-sulfonate) (ABTS). It was shown that the reaction of ABTS oxidation by hydrogen peroxide catalyzed by recombinant tobacco peroxidase proceeds via the ping-pong kinetic mechanism as for the native enzyme. In the presence of calcium ions, the recombinant peroxidase exhibits a 2.5-fold decrease in the second order rate constant for hydrogen peroxide and 1.5-fold decrease for ABTS. Thus, calcium ions have an inhibitory effect on the recombinant enzyme like that observed earlier for the native tobacco peroxidase. The data demonstrate that the oligosaccharide part of the enzyme has no effect on the kinetic properties and calcium inhibition of tobacco peroxidase. 相似文献
16.
Dmitri M. Hushpulian Andrew A. Poloznikov Pavel A. Savitski Alexandra M. Rozhkova Tatyana A. Chubar Victoria A. Fechina 《Biocatalysis and Biotransformation》2013,31(2-4):163-170
The wild-type anionic tobacco peroxidase and its Glu141Phe mutant have been expressed in Escherichia coli, and reactivated to yield active enzymes. A Glu141Phe substitution was made with the tobacco anionic peroxidase (TOP) to mimic neutral plant peroxidases, such as horseradish peroxidase (HRP). Both recombinant forms of tobacco peroxidase show extremely high activity in luminol oxidation with hydrogen peroxide, and thus, preserve the unique property of the native tobacco peroxidase, a superior chemiluminescent reagent. The chemiluminescent signal intensity for both recombinant forms of TOP is orders of magnitude higher than that for wild-type recombinant HRP. The substitution slightly increases TOP activity and stability in the reaction course, but has almost no effect on the optimal parameters of the reaction (pH, luminol and hydrogen peroxide concentrations) and calibration plot. Comparison of substrate specificity profiles for recombinant TOP and HRP demonstrates that Glu141 has no principal effect on the enzyme activity. It is not the presence of the negative charge at the haem edge, but the high redox potential of TOP Compounds I and II that provides high activity towards aromatic amines and aminophenols, and luminol in particular. 相似文献
17.
棉铃虫组织蛋白酶B多克隆抗体制备及鉴定 总被引:3,自引:0,他引:3
介绍了用重组的棉铃虫组织蛋白酶B(HCB)制备兔多克隆抗体的方法。用已经克隆得到的pGEX 4T 1 HCB重组质粒转化的大肠杆菌 (Escherichiacoli)BL2 1菌株进行诱导表达 ,获得融合表达的包涵体产物。经过变性、复性等方法处理包涵体 ,获得可溶性融合蛋白。用凝血酶裂解融合蛋白 ,再经SDS -聚丙烯酰胺凝胶电泳分离纯化目的蛋白HCB ,用做抗原免疫家兔。产生的抗血清经硫酸铵沉淀 ,最后得到了抗棉铃虫组织蛋白酶B的多克隆抗体。用间接酶联免疫吸附测定法测得该抗体对重组表达的组织蛋白酶B和棉铃虫体内的组织蛋白酶B的效价分别为 1∶5 1 2 0 0和 1∶2 5 60 0。通过免疫印迹检测证明此抗体不仅可以识别重组表达的棉铃虫组织蛋白B ,而且可以识别棉铃虫卵巢匀浆液中的棉铃虫组织蛋白酶B。 相似文献
18.
