Sulfate assimilation in Rhodopseudomonas globiformis

Rhodopseudomonas globiformis is able to grow on sulfate as sole source of sulfur, but only at concentrations below 1 mM. Good growth was observed with thiosulfate, cysteine or methionine as sulfur sources. Tetrathionate supported slow growth. Sulfide and sulfite were growth inhibitory. Growth inhibition by higher sulfate concentrations was overcome by the addition of O-acetylserine, which is known as derepressor of sulfate-assimilating enzymes, and by reduced glutathione. All enzymes of the sulfate assimilation pathway. ATP-sulfurylase, adenylylphosphate-sulfotransferase, thiosulfonate reductase and O-acetylserine sulfhydrylase are present in R. globiformis. Sulfate was taken up by the cells and the sulfur incorporated into the amino acids cysteine, methionine and homocysteine. It is concluded, that the failure of R. globiformis to grow on higher concentrations of sulfate is caused by disregulation of the sulfate assimilation pathway. Some preliminary evidence for this view is given in comparing the activities of some of the involved enzymes after growth on different sulfur sources and by examining the effect of O-acetylserine on these activities.Abbreviations DTE dl-dithioerythritol - APS adenosine 5-phosphosulfate, adenylyl sulfate - PAPS 3-phosphoadenosine 5-phosphosulfate, 3-phosphoadenylylsulfate     

Studies on the incorporation of labelled sulphate into cells and cell-free extracts of Nitrosomonas europaea

Summary The incorporation of [³⁵S]sulphate was followed into the washed cell suspensions of Nitrosomonas europaea. Thus bound sulphate, sulphite, sulphide, cysteine, glutathione, homocysteine and methionine were found in the ethanol soluble fraction as well as in the residual hydrolysed protein fraction. Cysteic acid, methionine sulphoxide and methionine sulphone were detected in the residual protein. The reaction between sulphydryl groups and N-ethylmaleimide has been successfully used to stabilize the thiol compounds in cell-extracts and the derivatives thus obtained were separated by paper chromatography. As in other microorganisms, sulphate is first activated by ATP in Nitrosomonas before it is reduced. The formation of APS and PAPS has been studied. A pathway for the incorporation of [³⁵S]sulphate is proposed.Abbreviations POPOP 1,4-bis-(5-phenyloxazolyl-2)-benzene - PPO 2,5-diphenyloxazole - APS adenosine-5-phosphosulphate - PAPS adenosine-3-phosphate 5-phosphosulphate - ATP adenosine triphosphate - DNA-ase deoxyribonuclease - NEM N-ethylmaleimide - TCA trichloro-acetic acid - GSH glutathione     

Metabolism of Sulfur Compounds in Bacillus subtilis during Sporulation

Metabolism of various sulfur compounds in Bacillus subtilis during growth and sporulation was investigated by use of tracer techniques, in an attempt to clarify the mechanism involved in the formation of cystine rich protein of the spore coat.

Methionine, homocysteine, cystathionine, cysteine and some inorganic sulfur compounds (sulfate, sulfite and thiosulfate) were utilized by this organism as sulfur sources for its growth and sporulation. Biosynthesis of methionine from sulfate during growth was more or less inhibited by the addition of cysteine, homocysteine or cystathionine to the culture.

It is suggested from these results that in Bacillus subtilis methionine is synthesized from sulfate through cysteine, cystathionine and homocysteine as is the case in Salmonella or Neurospora. The results also suggest that the metabolism of sulfur-containing amino acids in Bacillus subtilis is strongly regulated by methionine and homocysteine.     

Structure of Colletochlorin from Colletotrichum nicotianae

Pseudomonas melanogenum ATCC 17806 required methionine, cysteine, cystine, cystathionine, homocysteine or homocystine for growth. However, the addition of these amino acids decreased remarkably l-DOPA (3,4-dihydroxyphenyl-l-alanine) production by the bacterium. l-DOPA production by the bacterium was further affected by the amount of the substrate, the method of its addition and by the addition of antioxidants, as was the case with Vibrio tyrosinaticus.

Under suitable conditions about 8 mg/ml of l-DOPA were produced from 8.6 mg/ml of l-tyrosine.     

