Biogenesis of Hydrogen Sulfide and Thioethers by Cystathionine Beta-Synthase (original) (raw)
Related papers
The Quantitative Significance of the Transsulfuration Enzymes for H 2 S Production in Murine Tissues
Antioxidants & Redox Signaling, 2011
The enzymes of the transsulfuration pathway, cystathionine b-synthase (CBS) and cystathionine c-lyase (CSE), are important for the endogenous production of hydrogen sulfide (H 2 S), a gaseous signaling molecule. The relative contributions of CBS and CSE to H 2 S generation in different tissues are not known. In this study, we report quantification of CBS and CSE in murine liver and kidney and their contribution to H 2 S generation in these tissues and in brain at saturating substrate concentrations. We show that CBS protein levels are significantly lower than those of CSE; 60-fold and 20-fold in liver and kidney, respectively. Each enzyme is more abundant in liver compared with kidney, twofold and sixfold for CBS and CSE, respectively. At high substrate concentrations (20 mM each cysteine and homocysteine), the capacity for liver H 2 S production is approximately equal for CBS and CSE, whereas in kidney and brain, CBS constitutes the major source of H 2 S, accounting for *80% and *95%, respectively, of the total output. At physiologically relevant concentrations of substrate, and adjusting for the differences in CBS versus CSE levels, we estimate that CBS accounts for only 3% of H 2 S production by the transsulfuration pathway enzymes in liver. Antioxid. Redox Signal. 15, 363-372.
Enzymology of H 2 S Biogenesis, Decay and Signaling
Antioxidants & Redox Signaling, 2014
Significance: Hydrogen sulfide (H 2 S), produced by the desulfuration of cysteine or homocysteine, functions as a signaling molecule in an array of physiological processes including regulation of vascular tone, the cellular stress response, apoptosis, and inflammation. Recent Advances: The low steady-state levels of H 2 S in mammalian cells have been recently shown to reflect a balance between its synthesis and its clearance. The subversion of enzymes in the cytoplasmic trans-sulfuration pathway for producing H 2 S from cysteine and/or homocysteine versus producing cysteine from homocysteine, presents an interesting regulatory problem. Critical Issues: It is not known under what conditions the enzymes operate in the canonical trans-sulfuration pathway and how their specificity is switched to catalyze the alternative H 2 S-producing reactions. Similarly, it is not known if and whether the mitochondrial enzymes, which oxidize sulfide and persulfide (or sulfane sulfur), are regulated to increase or decrease H 2 S or sulfane-sulfur pools. Future Directions: In this review, we focus on the enzymology of H 2 S homeostasis and discuss H 2 S-based signaling via persulfidation and thionitrous acid. Antioxid. Redox Signal. 20, 770-782.
Biochemistry, 2011
Cystathionine β-synthase (CBS) catalyzes the first step in the transsulfuration pathway in mammals, i.e., the condensation of serine and homocysteine to produce cystathionine and water. Recently, we have reported a steady-state kinetic analysis of the three hydrogen sulfide (H 2 S)generating reactions that are catalyzed by human and yeast CBS (Singh et al (2009) J Biol Chem 284: 22457-66). In the current study, we report a pre-steady-state kinetic analysis of intermediates in the H 2 S-generating reactions catalyzed by yeast CBS (yCBS). Because yCBS does not have a heme cofactor, in contrast to human CBS, it is easier to observe reaction intermediates with yCBS. The most efficient route for H 2 S generation by yCBS is the β-replacement of the cysteine thiol by homocysteine. In this reaction, yCBS first reacts with cysteine to release H 2 S and forms an aminoacrylate intermediate (k obs =1.61 ± 0.04 mM −1 s −1 at low cysteine and 2.8 ± 0.1 mM −1 s −1 at high cysteine concentrations, at 20 °C), which has an absorption maximum at 465 nm. Homocysteine binds to the E•aminoacrylate intermediate with a bimolecular rate constant of 142 mM −1 s −1 and rapidly condenses to form the enzyme-bound external aldimine of cystathionine. The reactions could be partially rate limited by release of the products, cystathionine and H 2 S. Hydrogen sulfide (H 2 S)1, like nitric oxide and carbon monoxide, is a gaseous signaling molecule (1-3) that elicits a variety of physiological effects. In the cardiovascular system, H 2 S apparently functions as a vasorelaxant (4) and as a cardioprotective agent (5). A dosedependent decrease in murine blood pressure by sodium hydrosulfide has been reported (4). Other reported effects of H 2 S include protection against ischemia reperfusion injury and anti-inflammatory effects in tissues (5,6). In lower organisms like yeast, H 2 S plays a role in population synchronization during ultradian oscillations (7). There are two known mammalian enzymes that can directly generate H 2 S: cystathionase γ-lyase (CGL) and CBS (8). A third enzyme pair, 3-mercaptopyruvate sulfurtransferase together with cysteine aminotransferase, catalyzes the transfer of sulfur to an unknown acceptor and, in the presence of a reductant, can liberate H 2 S (3,9). The role of CGL-dependent H 2 S production in the vasculature is controversial with one group reporting development of age-related hypertension in CGL knockout mice (4) and another, a normotensive phenotype (10).
