Sequence of cDNA for rat cystathionine <i>γ</i>-lyase and comparison of deduced amino acid sequence with related <i>Escherichia coli</i> enzymes

Erickson, Paul F.; Maxwell, Ian H.; Su, Lih-Jen; Baumann, Marc; Glode, L. Michael

doi:10.1042/bj2690335

Cited by 128 publications

(93 citation statements)

References 44 publications

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“…Furthermore, we could see, by looking at Ca 2＋ imaging tests, that NaHS inhibited the [Ca 2＋ ]i oscillation in ICC. Two classes of pyridoxal-5'-phosphate-dependent enzymes, CBS and CSE are responsible for the majority of the endogenous production of H2S in mammalian tissues that use L-cysteine as the main substrate [19,20]. CSE is expressed in a range of mammalian cells and tissues, and it seems to be the main H2S-forming enzyme in the liver, kidney, and cardiovascular system and, CBS is mainly expressed in the liver and central nervous system [8,21].…”

Section: Discussionmentioning

confidence: 99%

The Inhibitory Effects of Hydrogen Sulfide on Pacemaker Activity of Interstitial Cells of Cajal from Mouse Small Intestine

Parajuli

Choi

Lee

et al. 2010

Korean J Physiol Pharmacol

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In this study, we studied whether hydrogen sulfide (H2S) has an effect on the pacemaker activity of interstitial cells of Cajal (ICC), in the small intestine of mice. The actions of H2S on pacemaker activity were investigated using whole-cell patch-clamp technique, intracellular Ca 2＋ analysis at 30 o C and RT-PCR in cultured mouse intestinal ICC. Exogenously applied sodium hydrogen sulfide (NaHS), a donor of hydrogen sulfide, caused a slight tonic inward current on pacemaker activity in ICC at low concentrations (50 and 100 μM), but at high concentration (500 μM and 1 mM) it seemed to cause light tonic inward currents and then inhibited pacemaker amplitude and pacemaker frequency, and also an increase in the resting currents in the outward direction. Glibenclamide or other potassium channel blockers (TEA, BaCl2, apamin or 4-aminopydirine) did not have an effect on NaHS-induced action in ICC. The exogenous application of carbonilcyanide p-triflouromethoxyphenylhydrazone (FCCP) and thapsigargin also inhibited the pacemaker activity of ICC as NaHS. Also, we found NaHS inhibited the spontaneous intracellular Ca 2＋ ([Ca 2＋ ]i) oscillations in cultured ICC. In doing an RT-PCR experiment, we found that ICC enriched population lacked mRNA for both CSE and CBS, but was prominently detected in unsorted muscle. In conclusion, H2S inhibited the pacemaker activity of ICC by modulating intracellular Ca 2＋ . These results can serve as evidence of the physiological action of H2S as acting on the ICC in gastrointestinal (GI) motility.

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Section: Discussionmentioning

confidence: 99%

The Inhibitory Effects of Hydrogen Sulfide on Pacemaker Activity of Interstitial Cells of Cajal from Mouse Small Intestine

Parajuli

Choi

Lee

et al. 2010

Korean J Physiol Pharmacol

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“…2) (39), the deduced amino acid sequence of CYS3 has highly significant homology with several enzymes involved in metabolism of sulfur-containing amino acids; E. coli CT 1y-synthase (8), ClT l-lyase (9), rat CIT ly-lyase (11), and S. cerevisiae OAS-OAH sulfhydrylase (6,24). All these proteins are known to have oligomeric structures composed of identical subunits of molecular weights ranging from 40,000 to 50,000 and to require PLP as a cofactor (11,17,44,47). No Figure 3 also shows the result of SDS-PAGE of the purified preparation of CYS3 gene product (lane P), indicating that the preparation obtained has a purity of 90% or higher.…”

Section: Methodsmentioning

confidence: 99%

Cloning and bacterial expression of the CYS3 gene encoding cystathionine gamma-lyase of Saccharomyces cerevisiae and the physicochemical and enzymatic properties of the protein