Interrelated regulation of sulphur-containing amino-acid biosynthetic enzymes and folate-metabolizing enzymes in Aspergillus nidulans

In Aspergillus nidulans homocysteine can be metabolized both to cysteine and methionine. Mutants impaired in the main pathway of cysteine synthesis or in the sulphate assimilation pathway show a low pool of glutathione and elevated levels of homocysteine synthase and of the homocysteine-to-cysteine pathway enzymes. On the other hand, the level of methionine synthase and other enzymes of folate metabolism is depressed in these mutants. This anticoordinated regulation provides a mechanism controlling the partition of homocysteine between the two diverging pathways. Homocysteine synthase was found derepressed, along with folate enzymes, in a strain carrying a mutation which suppresses mutations in metA, metB and metG genes. These results indicate that homocysteine synthase can be regarded as the enzyme of an alternative pathway of methionine synthesis and strongly suggest that the regulatory mechanisms governing sulphur-containing amino acid and folate metabolisms are interrelated.     

Effect of regulatory mutations of sulphur metabolism on the levels of cysteine- and homocysteine-synthesizing enzymes in Neurospora crassa

1. Regulation of four enzymes involved in cysteine and homocysteine synthesis, i.e. cysteine synthase (EC 4.2.99.8), homocysteine synthase (EC 4.1.99.10), cystathionine beta-synthase (EC 2.1.22) and gamma-cystathionase (EC 4.4.1.1) was studied in the wild type and sulphur regulatory mutants of Neurospora crassa. 2. Homocysteine synthase and cystathionine beta-synthase were found to be regulatory enzymes but only the former is under control of the cys-3 - scon system regulating several enzymes of sulphur metabolism, including gamma-cystathionase. 3. The results obtained with the mutants strongly suggest that homocysteine synthase plays a physiological role as an enzyme of the alternative pathway of methionine synthesis. Cysteine synthase activity was similar in all strains examined irrespective of growth conditions. 4. The sconc strain with derepressed enzymes of sulphur metabolism showed an increased pool of sulphur amino acids, except for methionine. Particularly characteristic for this pool is a high content of hypotaurine, a product of cysteine catabolism.     

Regulation of homocysteine metabolizing enzymes in Aspergillus nidulans

Summary Mutants of Aspergillus nidulans blocked in the main pathway of cysteine synthesis show an elevated level of the enzymes involved in the synthesis of cysteine from homocysteine i.e. cystathionine -synthase and -cystathionase and a depressed level of homocysteine methyltransferase. This results in a considerable change in the sulfur amino acids pool as compared to the wild type. Upon addition of cysteine to the growth medium the first two enzymes are repressed while the level of the third one increases. These data indicate that the two diverging pathways of homocysteine metabolism are anti-coordinately regulated.     

A highly selective phosphorescent chemodosimeter for cysteine and homocysteine based on platinum(II) complexes

A platinum(II) complex Pt(phen)CCC₆H₄CHO has been demonstrated as a highly selective phosphorescent chemodosimeter for cysteine and homocysteine. Upon addition of homocysteine or cysteine to its acetonitrile-water solution, an evident luminescent color variation from green to orange was observed. This red-shift of phosphorescence emission could be attributed to formation of a thiazinane through a selective reaction of aldehyde group of Pt(phen)CCC₆H₄CHO with cysteine/homocysteine, which was confirmed by ¹H NMR investigation and density functional theory (DFT) calculations. The further selectivity investigation indicates that it displays high selectivity for homocysteine and cysteine, and therefore can be utilized as cysteine/homocysteine phosphorescent chemodosimeter.     

Synergism between different transport systems stimulates the uptake of neutral amino acids by isolated brain microvessels

Summary The polar long-chain amino acids glutamine and methionine can be transported across the endothelial cells of brain microvessels either by an L-system which operates by a facilitated diffusion, exchanging mechanism, or by a concentrating, energy-dependent A-system. The presence of glutamine and/or of methionine can induce a synergism between the two transport systems which results, by a transstimulation mechanism, in a net increased uptake of neutral hydrophobic aminoacids. The methionine analog S-methylthiocysteine, which is the mixed disulfide resulting from the combination of methanethiol with cysteine, behaves similarly to methionine in stimulating the uptake of neutral hydrophobic amino acids. The same transstimulating effect can even be obtained in collagenase-treated, A-system-deprived microvessels by inducing the direct formation of S-methylthiocysteine within the cytoplasmic compartment of the endothelial cells.Abbreviations MeAIB -methylaminoisobutyric acid - BCH DL--aminobicyclo-2,2,1-heptanecarboxylic acid - HEPES N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid     

