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Enzymatic Pathways (enzymatic + pathway)
Selected AbstractsInterferon-stimulated gene 15 (ISG15) conjugates proteins in dermatomyositis muscle with perifascicular atrophyANNALS OF NEUROLOGY, Issue 1 2010Mohammad Salajegheh MD Objective We investigated interferon-stimulated gene 15 (ISG15), a poorly understood ubiquitin-like modifier, and its enzymatic pathway in dermatomyositis (DM), an autoimmune disease primarily involving muscle and skin. Methods We generated microarray data measuring transcript abundance for approximately 18,000 genes in each of 113 human muscle biopsy specimens, and studied biopsy specimens and cultured skeletal muscle using immunohistochemistry, immunoblotting proteomics, real-time quantitative polymerase chain reaction, and laser-capture microdissection. Results Transcripts encoding ISG15-conjugation pathway proteins were markedly upregulated in DM with perifascicular atrophy (DM-PFA) muscle (ISG15 339-fold, HERC5 62-fold, and USP18 68-fold) compared with 99 non-DM samples. Combined analysis with publicly available microarray datasets showed that >50-fold ISG15 transcript elevation had 100% sensitivity and specificity for 28 biopsies from adult DM-PFA and juvenile DM patients compared with 199 muscle samples from other muscle diseases. Free ISG15 and ISG15-conjugated proteins were only found on immunoblots from DM-PFA muscle. Cultured human skeletal muscle exposed to type 1 interferons produced similar transcripts and ISG15 protein and conjugates. Laser-capture microdissection followed by proteomic analysis showed deficiency of titin in DM perifascicular atrophic myofibers. Interpretation A large-scale microarray study of muscle samples demonstrated that among a diverse group of muscle diseases DM was uniquely associated with upregulation of the ISG15 conjugation pathway. Exposure of human skeletal muscle cell culture to type 1 interferons produced a molecular picture highly similar to that seen in human DM muscle. Perifascicular atrophic myofibers in DM were deficient in a number of skeletal muscle proteins including titin. ANN NEUROL 2010;67:53,63 [source] Nitric oxide generation from hydroxylamine in the presence of neutrophils and in the cell-free systemAPMIS, Issue 7-8 2001Magdalena Klink Conversion of hydroxylamine (HA) to nitric oxide (NO) has been studied in the presence or absence of human neutrophils with or without myristate acetate phorbol (PMA), catalase (CAT), hydrogen peroxide (H2O2), and superoxide dismutase (SOD) and nitric oxide synthase (NOS) inhibitors. The generation of NO from HA in the presence of neutrophils was higher than in the cell-free system. We found that catalase did not influence the nitrite generation from HA in the cell-free system and in the presence of neutrophils. The H2O2 enhanced the NO generation from HA in the presence of neutrophils only. When catalase and H2O2 were added together, a high increase of NO generation from HA in both systems was observed. The addition of SOD decreased whereas addition of PMA enhanced the NO generation from HA in the presence of neutrophils. The presented data show the possible role of oxygen radicals in the decomposition of HA to NO. The addition of NOS inhibitors to the culture of neutrophils decreased the generation of nitrite from HA. Our results suggest that NO generation from HA, which is an intermediate in NO production from L-arginine, may be supported by an enzymatic pathway in which cellular NO synthase is involved. [source] Involvement of 15-lipoxygenase and prostaglandin EP receptors in aspirin-triggered 15-hydroxyeicosatetraenoic acid generation in aspirin-sensitive asthmaticsCLINICAL & EXPERIMENTAL ALLERGY, Issue 7 2008M. Jedrzejczak-Czechowicz Summary Background The mechanism of aspirin (acetylsalicylic acid: ASA) hypersensitivity in asthmatic patients is related to arachidonic acid metabolism abnormalities, and specific triggering by ASA of 15-hydroxyeicosatetraenoic acid (15-HETE) generation was observed in leucocytes from aspirin-sensitive (AS) but not from aspirin-tolerant (AT) asthmatics. Objective The aim of this study was to identify the enzymatic pathway involved in ASA-induced 15-HETE generation in AS asthmatics and to assess the regulatory role of prostaglandin EP receptors. Methods Peripheral blood leucocytes (PBLs) were isolated from AS (n=18) and AT (n=20) asthmatics and challenged with ASA, with and without