The Therapeutic Potential of Cystathionine β-Synthetase/Hydrogen Sulfide Inhibition in Cancer
Antioxidants & Redox Signaling, 2015
Significance: Cancer represents a major socioeconomic problem; there is a significant need for novel therapeutic approaches targeting tumor-specific pathways. Recent Advances: In colorectal and ovarian cancers, an increase in the intratumor production of hydrogen sulfide (H 2 S) from cystathionine b-synthase (CBS) plays an important role in promoting the cellular bioenergetics, proliferation, and migration of cancer cells. It also stimulates peritumor angiogenesis inhibition or genetic silencing of CBS exerts antitumor effects both in vitro and in vivo, and potentiates the antitumor efficacy of anticancer therapeutics. Critical Issues: Recently published studies are reviewed, implicating CBS overexpression and H 2 S overproduction in tumor cells as a tumorgrowth promoting ''bioenergetic fuel'' and ''survival factor,'' followed by an overview of the experimental evidence demonstrating the anticancer effect of CBS inhibition. Next, the current state of the art of pharmacological CBS inhibitors is reviewed, with special reference to the complex pharmacological actions of aminooxyacetic acid. Finally, new experimental evidence is presented to reconcile a controversy in the literature regarding the effects of H 2 S donor on cancer cell proliferation and survival. Future Directions: From a basic science standpoint, future directions in the field include the delineation of the molecular mechanism of CBS upregulation of cancer cells and the delineation of the interactions of H 2 S with other intracellular pathways of cancer cell metabolism and proliferation. From the translational science standpoint, future directions include the translation of the recently emerging roles of H 2 S in cancer into human diagnostic and therapeutic approaches. Antioxid. Redox Signal. 22, 424-448. Biological Effects of H 2 S with Relevance for Cancer Biology H 2 S, as a vasodilator and pro-angiogenic mediator Vasorelaxation is one of the first recognized biological effects of H 2 S. Often compared with NO, H 2 S exerts a concentration-dependent vasodilatory effect in blood vessels. The mechanisms of H 2 S-mediated vasodilation include the activation of K ATP channels, a variety of other channels, inhibition of phosphodiesterases, and a synergy with NO (132).