Ethionine and Methionine Metabolism by the Chrysomonad Flagellate Prymnesium parvum

SYNOPSIS. Ethionine or methionine can serve as sole nitrogen source for growth of Prymnesium parvum. Both amino acids are taken up as such at a ratio of 2 : 1 methionine/ethionine. Ethionine is totally de-ethylated in the cell, while methionine is probably only partially de-methylated. The homocysteine moiety of both amino acids is similarly metabolised to form cysteine or re-methylated to form methionine. De-ethylation of ethionine seems how P. parvum avoids its antimetabolic effect     

Nutrition and health relevant regulation of intestinal sulfur amino acid metabolism

Sulfur amino acids (SAA), particularly methionine and cysteine, are critical for the gut to maintain its functions including the digestion, absorption and metabolism of nutrients, the immune surveillance of the intestinal epithelial layer and regulation of the mucosal response to foreign antigens. However, the metabolism of SAA in the gut, specifically the transmethylation of methionine, will result in a net release of homocysteine, which is shown to be associated with cardiovascular disease and stroke. Furthermore, the extensive catabolism of dietary methionine by the intestine or by luminal microbes may result in a decrease in nutritional efficiency. Therefore, the regulation of SAA metabolism in the gut is not only nutritionally relevant, but also relevant to the overall health and well-being. The superiority of dl-2-hydroxy-4-methylthiobutyrate to dl-methionine in decreasing homocysteine production, alleviating stress responses, and reducing the first-pass intestinal metabolism of dietary methionine may provide a promising implication for nutritional strategies to manipulate SAA metabolism and thus to improve the nutrition and health status of animals and perhaps humans.     

Ethionine and Methionine Metabolism by the Chrysomonad Flagellate Ochromonas danica

SYNOPSIS. Growth of Ochromonas danica is competitively inhibited by ethionine. Inhibition can be reversed by methionine. Inhibition indexes of the effect of ethionine on growth and methionine incorporation into proteins are 1 and 4, respectively. Inside the cell, methionine is partially de-methylated and metabolized to form cysteine. Ethionine is partially de-ethylated, and the homocysteine moiety is either re-methylated to form methionine or further metabolized to form cysteine. Ethionine is also incorporated into proteins of O. danica. The kind of metabolic interference, expressed by inhibition of growth, and correlated with incorporation of ethionine, is yet unknown.     

Enzymatic lesions in methionine mutants of Aspergillus nidulans: role and regulation of an alternative pathway for cysteine and methionine synthesis.

In Aspergillus nidulans the pathway involving cystathionine formation is the main one for homocysteine synthesis. Mutants lacking cystathionine gamma-synthase or beta-cystathionase are auxotrophs suppressible by: (i) mutations in the main pathway of cysteine synthesis (cysA1, cysB1, and cysC1), (ii) mutations causing stimulation of cysteine catabolism (su101), and (iii) mutations in a presumed regulatory gene (suAmeth). A relative shortage of cysteine in the first group of suppressors causes a derepression of homocysteine synthase, the enzyme involved in the alternative pathway of homocysteine synthesis. A similar derepression is observed in the suAmeth strain. Homocysteine synthesized by this pathway serves as precursor for cysteine and methionine synthesis. A mutant with altered homocysteine synthase is a prototroph, indicating that this enzyme is not essential for the fungus.     

Methionine metabolism and multiple sclerosis

Context: Methylation reactions are particularly important in the brain and their inhibition can lead to a number of serious pathologies. Multiple sclerosis is one of the most common neurological disorders caused by interaction of genetic and environmental factors, but little is known about its cause or factors that contribute to the disorder. Although multiple sclerosis is primarily regarded as demyelinating disorder, there are no many articles focusing on methionine determination.

Objective: The aim of this work was to investigate whether serum methionine and its related compounds like homocysteine, cysteine, glutathione and asymmetric dimethylarginine were changed in multiple sclerosis patients.

