Sulfur-containing amino acids block stretch-dependent K+ channels and nitrergic responses in the murine colon.
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1. Efforts to determine the role of stretch-dependent K(+) (SDK) channels in enteric inhibitory neural responses in gastrointestinal muscles are difficult due to a lack of blocking drugs for SDK channels. 2. SDK channels are blocked by sulfur-containing amino acids. These compounds reduced the open probability of SDK channels in on and off-cell patches of murine colonic myocytes. L-Methionine was the most selective and had little or no effect on other known K(+) conductances in colonic myocytes. 3. Application of L-cysteine, L-methionine or DL-homocysteine depolarized intact muscles and enhanced spontaneous contractions. D-Stereoisomers of these amino acids were less effective than L-stereoisomers. 4. Pretreatment of muscles with tetrodotoxin, N(W)-nitro-L-arginine or 1H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one reduced the depolarization responses to these compounds, suggesting that spontaneous neural activity and release of NO tonically activates SDK channels. 5. Nitrergic responses to nerve stimulation were reduced by sulfur-containing amino acids. 6. These data suggest that nitrergic inhibitory junction potentials are mediated, in part, by activation of SDK channels in murine colonic muscles. (+info)
Desulfonatronum cooperativum sp. nov., a novel hydrogenotrophic, alkaliphilic, sulfate-reducing bacterium, from a syntrophic culture growing on acetate.
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A novel alkaliphilic, sulfate-reducing bacterium was isolated from a syntrophic acetate-decomposing community enriched from samples of the soda lake Khadin, Tuva, Russia; the isolate was designated strain Z-7999(T). Cells of strain Z-7999(T) were vibrioid, Gram-negative, 0.4-0.5 x 1.0-2.5 microm and motile by means of a polar flagellum. The temperature range for growth was 15-40 degrees C, with an optimum of 35-38 degrees C. The pH range for growth was 6.7-10.3, with an optimum of pH 8.0-9.0. The NaCl concentration range for growth was 1-80 g l(-1). The novel isolate was obligately anaerobic, was alkaliphilic with a broad pH range and had an obligate requirement for carbonate ions in the growth medium. In the presence of sulfate as electron acceptor, it grew with hydrogen, formate and lactate. It was not able to ferment sugars, organic acids, amino acids or peptides. During growth on formate, strain Z-7999(T) reduced sulfite and thiosulfate to sulfide. It was able to grow lithoheterotrophically with sulfate and formate when acetate was added as a carbon source for biosynthesis of biomass. The G + C content of the genomic DNA of strain Z-7999(T) was 56.5 mol%. Results of comparative 16S rRNA gene sequence analyses revealed that strain Z-7999(T) was part of the delta-Proteobacteria and clustered with other members of the genus Desulfonatronum (similarity values of 95.2 and 95.3 % to Desulfonatronum lacustre and Desulfonatronum thiodismutans, respectively). DNA-DNA hybridization with D. lacustre was 37 %. On the basis of physiological and phylogenetic data, it is proposed that strain Z-7999(T) (= DSM 16749(T) = VKM B-2329(T)) should be placed in the genus Desulfonatronum as a representative of a novel species, Desulfonatronum cooperativum sp. nov. (+info)
Allyl disulfide as donor and cyanide as acceptor of sulfane sulfur in the mouse tissues.
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Cytoplasm of mammalian glial cells was reported to contain Gomori-positive cytoplasmic granulation (GPCG), whose biological role is unknown. The present study attempted to discover conditions facilitating GPCG formation and to elucidate their relationship with sulfane sulfur metabolism. To address these problems, we investigated in vivo the effect of both allyl disulfide (DADS), occurring in garlic (sulfane sulfur donor) and cyanide (sulfane sulfur acceptor) on number of GPCG-containing glial cells in the mouse brain. In parallel, sulfane sulfur level and activity of rhodanese and 3-mercaptopyruvate sulfurtransferase (MpST) were determined in the mouse brain and liver. Cyanide caused a drop in GPCG number in the brain, while activity of sulfurtransferases and sulfane sulfur level remained unchanged. Slight but significant cyanide-induced rise in MpST activity was observed only in the liver, which indicates a possibility of enhancement of its detoxification in reaction with mercapropyruvate in this organ. DADS, a sulfur donor, increased GPCG number in the brain, whereas activity of sulfurtransferases and sulfane sulfur level did not change. However, in the liver, DADS elevated both sulfurtransferase activity and sulfane sulfur level. These observations suggest that DADS can constitute a source of sulfane sulfur for the liver, thereby activating anaerobic sulfur metabolism and sulfane sulfur transfer. Consequently, this leads to the increase in sulfane sulfur level in plasma, in which it is transported in the form of albumin hydropersulfides and can be used for cyanide detoxification or stored in glial cells as GPCG. Therefore, it is not excluded that GPCG observed in the brain of mice and other mammals can be a source and a store of sulfane sulfur in mammals. (+info)
Sulfate-dependent repression of genes that function in organosulfur metabolism in Bacillus subtilis requires Spx.
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Oxidative stress in Bacillus subtilis results in the accumulation of Spx protein, which exerts both positive and negative transcriptional control over a genome-wide scale through its interaction with the RNA polymerase alpha subunit. Previous microarray transcriptome studies uncovered a unique class of genes that are controlled by Spx-RNA polymerase interaction under normal growth conditions that do not promote Spx overproduction. These genes were repressed by Spx when sulfate was present as a sole sulfur source. The genes include those of the ytmI, yxeI, and ssu operons, which encode products resembling proteins that function in the uptake and desulfurization of organic sulfur compounds. Primer extension and analysis of operon-lacZ fusion expression revealed that the operons are repressed by sulfate and cysteine; however, Spx functioned only in sulfate-dependent repression. Both the ytmI operon and the divergently transcribed ytlI, encoding a LysR-type regulator that positively controls ytmI operon transcription, are repressed by Spx in sulfate-containing media. The CXXC motif of Spx, which is necessary for redox sensitive control of Spx activity in response to oxidative stress, is not required for sulfate-dependent repression. The yxeL-lacZ and ssu-lacZ fusions were also repressed in an Spx-dependent manner in media containing sulfate as the sole sulfur source. This work uncovers a new role for Spx in the control of sulfur metabolism in a gram-positive bacterium under nonstressful growth conditions. (+info)
A simple sequential incubation method for deconvoluting the complicated sequential metabolism of capravirine in humans.