Scientific Reports, 2016
Hydrogen sulfide is an essential catabolite that intervenes in the pathophysiology of several diseases from hypertension to stroke, diabetes and pancreatitis. It is endogenously synthesized mainly by two pyridoxal-5′-phosphate-dependent enzymes involved in L-cysteine metabolism: cystathionine-ßsynthase (CBS) and cystathionine-γ-lyase (CSE). Research in this field is currently impaired by the lack of pharmacological tools such as selective enzymatic inhibitors that could target specifically only one of these pathways. We used a novel approach based on a hybrid method that includes drug design, synthetic biology, metabolomics and pharmacological assays to rationally design a new inhibitor selective for the CSE enzyme. The identification of this compound opens new frontiers towards a better understanding of the role of CSE over CBS in the pathophysiology of diseases where a role for the H 2 S pathway has been proposed and the development of new lead compounds that could target the CSE enzyme. Hydrogen sulfide (H 2 S), a colorless, flammable, water soluble gas with the characteristic smell of rotten eggs, has emerged as an important gaseous signaling molecule playing numerous roles in health and disease, along with CO and NO 1. Enzymatically generated H 2 S is mainly derived from two pyridoxal-5′-phosphate (PLP)-dependent enzymes responsible for the metabolism of L-cysteine (L-Cys): cystathionine-β-synthase (CBS) and cystathionine-γ-lyase (CSE) 2. A third pathway that catalyses the production of H 2 S from L-Cys via the combined action of 3-mercaptopyruvate sulfurtransferase and cysteine aminotransferase has also been described 3. This pathway is less well characterized and its role in determining the H 2 S levels in tissues still poorly understood. CBS and CSE are cytosolic enzymes which catalyse several H 2 S-generating reactions which convert L-Cys and/or homocysteine as substrates to L-cystathionine and pyruvate respectively 4-6. CBS was also originally considered to be the predominant enzyme for H 2 S production in the brain, indeed it is preferentially expressed in radial glia/astrocytes of adult and developing mouse brain 7,8 , whereas H 2 S synthesis in the heart and vasculature was attributed to CSE 9. More recent studies with improved markers have provided a broader picture of enzyme distribution. Because of the co-presence of both enzymes in specific pathway it is important to find inhibitors that selectively target only one enzyme. The most commonly used agents to inhibit H 2 S biosynthesis include propargylglycine (PAG), β-cyanoalanine (BCA) and aminooxyacetic acid (AOAA) 10,11. PAG is described as CSE selective inhibitor in fact it will not inhibit recombinant CBS even if used at 10 mM 12 ; moreover the crystal structure of the covalent complex PAG-CSE is the only described so far 13. However all of these compounds have a relatively low potency and cell permeability and are used at high concentrations (mM) 11. L-aminoethoxyvinylglycine (AVG) was also recently described as a potentially more potent and selective CSE inhibitor 12 but its mechanism is still uncharacterised. The inherent
Regulation of the redox metabolome and thiol proteome by hydrogen sulfide
Critical Reviews in Biochemistry and Molecular Biology, 2021
Overproduction of reactive oxygen species and compromised antioxidant defenses perturb intracellular redox homeostasis and is associated with a myriad of human diseases as well as with the natural process of aging. Hydrogen sulfide (H 2 S), which is biosynthesized by organisms ranging from bacteria to man, influences a broad range of physiological functions. A highly touted molecular mechanism by which H 2 S exerts its cellular effects is via post-translational modification of the thiol redox proteome, converting cysteine thiols to persulfides, in a process referred to as protein persulfidation. The physiological relevance of this modification in the context of specific signal transmission pathways remains to be rigorously established, while a general protective role for protein persulfidation against hyper-oxidation of the cysteine proteome is better supported. A second mechanism by which H 2 S modulates redox homeostasis is via remodeling the redox metabolome, targeting the electron transfer chain and perturbing the major redox nodes i.e., CoQ/ CoQH 2 , NAD + /NADH and FAD/FADH 2. The metabolic changes that result from H 2 S-induced redox changes fan out from the mitochondrion to other compartments. In this review, we discuss recent developments in elucidating the roles of H 2 S and its oxidation products on redox homeostasis and its role in protecting the thiol proteome.
Scientific Reports, 2023
Hydrogen sulfide (H 2 S) is a gaseous signaling molecule that participates in various signaling functions in health and diseases. The tetrameric cystathionine γ-lyase (CSE) contributes to H 2 S biogenesis and several investigations provide evidence on the pharmacological modulation of CSE as a potential target for the treatment of a multitude of conditions. D-penicillamine (D-pen) has recently been reported to selectively impede CSE-catalyzed H 2 S production but the molecular bases for such inhibitory effect have not been investigated. In this study, we report that D-pen follows a mixedinhibition mechanism to inhibit both cystathionine (CST) cleavage and H 2 S biogenesis by human CSE. To decipher the molecular mechanisms underlying such a mixed inhibition, we performed docking and molecular dynamics (MD) simulations. Interestingly, MD analysis of CST binding reveals a likely active site configuration prior to gem-diamine intermediate formation, particularly H-bond formation between the amino group of the substrate and the O3′ of PLP. Similar analyses realized with both CST and D-pen identified three potent interfacial ligand-binding sites for D-pen and offered a rational for D-pen effect. Thus, inhibitor binding not only induces the creation of an entirely new interacting network at the vicinity of the interface between enzyme subunits, but it also exerts long range effects by propagating to the active site. Overall, our study paves the way for the design of new allosteric interfacial inhibitory compounds that will specifically modulate H 2 S biogenesis by cystathionine γ-lyase. Abbreviations TSP Transsulfuration pathway PLP Pyridoxal 5′-phosphate CBS Cystathionine β-synthase CSE Cystathionine γ-lyase CST Cystathionine D-pen D-penicillamine PDB Protein data bank RMSD Root mean square deviation RMSF Root mean square fluctuation Hydrogen sulfide (H 2 S) is a gaseous signaling molecule that participates in various signaling functions in the vascular and neuronal systems, and in the regulation of the inflammatory response. It also contributes to the adaptive response and acts as a pleiotropic pro-resolving gaseous mediator in inflammatory diseases 1-6. In mammals, endogenous biogenesis of H 2 S is carried out by the transsulfuration pathway (TSP), constituted of two pyridoxal 5′-phosphate (PLP)-dependent enzymes, cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE), and by the mitochondrial catabolism of cysteine (Cys) that involves cysteine aminotransferase (CAT) in combination with 3-mercaptopyruvate sulfur transferase (3-MST). The TSP, with the methionine cycle to which it is connected, has singular significance in the metabolism of sulfur compounds and it also plays a role in the maintenance of cellular homeostasis by methylation through its control of S-adenosylmethionine (SAM) levels,
PLOS ONE, 2015
Urothelium, the epithelial lining the inner surface of human bladder, plays a key role in bladder physiology and pathology. It responds to chemical, mechanical and thermal stimuli by releasing several factors and mediators. Recently it has been shown that hydrogen sulfide contributes to human bladder homeostasis. Hydrogen sulfide is mainly produced in human bladder by the action of cystathionine-β-synthase. Here, we demonstrate that human cystathionine-β-synthase activity is regulated in a cGMP/PKG-dependent manner through phosphorylation at serine 227. Incubation of human urothelium or T24 cell line with 8-Bromo-cyclic-guanosine monophosphate (8-Br-cGMP) but not dibutyryl-cyclic-adenosine monophosphate (d-cAMP) causes an increase in hydrogen sulfide production. This result is congruous with the finding that PKG is robustly expressed but PKA only weakly present in human urothelium as well as in T24 cells. The cGMP/PKG-dependent phosphorylation elicited by 8-Br-cGMP is selectively reverted by KT5823, a specific PKG inhibitor. Moreover, the silencing of cystathionine-β-synthase in T24 cells leads to a marked decrease in hydrogen sulfide production either in basal condition or following 8-Br-cGMP challenge. In order to identify the phosphorylation site, recombinant mutant proteins of cystathionine-β-synthase in which Ser32, Ser227 or Ser525 was mutated in Ala were generated. The Ser227Ala mutant cystathionine-β-synthase shows a notable reduction in basal biosynthesis of hydrogen sulfide becoming unresponsive to the 8-Br-cGMP challenge. A specific antibody that recognizes the phosphorylated form of cystathionine-β-synthase has been produced and validated by using T24 cells and human urothelium. In conclusion, human cystathionine-β-synthase can be phosphorylated in a PKG-dependent manner at Ser227 leading to an increased catalytic activity.
Biogenesis of reactive sulfur species for signaling by hydrogen sulfide oxidation pathways
Nature chemical biology, 2015
The chemical species involved in H2S signaling remain elusive despite the profound and pleiotropic physiological effects elicited by this molecule. The dominant candidate mechanism for sulfide signaling is persulfidation of target proteins. However, the relatively poor reactivity of H2S toward oxidized thiols, such as disulfides, the low concentration of disulfides in the reducing milieu of the cell and the low steady-state concentration of H2S raise questions about the plausibility of persulfide formation via reaction between an oxidized thiol and a sulfide anion or a reduced thiol and oxidized hydrogen disulfide. In contrast, sulfide oxidation pathways, considered to be primarily mechanisms for disposing of excess sulfide, generate a series of reactive sulfur species, including persulfides, polysulfides and thiosulfate, that could modify target proteins. We posit that sulfide oxidation pathways mediate sulfide signaling and that sulfurtransferases ensure target specificity.