Materials and methods: Sulphur-containing compounds were determined by using high-performance liquid chromatography with electrochemical detection in a single run for providing more complex view on methionine metabolism and asymmetric dimetylarginine was measured by a commercial enzyme-linked immunosorbent assay kit.

Results: Methionine and glutathione were decreased, but homocysteine, asymmetric dimethylarginine and cysteine were unchanged in patients with multiple sclerosis compared with controls.

Conclusions: Methionine and glutathione seem to be potential biomarkers for prognosis of the disease.     

Alterations in the metabolomics of sulfur-containing substances in rat kidney by betaine

Earlier studies have shown that betaine administration may modulate the metabolism of sulfur amino acids in the liver. In this study, we determined the changes in the metabolomics of sulfur-containing substances induced by betaine in the kidney, the other major organ actively involved in the transsulfuration reactions. Male rats received betaine (1 %) in drinking water for 2 weeks before killing. Betaine intake did not affect betaine–homocysteine methyltransferase activity or its protein expression in the renal tissue. Expression of methionine synthase was also unchanged. However, methionine levels were increased significantly both in plasma and kidney. Renal methionine adenosyltransferase activity and S-adenosylmethionine concentrations were increased, but there were no changes in S-adenosylhomocysteine, homocysteine, cysteine levels or cystathionine β-synthase expression. γ-Glutamylcysteine synthetase expression or glutathione levels were not altered, but cysteine dioxygenase and taurine levels were decreased significantly. In contrast, betaine administration induced cysteine sulfinate decarboxylase and its metabolic product, hypotaurine. These results indicate that the metabolomics of sulfur-containing substances in the kidney is altered extensively by betaine, although the renal capacity for methionine synthesis is unresponsive to this substance unlike that of the liver. It is suggested that the increased methionine availability due to an enhancement of its uptake from plasma may account for the alterations in the metabolomics of sulfur-containing substances in the kidney. Further studies need to be conducted to clarify the physiological/pharmacological significance of these findings.     

Radiation Chemistry of Foods

When 10^?3m cysteine solution was irradiated in the presence of glucose at the concentration of ten-fold of cysteine, the G-values of products produced from cysteine were similar to those from 10^?3m cysteine solution. On the other hand, the yield of carbonyl compound from glucose was suppressed completely by interaction between cysteine and radicals which are secondarily produced from glucose.

Methionine could not suppress the yield of carbonyl compound from glucose, and, G-values of products from methionine varied in comparison with those from solution containing methionine only.

From the results using scavenger, it was concluded that oxidation to methionine sulfoxide and cleavage to α-aminobutyric acid was caused by OH and attack, respectively.     

Molecular cloning and expression analyses of mitochondrial and plastidic isoforms of cysteine synthase (O-acetylserine(thiol)lyase) fromArabidopsis thaliana

Summary Cysteme synthase, the key enzyme for fixation of inorganic sulfide, catalyses the formation of cysteine from O-acetylserine and inorganic sulfide. Here we report the cloning of cDNAs encoding cysteine synthase isoforms fromArabidopsis thaliana. The isolated cDNA clones encode for a mitochondrial and a plastidic isoform of cysteine synthase (O-acetylserine (thiol)-lyase, EC 4.2.99.8), designated cysteine synthase C (AtCS-C, CSase C) and B (AtCS-B; CSase B), respectively.AtCS-C andAtCS-B, having lengths of 1569-bp and 1421-bp, respectively, encode polypeptides of 430 amino acids (45.8 kD) and of 392 amino acids ( 41.8 kD), respectively. The deduced amino acid sequences of the mitochondrial and plastidic isoforms exhibit high homology even with respect to the presequences. The predicted presequence of AtCS-C has a N-terminal extension of 33 amino acids when compared to the plastidic isoform. Northern blot analysis showed thatAtCS-C is higher expressed in roots than in leaves whereas the expression ofAtCS-B is stronger in leaves. Furthermore, gene expression of both genes was enhanced by sulfur limitation which in turn led to an increase in enzyme activity in crude extracts of plants. Expression of theAtCS-B gene is regulated by light. The mitochondrial, plastidic and cytosolic (Hesse and Altmann, 1995) isoforms of cysteine synthase ofArabidopsis are able to complement a cysteine synthasedeficient mutant ofEscherichia coli unable to grow on minimal medium without cysteine, indicating synthesis of functional plant proteins in the bacterium. Two lines of evidence proved thatAtCS-C encodes a mitochondrial form of cysteine synthase; first, import ofin vitro translation products derived from AtCS-C in isolated intact mitochondria and second, Western blot analysis of mitochondria isolated from transgenic tobacco plants expressing AtCS-C cDNA/c-myc DNA fusion protein.Abbreviations CSase cysteine synthase The nucleotide sequence data reported will appear in the EMBL Database under the accession numbers X81973 forAtCS-C and X81698 forAtCS-B.     

