Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:4.2.1.22 (cystathionine beta-synthase)
965 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Four cDNA clones, rcs1, rcs2, rcs3 and rcs4, encoding cysteine synthase [O-acetylserine(thiol)lyase] were isolated from rice. The predicted amino acid sequences contain the conserved PXXSVKDR region characteristic of cysteine synthase, which includes the lysine residue that binds the cofactor, pyridoxal 5'-phosphate. Molecular phylogenic analysis suggests that, whereas rcs1 and rcs3 belong to the cytosolic isoform family, rcs2 and rcs4 form a new family of cysteine synthase. Transcript accumulation of each gene was examined for organ specificity, and also for response to sulfur, nitrogen and light. The rcs1 transcript accumulated in all organs examined, and was induced in shoots and roots upon sulfur starvation under non-limiting nitrogen conditions. The rcs2 transcript accumulated in shoots grown in the light, but disappeared almost completely by dark treatment. The rcs3 transcript was found more abundantly in roots than in shoots, and was reduced in the dark, as well as under sulfur and nitrogen deprivation. The rcs4 transcript was scarce in all organs examined. These observations indicate that cysteine synthase genes encode functionally distinct cysteine synthase isoforms, and that they are coordinately regulated by the availability of sulfur, nitrogen, and light.
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PMID:Four rice genes encoding cysteine synthase: isolation and differential responses to sulfur, nitrogen and light. 1009 15

Glutathione (GSH) levels are supposed to determine the vulnerability of many cells towards a wide array of insults. We investigated the effects of chronic inhibition of GSH synthesis and acute depletion of GSH on cerebellar granule neurons in vitro and determined cytoplasmic and mitochondrial GSH with relation to mitochondrial function and generation of reactive oxygen intermediates (ROI). l-buthionine sulfoximine (BSO), which irreversibly blocks gamma-glutamyl-cysteine synthase, led to a time- and concentration-dependent loss of cytoplasmic GSH, while mitochondrial GSH was relatively preserved. No increased generation of ROI was detected over 48 h and the mitochondrial membrane potential was largely maintained. Neuronal degeneration occurred when mitochondrial GSH levels had fallen below 50% of control after 24-36 h. In contrast, direct conjugation of mitochondrial and cytoplasmic GSH with etacrynic acid (EA), resulted in immediate loss of mitochondrial GSH, a large increase of ROI within 2 h, subsequent collapse of the mitochondrial membrane potential and complete cell death within 4-8 h. Electron microscopy studies revealed an as yet unknown change of the chromatin structure to a homogeneous granular pattern after BSO, while EA resulted in typical necrotic changes. No typical features of apoptosis, i.e., no chromatin condensation or DNA fragmentation were detected after GSH depletion after BSO or EA treatment.
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PMID:Glutathione depletion and neuronal cell death: the role of reactive oxygen intermediates and mitochondrial function. 1021 96

Cysteine 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 from Arabidopsis 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 and AtCS-B, having lengths of 1569-bp and 1421-bp, respectively, encode polypeptides of 430 amino acids (approximately 45.8 kD) and of 392 amino acids (approximately 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 that AtCS-C is higher expressed in roots than in leaves whereas the expression of AtCS-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 the AtCS-B gene is regulated by light. The mitochondrial, plastidic and cytosolic (Hesse and Altmann, 1995) isoforms of cysteine synthase of Arabidopsis are able to complement a cysteine synthase-deficient mutant of Escherichia coli unable to grow on minimal medium without cysteine, indicating synthesis of functional plant proteins in the bacterium. Two lines of evidence proved that AtCS-C encodes a mitochondrial form of cysteine synthase; first, import of in 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.
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PMID:Molecular cloning and expression analyses of mitochondrial and plastidic isoforms of cysteine synthase (O-acetylserine(thiol)lyase) from Arabidopsis thaliana. 1031 84

Cysteine and methionine biosynthesis was studied in Pseudomonas putida S-313 and Pseudomonas aeruginosa PAO1. Both these organisms used direct sulfhydrylation of O-succinylhomoserine for the synthesis of methionine but also contained substantial levels of O-acetylserine sulfhydrylase (cysteine synthase) activity. The enzymes of the transsulfuration pathway (cystathionine gamma-synthase and cystathionine beta-lyase) were expressed at low levels in both pseudomonads but were strongly upregulated during growth with cysteine as the sole sulfur source. In P. aeruginosa, the reverse transsulfuration pathway between homocysteine and cysteine, with cystathionine as the intermediate, allows P. aeruginosa to grow rapidly with methionine as the sole sulfur source. P. putida S-313 also grew well with methionine as the sulfur source, but no cystathionine gamma-lyase, the key enzyme of the reverse transsulfuration pathway, was found in this species. In the absence of the reverse transsulfuration pathway, P. putida desulfurized methionine by the conversion of methionine to methanethiol, catalyzed by methionine gamma-lyase, which was upregulated under these conditions. A transposon mutant of P. putida that was defective in the alkanesulfonatase locus (ssuD) was unable to grow with either methanesulfonate or methionine as the sulfur source. We therefore propose that in P. putida methionine is converted to methanethiol and then oxidized to methanesulfonate. The sulfonate is then desulfonated by alkanesulfonatase to release sulfite for reassimilation into cysteine.
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PMID:Pathways of assimilative sulfur metabolism in Pseudomonas putida. 1048 27