pre-incubation with caffeic acid (CA) [15-lipoxygenase (15-LO) inhibitor] or prostaglandin receptor non-specific (misoprostol, sulprostone) and specific EP1,4 receptors agonists. Eicosanoids were measured in supernatants using specific immunoassays. Results Aspirin triggered 15-HETE generation in PBLs of AS asthmatics (mean increase 292%) but not in AT asthmatics and inhibited prostaglandin2 (PGE2) generation in both groups of patients to the same degree. Leucocytes from AS patients produced less PGE2, both before and after ASA incubation. Pre-incubation of PBLs with CA decreased basal 15-HETE production in all patients and completely inhibited ASA-induced 15-HETE generation in AS asthmatics. CA did not change basal PGE2 production but enhanced induced by ASA inhibition of PGE2. Non-specific agonists of EP receptors (misoprostol and sulprostone) did not affect basal 15-HETE production but inhibited in a dose-dependent manner the ASA-induced increase of 15-HETE generation in AS asthmatics. On the contrary, in AT asthmatics, pre-incubation of PBLs with misoprostol or sulprostone resulted in a significant increase in 15-HETE generation after addition of ASA (200 ,m). EP1,3 receptor agonists inhibited (range 72,94%) the ASA-induced 15-HETE production significantly. Conclusion Our study demonstrated that ASA-triggered 15-HETE generation involves the activation of 15-LO and is modulated by prostaglandin EP1,3 receptors. The relevance of these observations to the mechanism of in vivo ASA-induced asthmatic attack remains to be established. [source] Diabetic neuropathy and oxidative stressDIABETES/METABOLISM: RESEARCH AND REVIEWS, Issue 4 2006Rodica Pop-Busui Abstract This review will focus on the impact of hyperglycemia-induced oxidative stress in the development of diabetes-related neural dysfunction. Oxidative stress occurs when the balance between the production of reactive oxygen species (ROS) and the ability of cells or tissues to detoxify the free radicals produced during metabolic activity is tilted in the favor of the former. Although hyperglycemia plays a key role in inducing oxidative stress in the diabetic nerve, the contribution of other factors, such as endoneurial hypoxia, transition metal imbalances, and hyperlipidemia have been also suggested. The possible sources for the overproduction of ROS in diabetes are widespread and include enzymatic pathways, auto-oxidation of glucose, and mitochondrial superoxide production. Increase in oxidative stress has clearly been shown to contribute to the pathology of neural and vascular dysfunction in diabetes. Potential therapies for preventing increased oxidative stress in diabetic nerve dysfunction will be discussed. Copyright © 2006 John Wiley & Sons, Ltd. [source] Bioaccumulation and biotransformation of arsenic in the Mediterranean polychaete Sabella spallanzanii experimental observationsENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 6 2007Alessandra Notti Abstract The Mediterranean fan worm Sabella spallanzanii is characterized by elevated basal levels of arsenic in branchial crowns (>1,000 ,g/g) and an unusual prevalence of dimethylarsinic acid (DMA), a relatively toxic compound with a possible antipredatory role. The aim of this work was to obtain further insights on the capability of this polychaete to accumulate arsenic from different compounds and to operate biotransformation reactions. Laboratory exposures to arsenate (AsV), dimethylarsinic acid (DMA), trimethylarsine (TMA), and arsenobetaine (AsB) revealed significant differences among tissues and kind of experiments. The highest increases of arsenic content were observed in branchial crowns of organisms treated with arsenate, which can enter the cell through the phosphate carrier system; lower variations were measured with DMA and TMA, while not-significant changes of total As occurred after treatments with AsB. In body tissues, exposure to AsV, DMA, and TMA confirmed a progressively lower accumulation of total arsenic, while a marked increase was caused by AsB. Obtained results suggested that accumulated arsenic could be chemically transformed, thus explaining the elevated basal levels of DMA typical of S. spallanzanii; during all the experiments, DMA was the most accumulated molecule, suggesting that this species possesses the enzymatic pathways for methylation and demethylation reactions of inorganic and trimethylated arsenicals. Only arsenobetaine was not converted into