Hydrogen sulfide as a gasotransmitter

Abstract

Nitric oxide (NO) and carbon monoxide (CO) are well established as messenger molecules throughout the body, gasotransmitters, based on striking alterations in mice lacking the appropriate biosynthetic enzymes. Hydrogen sulfide (H(2)S) is even more chemically reactive, but until recently there was little definitive evidence for its physiologic formation. Cystathionine beta-synthase (EC 4.2.1.22), and cystathionine gamma-lyase (CSE; EC 4.4.1.1), also known as cystathionine, can generate H(2)S from cyst(e)ine. Very recent studies with mice lacking these enzymes have established that CSE is responsible for H(2)S formation in the periphery, while in the brain cystathionine beta-synthase is the biosynthetic enzyme. Endothelial-derived relaxing factor activity is reduced 80% in the mesenteric artery of mice with deletion of CSE, establishing H(2)S as a major physiologic endothelial-derived relaxing factor. H(2)S appears to signal predominantly by S-sulfhydrating cysteines in its target proteins, analogous to S-nitrosylation by NO. Whereas S-nitrosylation typically inhibits enzymes, S-sulfhydration activates them. S-nitrosylation basally affects 1-2% of its target proteins, while 10-25% of H(2)S target proteins are S-sulfhydrated. In summary, H(2)S appears to be a physiologic gasotransmitter of comparable importance to NO and carbon monoxide.

Keywords: EDHF; EDRF; GAPDH; KATP; S-sulfhydration; cystathionase; cystathionine β-synthase; cystathionine γ-lyase; hydrogen sulfide; hydropersulfide.

Fig. 1

(a) The classically described roles of CBS and CSE in sulfur metabolism. CBS condenses homocysteine with serine to generate the thiol ether cystathionine. CSE hydrolyzes cystathionine into cysteine, α-ketobutyrate and ammonia. (b) H₂S producing reactions catalyzed by CBS and CSE. CBS catalyzes the β-replacement reaction of cysteine (Cy–SH) with a variety of thiols (R–SH) to generate H₂S and the corresponding thiol ether (R–S–Cy). CSE catalyzes the β-disulfide elimination reaction of cystine (Cy–S–S–Cy), this is followed by a reaction with a variety of thiols, to generate H₂S and the corresponding disulfide (R–S–S–Cy).

Fig. 2

A model protein with some of the possible states of the cysteine thiol groups. From the top to the bottom, a free thiol (–SH), an S-nitrosylated thiol (–SNO), an S-sulfhydrated thiol (hydropersulfide) (–SSH) and a disulfide is shown.

Fig. 3

(a) Sulfhydration physiologically increases the catalytic activity of GAPDH. GAPDH activity assay in vitro at 37 °C with increasing NaHS levels. NaHS dose-dependently activates GAPDH. (b) DTT (1 mM) reverses GAPDH activation by 10 μM NaHS in vitro. All results are mean ± SEM. **P < 0.01. (c) Wild-type versus C150S mutant GAPDH activity in vitro with 15 μM NaHS. Wild-type (wt) but not C150S GAPDH is activated by NaHS. All results are mean ± SEM. **P < 0.01. (d) GAPDH activity with increasing substrate, glyceraldehyde 3-phosphate (G3P), levels with or without 10 μM NaHS. NaHS increases overall V_max without affecting K_m (~0.8 mM). (e) GAPDH activity in HEK293 cells transfected with nothing, or plasmids endcoding wild-type CSE, or catalytically inactive CSE and incubated with increasing concentrations of L-cysteine in the media for 1 h at 37 °C. GAPDH is activated in a dose-dependent manner in the presence of wild-type CSE. (f) In vivo GAPDH activity from wild-type versus CSE^−/− liver. CSE^−/− mice show decreased GAPDH activity (n = 6 animals). All results are mean ± SEM. *P < 0.05. Reproduced with permission from Mustafa et al. 2009.

Fig. 4

(a) Age-dependent hypertensive phenotype of CSE male knockout mice. The hypertensive phenotype peaks at 12 weeks of age with blood pressures 18 mm Hg greater than wild-type control mice (+/+). Blood pressure of heterozygotes (−/+) resembles that of homozygous knockouts (−/−) at early ages, but by 10 weeks of age the homozygous knockout mice display levels 10 mm Hg greater than the heterozygotes (n = 12). (b) Immunohistochemical localization of CSE to the endothelium of arterial blood vessels (black arrows) in wild-type mice. The signal is abolished in CSE knockout mice. (c) The contactile effects of phenylephrine on the mesenteric artery is the same in wild-type, heterozygous and homozygous knockout mice (n = 15). (d) Methacholine relaxation of the mesenteric artery is reduced by about 80% in homozygous CSE knockout vessels and about 50% in heterozygotes (n = 15). All results are means ± SEM. *P < 0.05 versus wild-type; #P < 0.05 versus heterozygote. Reproduced with permission from Yang et al. 2008.