Fasciola hepatica: host responders and nonresponders to parasite glutathione S-transferase. 总被引:1,自引:0,他引:1
Fasciola hepatica glutathione S-transferase (FhGST) was isolated from adult worms by glutathione agarose affinity chromatography. SDS-PAGE shows three proteins of M(r) ranging from 29-27.8 kDa. Western immunoblot analyses using SDS-PAGE separated adult worm extracts and probed with a rabbit anti-FhGST antiserum reveal two bands in the same M(r) range. Mice and rabbits immunized with purified FhGST develop copious amounts of anti-FhGST antibodies. Moreover, antisera to F. hepatica adult worms and excretion-secretion products also react with FhGST. Cross-reactivity with schistosomes is evidenced in the reactivity with FhGST of anti-Schistosoma mansoni adult worm antisera and, to a lesser extent, antisera to S. mansoni-soluble egg antigens. The time of appearance of anti-FhGST antibodies in different species of animals infected with F. hepatica was determined. Sheep and a New Zealand white rabbit developed anti-FhGST antibodies detectable by ELISA as early as 2 weeks postexposure with F. hepatica. However, neither mice nor calves infected with F. hepatica developed antibodies to FhGST through the 5-10 weeks of infection tested. But mice infected with S. mansoni developed anti-FhGST cross-reacting antibodies by 6 weeks of infection. Calves immunized with a Fasciola/Schistosoma cross-reactive, cross-protective antigen complex in which a 12,000-kDa protein (Fh12) has been shown to contain immunoprophylactic activity, also developed antibodies to FhGST. Since FhGST is a novel potential vaccine, its protection-inducing capability in a multivalent vaccine combined with Fh12 clearly warrants study. In summary, it appears that hosts with fascioliasis are either responders to FhGST (rabbits, sheep) or nonresponders (mice, cattle), offering interesting models for studying the immune response. 相似文献
19.
Activated leukocytes participate in immunity to infection by the parasitic blood fluke Schistosoma mansoni. They attach to the surface of schistosomes and secrete schistosomicidal substances. Cationic proteins, hydrolytic enzymes, and oxidants, produced by the leukocytes, have been implicated in the damage to the schistosomes. To examine the possible involvement of elastase in the killing of schistosomes by leukocytes, young and adult stages of S. mansoni were treated in vitro with pancreatic elastase (PE) and neutrophil elastase (NE). Schistosomula, lung-stage schistosomula (LSS), and adult worms (AW) have been found to be sensitive to both PE and NE. Male AW were more sensitive to PE than female AW. The enzymatic activity of elastase is essential for its toxic effect because heat-inactivation and specific elastase inhibitors prevented elastase-mediated schistosome killing. Thus, alpha1-antitrypsin and the chloromethyl ketone (CMK)-derived tetrapeptides Ala-Ala-Pro-Val-CMK and Ala-Ala-Pro-Ala-CMK but not Ala-Ala-Pro-Phe-CMK and Ala-Ala-Pro-Leu-CMK blocked PE caseinolytic and schistosomulicidal activities. As shown previously, schistosomes are also efficiently killed by hydrogen peroxide. LSS appear to be more resistant than AW and early-stage schistosomula to the lytic effects of hydrogen peroxide. Cotreatment experiments with both elastase and hydrogen peroxide indicated that they exert an additive toxic effect and that hydrogen peroxide sensitizes schistosomula to the toxic effect of elastase but not vice versa. These results demonstrate, for the first time, that elastases may be toxic molecules used by neutrophils, eosinophils, and macrophages to kill various developmental stages of S. mansoni. 相似文献
20.
Stéphane Herbette Catherine Lenne Nathalie Leblanc Jean-Louis Julien Jo?l R Drevet Patricia Roeckel-Drevet 《European journal of biochemistry》2002,269(9):2414-2420
This study investigated the enzymatic function of two putative plant GPXs, GPXle1 from Lycopersicon esculentum and GPXha2 from Helianthus annuus, which show sequence identities with the mammalian phospholipid hydroperoxide glutathione peroxidase (PHGPX). Both purified recombinant proteins expressed in Escherichia coli show PHGPX activity by reducing alkyl, fatty acid and phospholipid hydroperoxides but not hydrogen peroxide in the presence of glutathione. Interestingly, both recombinant GPXle1 and GPXha2 proteins also reduce alkyl, fatty acid and phospholipid hydroperoxides as well as hydrogen peroxide using thioredoxin as reducing substrate. Moreover, thioredoxin peroxidase (TPX) activities were found to be higher than PHGPX activities in terms of efficiency and substrate affinities, as revealed by their respective Vmax and Km values. We therefore conclude that these two plant GPX-like proteins are antioxidant enzymes showing PHGPX and TPX activities. 相似文献