DMA, which would confirm a microbial pathway for degradation for this molecule, particularly important in body tissues of S. spallanzanii for the presence of bacteria associated to digestive tracts. Overall, the present study suggests future investigations on the biological role of arsenic and DMA in S. spallanzanii as a potential adaptive mechanism against predation in more vulnerable tissues. [source] The role of cysteine and cysteine,S conjugates as odour precursors in the flavour and fragrance industry ,FLAVOUR AND FRAGRANCE JOURNAL, Issue 6 2008Christian Starkenmann Abstract Volatile sulphur compounds are important odorants in the flavour and fragrance industries. Recent developments in the field have reinvigorated sulphur compound research, particularly with regard to the precursor compounds of volatile thiols. The present review concentrates on the role of cysteine and cysteine,S conjugates as precursor compounds for a variety of aromas and examines the chemical and enzymatic pathways of degradation and biotransformation. Cysteine is an important source of sulphur in flavour chemistry and may be degraded into hydrogen sulphide, which can further participate in other reactions. Direct reactions of cysteine and the pathways particular to various flavours and fragrances are discussed, with an emphasis on the precursor compounds cysteine,S conjugates. The importance of cysteine precursors in plants, including onion, wine, passion fruit, bitter orange, asparagus and bell pepper and the formation of natural scents in cats and humans are discussed. Copyright © 2008 John Wiley & Sons, Ltd. [source] Genes involved in the determination of the rate of inversions at short inverted repeatsGENES TO CELLS, Issue 6 2000Malgorzata M. Slupska Background Not all of the enzymatic pathways involved in genetic rearrangements have been elucidated. While some rearrangements occur by recombination at areas of high homology, others are mediated by short, often interrupted homologies. We have previously constructed an Escherichia coli strain that allows us to examine inversions at microhomologies, and have shown that inversions can occur at short inverted repeats in a recB,C -dependent fashion. Results Here, we report on the use of this strain to define genetic loci involved in limiting rearrangements on an F, plasmid carrying the lac genes. Employing mini-Tn10 derivatives to generate insertions near or into genes of interest, we detected three loci (rmuA,B,C) that, when mutated, increase inversions. We have mapped, cloned and sequenced these mutator loci. In one case, inactivation of the sbcC gene leads to an increase in rearrangements, and in another, insertions near the recE gene lead to an even larger increase. The third gene involved in limiting inversions, rmuC, has been mapped at 86 min on the E. coli chromosome and encodes a protein of unknown function with a limited homology to myosins, and some of the SMC (structural maintenance of chromosomes) proteins. Conclusions This work presents the first example of an anti-mutator role of the sbcC,D genes, and defines a new gene (rmuC) involved in DNA recombination. [source] Electrosprayed Enzyme Coatings as Bioinspired Alternatives to Bioceramic Coatings for Orthopedic and Oral ImplantsADVANCED FUNCTIONAL MATERIALS, Issue 5 2009Lise T. de Jonge Abstract The biological performance of orthopedic and oral implants can be significantly improved by functionalizing the non-physiological metallic implant surface through the application of biologically active coatings. In this paper, a cost-effective alternative to traditional biomedical coatings for bone substitution through exploitation of the specific advantages of the electrospray deposition technique for the immobilization of the enzyme alkaline phosphatase (ALP) onto the implant surface is presented. Since ALP increases the local inorganic phosphate concentration required for physiological mineralization of hard tissues, ALP coatings will enable enzyme-mediated mineralization onto titanium surfaces. To evaluate the bone-bioactive capacity of the ALP-coated titanium surface, soaking experiments are performed. Although the purely inorganic so-called simulated body fluid is the standard in vitro procedure for predictive studies on potential bone bonding in vivo, an alternative testing solution is proposed that also contains organic phosphates (cell culture medium supplemented with the organic ,-b; -glycerophosphate (,-b; -GP) and serum proteins), thereby resembling the in vivo conditions more closely. Under these physiological conditions, the electrosprayed ALP coatings accelerated mineralization onto the titanium surface as compared to noncoated implant material by means of enzymatic pathways. Therefore, this novel approach toward implant fixation holds significant promise. [source] Pathways that produce volatile sulphur compounds from methionine in Oenococcus oeniJOURNAL OF APPLIED MICROBIOLOGY, Issue 6 2008A. Vallet Abstract Aims:, Determination of pathways involved in synthesis of volatile sulphur compounds (VSC) from methionine by Oenococcus oeni isolated from wine. Methods and Results:, Production of VSC by O. oeni from methionine was investigated during bacterial cultures and in assays performed in the presence of resting cells or protein fractions. Cells of O. oeni grown in a medium supplemented with methionine produced methanethiol, dimethyl disulphide, methionol and 3-(methylthio)propionic acid. Methional was also detected, but only transiently during the exponential growth phase. It was converted to methionol and 3-(methylthio) propionic acid in assays. Although this acid could be produced alternatively from 2-oxo-4-(methylthio) butyric acid (KMBA) by oxidative decarboxylation. In addition, KMBA was a precursor for methanethiol and dimethyl disulphide synthesis. Interestingly, assays with resting cells and protein fractions suggested that a specific enzyme could be involved in this conversion in O. oeni. Conclusion:, This work shows that methional and KMBA are the key intermediates for VSC synthesis from methionine in O. oeni. Putative enzymatic and chemical pathways responsible for the production of these VSC are discussed. Significance and impact of the study:, This work confirms the capacity of O. oeni to metabolize methionine and describes the involvement of potential enzymatic pathways. [source] Resveratrol: Preventing properties against vascular alterations and ageingMOLECULAR NUTRITION & FOOD RESEARCH (FORMERLY NAHRUNG/FOOD), Issue 5 2005Dominique Delmas Abstract Cardiovascular diseases are the leading cause of death in developed countries where the common pathological substrate underlying this process is atherosclerosis. Several new concepts have emerged in relation to mechanisms that contribute to the regulation of the vascular diseases and associated inflammatory effects. Recently, potential antioxidants (vitamin E, polyphenols) have received much attention as potential anti-atherosclerotic agents. Among the polyphenols with health benefic properties, resveratrol, a phytoalexin of grape, seem to be a good candidate protecting the vascular walls from oxidation, inflammation, platelet aggregation, and thrombus formation. In this review, we focus on the mechanism of resveratrol cardiovascular benefic effects. We analyze, in relation with the different steps of atherosclerotic process, the resveratrol properties at multiple levels, such as cellular signaling, enzymatic pathways, apoptosis, and gene expression. We show and discuss the relationship with reactive oxygen species, regulation of pro-inflammatory genes including cycloxygenases and cytokines in molecular inflammatory and aging processes, and how the regulation of these activites by resveratrol can lead to a prevention of vascular diseases. [source] Metabolism of fungicidal cyanooximes, cymoxanil and analogues in various strains of Botrytis cinereaPEST MANAGEMENT SCIENCE (FORMERLY: PESTICIDE SCIENCE), Issue 2 2009Frédérique Tellier Abstract BACKGROUND: The metabolism of cymoxanil [1-(2-cyano-2-methoxyiminoacetyl)-3-ethylurea] and fungicidal cyanooxime analogues was monitored on three phenotypes of Botrytis cinerea Pers. ex Fr. differing in their sensitivity towards cymoxanil. For this purpose, labelled [2- 14C]cymoxanil was added either to the culture medium of these strains or to its cell-free extract. RESULTS: In the culture medium of the most sensitive strain, four main metabolites were detected. Three were isolated and identified. Cymoxanil was quickly metabolised by at least three concurrent enzymatic pathways: (i) cyclisation leading, after hydrolysis, to ethylparabanic acid, (ii) reduction giving demethoxylated cymoxanil, (iii) hydrolysis followed by reduction and then acetylation leading to N -acetylcyanoglycine. In the cell-free extract of the same strain, only the first and the second of these enzymatic