The enteric protist parasites Entamoeba histolytica and Entamoeba dispar possess a cysteine biosynthetic pathway, unlike their mammalian host, and are capable of de novo production of L-cysteine. We cloned and characterized cDNAs that encode the regulated enzyme serine acetyltransferase (SAT) in this pathway from these amoebae by genetic complementation of a cysteine-auxotrophic Escherichia coli strain with the amoebic cDNA libraries. The deduced amino acid sequences of the amoebic SATs exhibited, within the most conserved region, 36-52% identities with the bacterial and plant SATs. The amoebic SATs contain a unique insertion of eight amino acids, also found in the corresponding region of a plasmid-encoded SAT from Synechococcus sp., which showed the highest overall identities to the amoebic SATs. Phylogenetic reconstruction also revealed a close kinship of the amoebic SATs with cyanobacterial SATs. Biochemical characterization of the recombinant E. histolytica SAT revealed several enzymatic features that distinguished the amoebic enzyme from the bacterial and plant enzymes: 1) inhibition by L-cysteine in a competitive manner with L-serine; 2) inhibition by L-cystine; and 3) no association with cysteine synthase. Genetically engineered amoeba strains that overproduced cysteine synthase and SAT were created. The cysteine synthase-overproducing amoebae had a higher level of cysteine synthase activity and total thiol content and revealed increased resistance to hydrogen peroxide. These results indicate that the cysteine biosynthetic pathway plays an important role in antioxidative defense of these enteric parasites.
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PMID:Characterization of the gene encoding serine acetyltransferase, a regulated enzyme of cysteine biosynthesis from the protist parasites Entamoeba histolytica and Entamoeba dispar. Regulation and possible function of the cysteine biosynthetic pathway in Entamoeba. 1054 89

The enzymatic degradation of amino acids in cheese is believed to generate aroma compounds and therefore to be essential for flavor development. Cystathionine beta-lyase (CBL) can convert cystathionine to homocysteine but is also able to catalyze an alpha, gamma elimination. With methionine as a substrate, it produces volatile sulfur compounds which are important for flavor formation in Gouda cheese. The metC gene, which encodes CBL, was cloned from the Lactococcus lactis model strain MG1363 and from strain B78, isolated from a cheese starter culture and known to have a high capacity to produce volatile compounds. The metC gene was found to be cotranscribed with a downstream cysK gene, which encodes a putative cysteine synthase. The MetC proteins of both strains were overproduced in strain MG1363 with the NICE (nisin-controlled expression) system, resulting in a >25-fold increase in cystathionine lyase activity. A disruption of the metC gene was achieved in strain MG1363. Determination of enzymatic activities in the overproducing and knockout strains revealed that MetC is essential for the degradation of cystathionine but that at least one lyase other than CBL contributes to methionine degradation via alpha, gamma elimination to form volatile aroma compounds.
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PMID:Molecular and functional analyses of the metC gene of Lactococcus lactis, encoding cystathionine beta-lyase. 1061 1

Novel genes that are regulated in Clostridium perfringens by the two-component regulatory system, VirR/VirS, were identified using a differential display method. A plasmid library was constructed from C. perfringens chromosomal DNA, and the plasmids were hybridized with cDNA probes prepared from total RNA of wild-type strain 13 and its virR mutant derivative TS133. Three clones were identified that carry newly identified VirR/VirS-regulated genes, two of which were positively regulated and one of which was negatively regulated. Genes located on the identified clones were deduced by nucleotide sequencing, and the target genes of the VirR/VirS system were identified with a set of Northern hybridizations. A 4.9 kb mRNA transcribing the metB (cystathionine gamma-synthase), cysK (cysteine synthase) and ygaG (hypothetical protein) genes was negatively regulated, whereas 1.6 and 6.0 kb transcripts encoding ptp (protein tyrosine phosphatase) and cpd (2',3'-cyclic nucleotide 2'-phosphodiesterase) respectively, were shown to be positively regulated by the VirR/VirS system. The other gene, hyp7, whose transcript was positively regulated by the VirR/VirS system, was shown to activate the transcription of the colA (kappa-toxin) and plc (alpha-toxin) genes, but not the pfoA (theta-toxin) gene in C. perfringens. These results suggested that the global regulatory system VirR/VirS could regulate various genes, other than toxin genes, both positively and negatively and that the hyp7 gene might encode a novel regulatory factor for toxin production in C. perfringens.
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PMID:Identification of novel VirR/VirS-regulated genes in Clostridium perfringens. 1069 62