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Capravirine, a non-nucleoside reverse transcriptase inhibitor for the treatment of human immunodeficiency virus type 1, undergoes extensive oxygenations to numerous sequential metabolites in humans. Because several possible oxygenation pathways may be involved in the formation and/or sequential metabolism of a single metabolite, it is very difficult or even impossible to determine the definitive pathways and their relative contributions to the overall metabolism of capravirine using conventional approaches. For this reason, a human liver microsome-based "sequential incubation" method has been developed to deconvolute the complicated sequential metabolism of capravirine. In brief, the method includes three fundamental steps: 1) 30-min primary incubation of [(14)C]capravirine, 2) isolation of (14)C metabolites from the primary incubate, and 3) 30-min sequential incubation of each isolated (14)C metabolite supplemented with an ongoing (30 min) microsomal incubation with nonlabeled capravirine. Based on the extent of both the disappearance of the isolated precursor (14)C metabolites and the formation of sequential (14)C metabolites, definitive oxygenation pathways of capravirine were assigned. In addition, the percentage contribution of a precursor metabolite to the formation of each of its sequential metabolites (called sequential contribution) and the percentage contribution of a sequential metabolite formed from each of its precursor metabolites (called precursor contribution) were determined. An advantage of this system is that the sequential metabolism of each isolated (14)C metabolite can be monitored selectively by radioactivity in the presence of all relevant metabolic components (i.e., nonlabeled parent and its other metabolites). This methodology should be applicable to mechanistic studies of other compounds involving complicated sequential metabolic reactions when radiolabeled materials are available. (+info)
Composition, stability, and bioavailability of garlic products used in a clinical trial.
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In support of a new clinical trial designed to compare the effects of crushed fresh garlic and two types of garlic supplement tablets (enteric-coated dried fresh garlic and dried aged garlic extract) on serum lipids, the three garlic products have been characterized for (a) composition (14 sulfur and 2 non-sulfur compounds), (b) stability of suspected active compounds, and (c) availability of allyl thiosulfinates (mainly allicin) under both simulated gastrointestinal (tablet dissolution) conditions and in vivo. The allyl thiosulfinates of blended fresh garlic were stable for at least 2 years when stored at -80 degrees C. The dissolution release of thiosulfinates from the enteric-coated garlic tablets was found to be >95%. The bioavailability of allyl thiosulfinates from these tablets, measured as breath allyl methyl sulfide, was found to be complete and equivalent to that of crushed fresh garlic. S-Allylcysteine was stable for 12 months at ambient temperature. The stability of the suspected active compounds under the conditions of the study and the bioavailability of allyl thiosulfinates from the dried garlic supplement have validated the use of these preparations for comparison in a clinical trial. (+info)
Involvement of a branched-chain aminotransferase in production of volatile sulfur compounds in Yarrowia lipolytica.
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The enzymatic degradation of L-methionine and the subsequent formation of volatile sulfur compounds (VSCs) are essential for the development of the typical flavor in cheese. In the yeast Yarrowia lipolytica, the degradation of L-methionine was accompanied by the formation of the transamination product 4-methylthio-2-oxobutyric acid. A branched-chain aminotransferase gene (YlBCA1) of Y. lipolytica was amplified, and the L-methionine-degrading activity and the aminotransferase activity were measured in a genetically modified strain and compared to those of the parental strain. Our work shows that L-methionine degradation via transamination is involved in formation of VSCs in Y. lipolytica. (+info)
Effects of a series of organosulfur compounds on mitotic arrest and induction of apoptosis in colon cancer cells.
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We previously reported that the garlic-derived compound S-allylmercaptocysteine (SAMC) causes growth inhibition, mitotic arrest, and induction of apoptosis in SW480 human colon cancer cells by inducing microtubule depolymerization and c-Jun NH(2) terminus kinase-1 activation. In the present study, we compared the aforementioned effects of SAMC to those of a series of garlic-derived and other organosulfur compounds. Among the 10 compounds tested, only SAMC, diallyl disulfide (DADS), and S-trityl-L-cysteine (trityl-cys) cause significant inhibition of cell growth with IC(50) values of 150, 56, and 0.9 micromol/L, respectively. These three compounds also induce G(2)-M cell cycle arrest and apoptosis. Further studies reveal that, like SAMC, the garlic-derived compound DADS exerts antiproliferative effects by binding directly to tubulin and disrupting the microtubule assembly, thus arresting cells in mitosis and triggering mitochondria-mediated signaling pathways that lead to apoptosis. However, the synthetic compound trityl-cys exerts its effect on M-phase arrest and growth inhibition by mechanisms that involve spindle impairment but do not involve disruption of microtubule structure or dynamics. Furthermore, trityl-cys does not induce marked loss of mitochondrial membrane potential or release of cytochrome c, but it does induce caspase-3 activation and poly(ADP-ribose) polymerase cleavage. Structure-function analysis suggests that both the allyl and the disulfide moieties are important features for the antiproliferative effects of SAMC and DADS. These findings may be useful in the identification, synthesis, and development of organosulfur compounds that have anticancer activity. (+info)