A Novel Enhancing Mechanism for Hydrogen Sulfide-producing Activity of Cystathionine β-Synthase

Abstract

H 2 S is produced from cysteine by cystathionine ␤-synthase (CBS) in the brain and functions as a neuromodulator. Although the production of H 2 S is regulated by Ca 2؉ and calmodulin in response to neuronal excitation, little is known about the molecular mechanism for the regulation in CBS activity. Here we show that four cysteine residues of CBS are involved in the regulation of its activity in the presence of Ca 2؉ and calmodulin. Sodium nitroprusside (SNP), a modifying agent for cysteine residues, enhances CBS activity, whereas N-ethylmaleimide, an alkylating agent for cysteine residues, completely abolished the effect of SNP. Site-directed mutagenesis of the 13 cysteine residues of CBS identified four cysteine residues that are involved in the regulation of CBS activity by SNP, and two of the four residues are involved in the regulation of the basal CBS activity. The enhancement of CBS activity by SNP is independent of nitric oxide production. In the presence of Staphylococcus aureus ␣-hemolysin, which permeabilizes the cell membrane, exogenously applied SNP enhances the activity of CBS in intact cells. The present study demonstrates a novel mechanism for the regulation of CBS activity and provides a possible therapeutic application of SNP for the diseases in which CBS activity is deficient.Relatively high endogenous levels of H 2 S, which is well known toxic gas, have been found in the brains of rats, humans and bovine (1-3), suggesting that H 2 S may have a physiological function. Endogenous H 2 S in the brain is produced from L-cysteine by the pyridoxal 5Ј-phosphate-dependent enzyme, cystathionine ␤-synthase (CBS) 1 (4 -7). CBS is expressed in the brain, and a CBS activator, S-adenosylmethionine (AdoMet), enhances H 2 S production (6). These observations, together with our recent finding that endogenous H 2 S is under detectable levels in the brains of CBS knock-out mice, indicate that CBS is a major H 2 S-producing enzyme in the brain (7).The production of H 2 S is regulated by a Ca 2ϩ -and calmodulin-mediated pathway that is activated in response to neuronal excitation (7). Physiological concentrations of H 2 S specifically potentiate the activity of the N-methyl-D-aspartate (NMDA) receptor, and hippocampal long term potentiation is altered in CBS knock-out mice (6, 7). H 2 S can regulate the release of corticotropin-releasing hormone from the hypothalamus (8). In addition to the function in the brain H 2 S relaxes smooth muscle in synergy with nitric oxide (NO) by activating ATP-dependent potassium channels (9, 10). Based upon these observations it has been proposed that H 2 S may function as a neuromodulator, as well as a smooth muscle relaxant (6, 9).Sodium nitroprusside (SNP), which is well known as an NO donor, activates guanylyl cyclase and relaxes smooth muscle (11,12). SNP has also been used to detect cysteine, cystine, acetophenone, and sulfite in a reaction known as the Legal and Boedeker reaction, as well as secondary amines (13-15). The reactivity of SNP on cysteine residues modifies biolog...

Overexpression of Cystathionine γ-Lyase Suppresses Detrimental Effects of Spinocerebellar Ataxia Type 3

“…There is evidence that myeloid zinc finger 1 and specificity protein 1 transcription factor affect the transcription of CSE (66). Furthermore, studies suggest that CSE can be upregulated by bacterial endotoxin (66,67) and by nitric oxide (68). S-adenosylmethionine and pyridoxal-5′-phosphate stimulate CSE activity to increase H 2 S production (69,70).…”

Redox Biochemistry of Hydrogen Sulfide

“…Mechanisms for transient up-regulation of H 2 S production, as is expected for a signaling molecule, remain to be elucidated. Neither nitric oxide (21) nor calmodulin (22,23), reported to modulate the activities of CSE and CBS, appears to do so (13,15). 3 Peroxynitrite (24) and carbon monoxide (25) inhibit human CBS.…”

Commonalities in Metabolism of Arsenicals

“…[35,36] Formation in vitro or in vivo of thioylated metabolites by rat liver [37,38] may require enzymatic generation of hydrogen sulfide. [39] Understanding formation of arsinothioyls and their biological activities will provide more information about roles of metabolism in the actions of arsenicals as toxins and carcinogens.…”