Our studies of cystathionine beta-synthase from Saccharomyces cerevisiae (yeast) are aimed at clarifying the cofactor dependence and catalytic mechanism and obtaining a system for future investigations of the effects of mutations that cause human disease (homocystinuria or coronary heart disease). We report methods that yielded high expression of the yeast gene in Escherichia coli and of purified yeast cystathionine beta-synthase. The absorption and circular dichroism spectra of the homogeneous enzyme were characteristic of a pyridoxal phosphate enzyme and showed the absence of heme, which is found in human and rat cystathionine beta-synthase. The absence of heme in the yeast enzyme facilitates spectroscopic studies to probe the catalytic mechanism. The reaction of the enzyme with L-serine in the absence of L-homocysteine produced the aldimine of aminoacrylate, which absorbed at 460 nm and had a strong negative circular dichroism band at 460 nm. The formation of this intermediate from the product, L-cystathionine, demonstrates the partial reversibility of the reaction. Our results establish the overall catalytic mechanism of yeast cystathionine beta-synthase and provide a useful system for future studies of structure and function. The absence of heme in the functional yeast enzyme suggests that heme does not play an essential catalytic role in the rat and human enzymes. The results are consistent with the absence of heme in the closely related enzymes O-acetylserine sulfhydrylase, threonine deaminase, and tryptophan synthase.
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PMID:Yeast cystathionine beta-synthase is a pyridoxal phosphate enzyme but, unlike the human enzyme, is not a heme protein. 1076 67

Three cDNA clones encoding putative cysteine synthases (O-acetylserine (thiol) lyase, EC 4.2.99.8) were isolated from Arabidopsis thaliana and designated AtcysC1, AtcysD1 and AtcysD2, respectively. Southern blot analyses suggested that the corresponding genes were present as a single copy, or at most two copies, in the A. thaliana genome. Escherichia coli complementation analyses confirmed that the cDNAs encode cysteine synthase and the corresponding proteins produced in E. coli clearly showed cysteine synthase activity. In addition, AtcysC1 protein showed beta-cyanoalanine synthase (EC 4.4.1.9) activity, but the other two did not. Kinetic analysis suggests that AtcysC1 actually functions as beta-cyanoalanine synthase rather than cysteine synthase in vivo. The mRNA accumulation of AtcysC1, AtcysD1 and AtcysD2 differed in various organs, but did not change markedly when A. thaliana seedlings were subjected to various stresses, including nutrient deprivation. In vivo targeting experiments indicated that AtcysD1 and AtcysD2 are cytoplasmic isozymes, and AtcysC1 is a mitochondrial isozyme.
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PMID:Three Arabidopsis genes encoding proteins with differential activities for cysteine synthase and beta-cyanoalanine synthase. 1084 60

beta-Cyano-alanine synthase (CAS; EC 4.4.1.9) plays an important role in cyanide metabolism in plants. Although the enzymatic activity of beta-cyano-Ala synthase has been detected in a variety of plants, no cDNA or gene has been identified so far. We hypothesized that the mitochondrial cysteine synthase (CS; EC 4.2.99. 8) isoform, Bsas3, could actually be identical to CAS in spinach (Spinacia oleracea) and Arabidopsis. An Arabidopsis expressed sequence tag database was searched for putative Bsas3 homologs and four new CS-like isoforms, ARAth;Bsas1;1, ARAth;Bsas3;1, ARAth;Bsas4;1, and ARAth;Bsas4;2, were identified in the process. ARAth;Bsas3;1 protein was homologous to the mitochondrial SPIol;Bsas3;1 isoform from spinach, whereas ARAth;Bsas4;1 and ARAth;Bsas4;2 proteins defined a new class within the CS-like proteins family. In contrast to spinach SPIol;Bsas1;1 and SPIol;Bsas2;1 recombinant proteins, spinach SPIol;Bsas3;1 and Arabidopsis ARAth;Bsas3;1 recombinant proteins exhibited preferred substrate specificities for the CAS reaction rather than for the CS reaction, which identified these Bsas3 isoforms as CAS. Immunoblot studies supported this conclusion. This is the first report of the identification of CAS synthase-encoding cDNAs in a living organism. A new nomenclature for CS-like proteins in plants is also proposed.
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PMID:beta-Cyanoalanine synthase is a mitochondrial cysteine synthase-like protein in spinach and Arabidopsis. 1088 65


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