reactions occurred. By comparing the metabolic profile of the most sensitive strain with that of the less sensitive ones, it was shown that the decrease in sensitivity to cymoxanil correlates with a reduced acetylcyanoglycine formation. Among all metabolites, only N -acetylcyanoglycine is active against the most sensitive strain. Moreover, in a culture of this strain, two other fungicidal cyanooximes were also metabolised into this metabolite. CONCLUSION: The formation of N -acetylcyanoglycine may play an important role in the fungitoxicity of cymoxanil and cyanooxime derivatives. Copyright © 2008 Society of Chemical Industry [source] Neuronal nitric oxide synthase activity in rat urinary bladder detrusor: participation in M3 and M4 muscarinic receptor functionAUTONOMIC & AUTACOID PHARMACOLOGY, Issue 3 2005B. Orman Summary 1,The aim of this paper was to determine the different signalling cascades involved in contraction of the rat urinary bladder detrusor muscle mediated via muscarinic acetylcholine receptors (muscarinic AChR). Contractile responses, phosphoinositides (IPs) accumulation, nitric oxide synthase (NOS) activity and cyclic GMP (cGMP) production were measured to determine the reactions associated with the effect of cholinergic agonist carbachol. The specific muscarinic AChR subtype antagonists and different inhibitors of the enzymatic pathways involved in muscarinic receptor-dependent activation of NOS and cGMP were tested. 2,Carbachol stimulation of M3 and M4 muscarinic AChR increased contractility, IPs accumulation, NOS activity and cGMP production. All of these effects were selectively blunted by 4-DAMP and tropicamide, M3 and M4 antagonists respectively. 3,The inhibitors of phospholipase C (PLC), calcium/calmodulin (CaM), neuronal NOS (nNOS) and soluble guanylate cyclase, but not of protein kinase C and endothelial NOS (eNOS), inhibited the carbachol action on detrusor contractility. These inhibitors also attenuated the muscarinic receptor-dependent increase in cGMP and activation of NOS. 4,In addition, sodium nitroprusside and 8-bromo-cGMP, induced negative relaxant effect. 5,The results obtained suggest that carbachol activation of M3 and M4 muscarinic AChRs, exerts a contractile effect on rat detrusor that is accompanied by an increased production of cGMP and nNOS activity. The mechanism appears to occur secondarily to stimulation of IPs turnover via PLC activation. This in turn, triggers cascade reactions involving CaM, leading to activation of nNOS and soluble guanylate cyclase. They, in turn, exert a modulator inhibitory cGMP-mediated mechanism limiting the effect of muscarinic AChR stimulation of the bladder. [source] Proteomics: Recent Applications and New TechnologiesBASIC AND CLINICAL PHARMACOLOGY & TOXICOLOGY, Issue 5 2006Mollisa M. Elrick Proteomic analyses have recently been conducted on tissues, biofluids, subcellular components and enzymatic pathways as well as various disease and toxicological states, in both animal models and man. In addition, several recent studies have attempted to integrate proteomics data with genomics and/or metabonomics data in a systems biology approach. The translation of proteomic technology and bioinformatics tools to clinical samples, such as in the areas of disease and toxicity biomarkers, represents one of the major opportunities and challenges facing this field. An ongoing challenge in proteomics continues to be the analysis of the serum proteome due to the vast number and complexity of proteins estimated to be present in this biofluid. Aside from the removal of the most abundant proteins, a number of interesting approaches have recently been suggested that may help reduce the overall complexity of serum analysis. In keeping with the increasing interest in applications of proteomics, the tools available for proteomic analyses continue to improve and expand. For example, enhanced tools (such as software and labeling procedures) continue to be developed for the analysis of 2D gels and protein quantification. In addition, activity-based probes are now being used to tag, enrich and isolate distinct sets of proteins based on enzymatic activity. One of the most active areas of development involves microarrays. Antibody-based microarrays have recently been released as commercial products while numerous additional capture agents (e.g. aptamers) and many additional types of microarrays are being explored. [source] | ||||||||||||||||