et al. 1993

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By screening a yeast genomic library, we isolated and characterized a gene rescuing the cysteine requirement in a "cys1" strain of Saccharomyces cerevisiae. Except for four residues in the open reading frame composed of 1,182 nucleotides, the DNA sequence was the same as that for the CYS3 (CYI1) gene, encoding cystathionine gamma-lyase (EC 4.4.1.1), and isolated previously as a cycloheximide-induced gene (B. Ono, K. Tanaka, K. Naito, C. Heike, S. Shinoda, S. Yamamoto, S. Ohmori, T. Oshima, and A. Toh-e, J. Bacteriol. 174:pp.3339-3347, 1992). S. cerevisiae "cys1" strains carry two closely linked mutations; one (cys1) causes a defect in serine O-acetyltransferase (EC 2.3.1.30), and another, designated cys3, impairs cystathionine gamma-lyase activity. Rescue of the cysteine requirement by the gene encoding cystathionine gamma-lyase is consistent with both defects being responsible for the cysteine auxotrophy. In an effort to further determine the physicochemical and enzymatic properties of this enzyme, a coding fragment was cloned into an Escherichia coli expression plasmid, and the protein was produced in the bacteria. The induced protein was extracted by sonication and purified to homogeneity through one course of DEAE-cellulose column chromatography. The yield of the protein was approximately 150 mg from cells cultured in 1 liter of L broth. The protein showed molecular weights of approximately 194,000 and 48,000 (for the subunit), suggesting a tetrameric structure. An s20,w value of 8.8 was estimated by centrifugation in a sucrose concentration gradient. No sulfhydryl groups were detected, which is consistent with the absence of cysteine residues in the coding sequence. The isoelectric point was at pH 5.2. The protein showed a number of cystathionine-related activities, i.e., cystathionine beta-lyase (EC 4.4.1.8), cystathionine gamma-lyase, and cystathionine gamma-synthase (EC 4.2.99.9) with L-homoserine as substrate. In addition, we demonstrated L-homoserine sulfhydrylase (adding H2S) activity but could find no detectable serine O-acteyltransferease activity. In this paper, we compare the enzymatic properties of the protein with those of homologous enzymes previously reported and discuss the possibility that this enzyme has a physiological role as cystathionine Beta-lyase and cystathionine gamma-synthase in addition to its previously described role as cystathionine gamma-lyase.

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“…Two pyridoxal-5Ј-phosphate-dependent enzymes, cystathionine ␤-synthase (EC 4.2.1.22) and cystathionine ␥-lyase (CSE) 1 (EC 4.4.1.1), are responsible for the endogenous production of H 2 S in mammalian tissues, which use L-cysteine as the main substrate (3)(4)(5). Cystathionine ␤-synthase is a predominant H 2 S-generating enzyme in the brain and nervous system (6), and CSE is mainly expressed in liver, kidney, and vascular smooth muscles (7,8).…”

mentioning

confidence: 99%

Cystathionine γ-Lyase Overexpression Inhibits Cell Proliferation via a H2S-dependent Modulation of ERK1/2 Phosphorylation and p21Cip/WAK-1

Yang¹,

Cao²,

et al. 2004

Journal of Biological Chemistry

150

117

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Cystathionine ␥-lyase (CSE) is a key enzyme in the trans-sulfuration pathway. CSE uses L-cysteine as a substrate to produce hydrogen sulfide (H 2 S). The CSE/H 2 S system has been shown to play an important role in regulating cellular functions in different systems. In the present study, we used CSE stably overexpressed HEK-293 cells to explore the effect of the CSE/H 2 S system on cell growth and proliferation. The overexpression of CSE resulted in increases in CSE mRNA levels, CSE proteins, and intracellular H 2 S production rates, as well as the inhibition of cell proliferation and DNA synthesis. These effects were accompanied by a sustained ERK activation and up-regulation of the cyclin-dependent kinase inhibitor p21Cip/WAK-1 . Blocking the action of ERK with U0126 inhibited the induction of p21Cip/WAK-1 , suggesting that ERK activation functions upstream of p21 Cip/WAK-1 activation to initiate the CSE overexpression-induced cell growth inhibition. The antiproliferative effect of CSE is likely mediated by endogenously produced H 2 S because the H 2 S scavenger methemoglobin (10 M) significantly decreased the H 2 S production rate and reversed the antiproliferative effect afforded by CSE. Exogenous H 2 S (100 M) also inhibited cell proliferation. However, the other CSE-catalyzed products, ammonium and pyruvate, failed to inhibit cell proliferation. Methemoglobin also abolished the inhibitory effect of exogenous H 2 S on cell proliferation. Moreover, exogenous H 2 S induced a sustained ERK and p21Cip/WAK-1 activation. These findings support the hypothesis that endogenously produced H 2 S may play a fundamental role in cell proliferation and survival.

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Sequence of cDNA for rat cystathionine γ-lyase and comparison of deduced amino acid sequence with related Escherichia coli enzymes

Cited by 128 publications

References 44 publications

The Inhibitory Effects of Hydrogen Sulfide on Pacemaker Activity of Interstitial Cells of Cajal from Mouse Small Intestine

The Inhibitory Effects of Hydrogen Sulfide on Pacemaker Activity of Interstitial Cells of Cajal from Mouse Small Intestine

Cloning and bacterial expression of the CYS3 gene encoding cystathionine gamma-lyase of Saccharomyces cerevisiae and the physicochemical and enzymatic properties of the protein

Cystathionine γ-Lyase Overexpression Inhibits Cell Proliferation via a H2S-dependent Modulation of ERK1/2 Phosphorylation and p21Cip/WAK-1

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