Methylation demand: a key determinant of homocysteine metabolism

Elevated plasma homocysteine is a risk factor for cardiovascular disease and Alzheimer's disease. To understand the factors that determine the plasma homocysteine level it is necessary to appreciate the processes that produce homocysteine and those that remove it. Homocysteine is produced as a result of methylation reactions. Of the many methyltransferases, two are, normally, of the greatest quantitative importance. These are guanidinoacetate methyltransferase (that produces creatine) and phosphatidylethanolamine N-methyltransferase (that produces phosphatidylcholine). In addition, methylation of DOPA in patients with Parkinson's disease leads to increased homocysteine production. Homocysteine is removed either by its irreversible conversion to cysteine (transsulfuration) or by remethylation to methionine. There are two separate remethylation reactions, catalyzed by betaine:homocysteine methyltransferase and methionine synthase, respectively. The reactions that remove homocysteine are very sensitive to B vitamin status as both the transsulfuration enzymes contain pyridoxal phosphate, while methionine synthase contains cobalamin and receives its methyl group from the folic acid one-carbon pool. There are also important genetic influences on homocysteine metabolism.     

Iron inhibits neurotoxicity induced by trace copper and biological reductants

The extracellular microenvironment of the brain contains numerous biological redox agents, including ascorbate, glutathione, cysteine and homocysteine. During ischemia/reperfusion, aging or neurological disease, extracellular levels of reductants can increase dramatically owing to dysregulated homeostasis. The extracellular concentrations of transition metals such as copper and iron are also substantially elevated during aging and in some neurodegenerative disorders. Increases in the extracellular redox capacity can potentially generate neurotoxic free radicals from reduction of Cu(II) or Fe(III), resulting in neuronal cell death. To investigate this in vitro, the effects of extracellular reductants (ascorbate, glutathione, cysteine, homocysteine or methionine) on primary cortical neurons was examined. All redox agents except methionine induced widespread neuronal oxidative stress and subsequent cell death at concentrations occurring in normal conditions or during neurological insults. This neurotoxicity was totally dependent on trace Cu (0.4 M) already present in the culture medium and did not require addition of exogenous Cu. Toxicity involved generation of Cu(I) and H₂O₂, while other trace metals did not induce toxicity. Surprisingly, administration of Fe(II) or Fe(III) (2.5 M) completely abrogated reductant-mediated neurotoxicity. The potent protective activity of Fe correlated with Fe inhibiting reductant-mediated Cu(I) and H₂O₂ generation in cell-free assays and reduced cellular Cu uptake by neurons. This demonstrates a novel role for Fe in blocking Cu-mediated neurotoxicity in a high reducing environment. A possible pathogenic consequence for these phenomena was demonstrated by abrogation of Fe neuroprotection after pre-exposure of cultures to the Alzheimers amyloid beta peptide (A). The loss of Fe neuroprotection against reductant toxicity was greater after treatment with human A1–42 than with human A1–40 or rodent A1–42, consistent with the central role of A1–42 in Alzheimers disease. These findings have important implications for trace biometal interactions and free radical-mediated damage during neurodegenerative illnesses such as Alzheimers disease and old-age dementia.Abbreviations A amyloid beta - AD Alzheimers disease - Asc ascorbate - BC bathocuproine disulfonate - Cys cysteine - DCF 2,7-dichlorofluorescein - DTNB 5,5-dithiobis(2-nitrobenzoic acid) - FCS fetal calf serum - Glut glutamate - GSH reduced glutathione - GSSG oxidized glutathione - Hcys homocysteine - ICP-MS inductively coupled plasma mass spectrometry - MEM minimal essential media - Met methionine - MnTMPyP manganese tetrakis(1-methyl-4-pyridyl)porphyrin - MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide