Hydrogen Sulfide: Redox Metabolism and Signaling

The recognition of hydrogen sulfide (H2S) as an endogenously produced gas with signaling potential has stimulated research on a multitude of physiological effects mediated in the cardiovascular, immune, gastrointestinal, genitourinary, endocrine, and central nervous systems. The heightened activity in the area of H2S biology led to convening of the first international conference on H2S in Shanghai in the summer of 2009 and to two Forum issues published in 2010 by Antioxidants & Redox Signaling on the physiological effects of H2S. Yet, fundamental questions regarding the biogenesis and regulation of H2S, the bioenergetics of its catabolism, its tissue concentrations, and elucidation of its molecular targets remain. Some of these issues are the subject of the current Forum on H2S. Antioxid. Redox Signal. 15, 339-341.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/90431/1/ars-2E2011-2E3912.pd

Enzymology of H2S Biogenesis, Decay and Signaling

Significance: Hydrogen sulfide (H2S), 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 H2S 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 H2S 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 H2S-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 H2S or sulfane-sulfur pools. Future Directions: In this review, we focus on the enzymology of H2S homeostasis and discuss H2S-based signaling via persulfidation and thionitrous acid. Antioxid. Redox Signal. 20, 770?782.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/140254/1/ars.2013.5339.pd

THE MANY FACES OF VITAMIN B12: CATALYSIS BY COBALAMIN-DEPENDENT ENZYMES

Vitamin B12 is a complex organometallic cofactor associated with three subfamilies of enzymes: the adenosylcobalamin-dependent isomerases, the methylcobalamin-dependent methyltransferases, and the dehalogenases. Different chemical aspects of the cofactor are exploited during catalysis by the isomerases and the methyltransferases. Thus, the cobalt-carbon bond ruptures homolytically in the isomerases, whereas it is cleaved heterolytically in the methyltransferases. The reaction mechanism of the dehalogenases, the most recently discovered class of B12 enzymes, is poorly understood. Over the past decade our understanding of the reaction mechanisms of B12 enzymes has been greatly enhanced by the availability of large amounts of enzyme that have afforded detailed structure-function studies, and these recent advances are the subject of this review

Cobalamin‐dependent methionine synthase

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/154369/1/fsb2004005005.pd

High Turnover Rates for Hydrogen Sulfide Allow for Rapid Regulation of Its Tissue Concentrations

Abstract Aims: Hydrogen sulfide (H2S) is a signaling molecule, which influences many physiological processes. While H2S is produced and degraded in many cell types, the kinetics of its turnover in different tissues has not been reported. In this study, we have assessed the rates of H2S production in murine liver, kidney, and brain homogenates at pH 7.4, 37°C, and at physiologically relevant cysteine concentrations. We have also studied the kinetics of H2S clearance by liver, kidney, and brain homogenates under aerobic and anaerobic conditions. Results: We find that the rate of H2S production by these tissue homogenates is considerably higher than background rates observed in the absence of exogenous substrates. An exponential decay of H2S with time is observed and, as expected, is significantly faster under aerobic conditions. The half-life for H2S under aerobic conditions is 2.0, 2.8, and 10.0?min with liver, kidney, and brain homogenate, respectively. Western-blot analysis of the sulfur dioxygenase, ETHE1, involved in H2S catabolism, demonstrates higher steady-state protein levels in liver and kidney versus brain. Innovation: By combining experimental and simulation approaches, we demonstrate high rates of tissue H2S turnover and provide estimates of steady-state H2S levels. Conclusion: Our study reveals that tissues maintain a high metabolic flux of sulfur through H2S, providing a rationale for how H2S levels can be rapidly regulated. Antioxid. Redox Signal. 17, 22?31.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/98481/1/ars%2E2011%2E4310.pd

Astrocytic Redox Remodeling by Amyloid Beta Peptide

Abstract Astrocytes are critical for neuronal redox homeostasis providing them with cysteine needed for glutathione synthesis. In this study, we demonstrate that the astrocytic redox response signature provoked by amyloid beta (A-) is distinct from that of a general oxidant (tertiary-butylhydroperoxide [t-BuOOH]). Acute A- treatment increased cystathionine --synthase (CBS) levels and enhanced transsulfuration flux in contrast to repeated A- exposure, which decreased CBS and catalase protein levels. Although t-BuOOH also increased transsulfuration flux, CBS levels were unaffected. The net effect of A- treatment was an oxidative shift in the intracellular glutathione/glutathione disulfide redox potential in contrast to a reductive shift in response to peroxide. In the extracellular compartment, A-, but not t-BuOOH, enhanced cystine uptake and cysteine accumulation, and resulted in remodeling of the extracellular cysteine/cystine redox potential in the reductive direction. The redox changes elicited by A- but not peroxide were associated with enhanced DNA synthesis. CBS activity and protein levels tended to be lower in cerebellum from patients with Alzheimer's disease than in age-matched controls. Our study suggests that the alterations in astrocytic redox status could compromise the neuroprotective potential of astrocytes and may be a potential new target for therapeutic intervention in Alzheimer's disease. Antioxid. Redox Signal. 14, 2385-2397.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/90483/1/ars-2E2010-2E3681.pd

Heme-Thiolate perturbation in cystathionine β-Synthase by mercury compounds

Cystathionine β-synthase (CBS) is an enzyme involved in sulfur metabolism that catalyzes the pyridoxal phosphate-dependent condensation of homocysteine with serine or cysteine to form cystathionine and water or hydrogen sulfide (H2S), respectively. CBS possesses a b-type heme coordinated by histidine and cysteine. Fe(III)-CBS is inert toward exogenous ligands, while Fe(II)-CBS is reactive. Both Fe(III)- and Fe(II)-CBS are sensitive to mercury compounds. In this study, we describe the kinetics of the reactions with mercuric chloride (HgCl2) and p-chloromercuribenzoic acid. These reactions were multiphasic and resulted in five-coordinate CBS lacking thiolate ligation, with six-coordinate species as intermediates. Computational QM/MM studies supported the feasibility of formation of species in which the thiolate is proximal to both the iron ion and the mercury compound. The reactions of Fe(II)-CBS were faster than those of Fe(III)-CBS. The observed rate constants of the first phase increased hyperbolically with concentration of the mercury compounds, with limiting values of 0.3–0.4 s–1 for Fe(III)-CBS and 40 ± 4 s–1 for Fe(II)-CBS. The data were interpreted in terms of alternative models of conformational selection or induced fit. Exposure of Fe(III)-CBS to HgCl2 led to heme release and activity loss. Our study reveals the complexity of the interactions between mercury compounds and CBS

Heme-Thiolate Perturbation in Cystathionine β-Synthase by Mercury Compounds

Cystathionine β-synthase (CBS) is an enzyme involved in sulfur metabolism that catalyzes the pyridoxal phosphate-dependent condensation of homocysteine with serine or cysteine to form cystathionine and water or hydrogen sulfide (H2S), respectively. CBS possesses a b-type heme coordinated by histidine and cysteine. Fe(III)-CBS is inert toward exogenous ligands, while Fe(II)-CBS is reactive. Both Fe(III)-and Fe(II)-CBS are sensitive to mercury compounds. In this study, we describe the kinetics of the reactions with mercuric chloride (HgCl2) and p-chloromercuribenzoic acid. These reactions were multiphasic and resulted in five-coordinate CBS lacking thiolate ligation, with six-coordinate species as intermediates. Computational QM/MM studies supported the feasibility of formation of species in which the thiolate is proximal to both the iron ion and the mercury compound. The reactions of Fe(II)-CBS were faster than those of Fe(III)-CBS. The observed rate constants of the first phase increased hyperbolically with concentration of the mercury compounds, with limiting values of 0.3-0.4 s-1 for Fe(III)-CBS and 40 ± 4 s-1 for Fe(II)-CBS. The data were interpreted in terms of alternative models of conformational selection or induced fit. Exposure of Fe(III)-CBS to HgCl2 led to heme release and activity loss. Our study reveals the complexity of the interactions between mercury compounds and CBS.Fil: Benchoam, Dayana. Universidad de la Republica; UruguayFil: Cuevasanta, Ernesto. Universidad de la Republica; UruguayFil: Julió Plana, Laia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; ArgentinaFil: Capece, Luciana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; ArgentinaFil: Banerjee, Ruma. University of Michigan; Estados UnidosFil: Alvarez, Beatriz. Universidad de la República; Urugua

The Quantitative Significance of the Transsulfuration Enzymes for H2S Production in Murine Tissues

The enzymes of the transsulfuration pathway, cystathionine --synthase (CBS) and cystathionine --lyase (CSE), are important for the endogenous production of hydrogen sulfide (H2S), a gaseous signaling molecule. The relative contributions of CBS and CSE to H2S 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 H2S 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 H2S production is approximately equal for CBS and CSE, whereas in kidney and brain, CBS constitutes the major source of H2S, 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 H2S production by the transsulfuration pathway enzymes in liver. Antioxid. Redox Signal. 15, 363-372.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/90506/1/ars-2E2010-2E3781.pd

Differential Dependence on Cysteine from Transsulfuration versus Transport During T Cell Activation

The synthesis of glutathione, a major cellular antioxidant with a critical role in T cell proliferation, is limited by cysteine. In this study, we evaluated the contributions of the xC- cystine transporter and the transsulfuration pathway to cysteine provision for glutathione synthesis and antioxidant defense in naive versus activated T cells and in the immortalized T lymphocyte cell line, Jurkat. We show that the xC- transporter, although absent in naive T cells, is induced after activation, releasing T cells from their cysteine dependence on antigen-presenting cells. We also demonstrate the existence of an intact transsulfuration pathway in naive and activated T cells and in Jurkat cells. The flux through the transsulfuration pathway increases in primary but not in transformed T cells in response to oxidative challenge by peroxide. Inhibition of the transsulfuration pathway in both primary and transformed T cells decreases cell viability under oxidative-stress conditions. Antioxid. Redox Signal. 15, 39-47.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/90472/1/ars-2E2010-2E3496.pd