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Identification of H2S3 and H2S produced by 3-mercaptopyruvate sulfurtransferase in the brain.

Kimura Y, Toyofuku Y, Koike S, Shibuya N, Nagahara N, Lefer D, Ogasawara Y, Kimura H - Sci Rep (2015)

Bottom Line: We recently found H2Sn in the brain.Purified recombinant 3MST and lysates of COS cells expressing 3MST produced H2S3 from 3 MP, while those expressing defective 3MST mutants did not.The present study provides new insights into the physiology of H2S3 and H2S, as well as novel therapeutic targets for diseases in which these molecules are involved.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Pharmacology, National Institute of Neuroscience, National Center of Neurology and Psychiatry, 4-1-1 Ogawahigashi, Kodaira, Tokyo 187-8502, Japan.

ABSTRACT
Hydrogen polysulfides (H2Sn) have a higher number of sulfane sulfur atoms than hydrogen sulfide (H2S), which has various physiological roles. We recently found H2Sn in the brain. H2Sn induced some responses previously attributed to H2S but with much greater potency than H2S. However, the number of sulfur atoms in H2Sn and its producing enzyme were unknown. Here, we detected H2S3 and H2S, which were produced from 3-mercaptopyruvate (3 MP) by 3-mercaptopyruvate sulfurtransferase (3MST), in the brain. High performance liquid chromatography with fluorescence detection (LC-FL) and tandem mass spectrometry (LC-MS/MS) analyses showed that H2S3 and H2S were produced from 3 MP in the brain cells of wild-type mice but not 3MST knockout (3MST-KO) mice. Purified recombinant 3MST and lysates of COS cells expressing 3MST produced H2S3 from 3 MP, while those expressing defective 3MST mutants did not. H2S3 was localized in the cytosol of cells. H2S3 was also produced from H2S by 3MST and rhodanese. H2S2 was identified as a minor H2Sn, and 3 MP did not affect the H2S5 level. The present study provides new insights into the physiology of H2S3 and H2S, as well as novel therapeutic targets for diseases in which these molecules are involved.

No MeSH data available.


Related in: MedlinePlus

H2S3 and H2S production depends on 3MST catalytic site.(a,b) The kinetics of H2S3 generation by 3MST in the presence of 3 MP. The production of H2S3 from 3 MP by purified 3MST (a), and its Lineweaver-Burk plot (b). (c,d) Production of H2S3 (c) and H2S (d) from 3 MP by various defective 3MST mutants. Lysates of COS cells expressing 3MST mutants as a source of the enzymes were incubated with 100 μM 3 MP for 15 min, and the H2S3 (c) and H2S (d) produced were measured. Control: lysates of cells transfected with an empty vector. #p < 0.05, ** and ##p < 0.01. **: the comparison with a value of an empty vector for (c,d). All data represent the mean ± standard error of the mean (SEM) of at least three experiments.
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f3: H2S3 and H2S production depends on 3MST catalytic site.(a,b) The kinetics of H2S3 generation by 3MST in the presence of 3 MP. The production of H2S3 from 3 MP by purified 3MST (a), and its Lineweaver-Burk plot (b). (c,d) Production of H2S3 (c) and H2S (d) from 3 MP by various defective 3MST mutants. Lysates of COS cells expressing 3MST mutants as a source of the enzymes were incubated with 100 μM 3 MP for 15 min, and the H2S3 (c) and H2S (d) produced were measured. Control: lysates of cells transfected with an empty vector. #p < 0.05, ** and ##p < 0.01. **: the comparison with a value of an empty vector for (c,d). All data represent the mean ± standard error of the mean (SEM) of at least three experiments.

Mentions: We examined the affinity of 3MST for 3 MP using purified recombinant 3MST produced by bacteria (Fig. 3a,b). 3MST produced H2S3 from 3 MP in a concentration-dependent manner up to 1.5 mM. The Km value of 3MST for 3 MP was 4.5 ± 1.5 mM.


Identification of H2S3 and H2S produced by 3-mercaptopyruvate sulfurtransferase in the brain.

Kimura Y, Toyofuku Y, Koike S, Shibuya N, Nagahara N, Lefer D, Ogasawara Y, Kimura H - Sci Rep (2015)

H2S3 and H2S production depends on 3MST catalytic site.(a,b) The kinetics of H2S3 generation by 3MST in the presence of 3 MP. The production of H2S3 from 3 MP by purified 3MST (a), and its Lineweaver-Burk plot (b). (c,d) Production of H2S3 (c) and H2S (d) from 3 MP by various defective 3MST mutants. Lysates of COS cells expressing 3MST mutants as a source of the enzymes were incubated with 100 μM 3 MP for 15 min, and the H2S3 (c) and H2S (d) produced were measured. Control: lysates of cells transfected with an empty vector. #p < 0.05, ** and ##p < 0.01. **: the comparison with a value of an empty vector for (c,d). All data represent the mean ± standard error of the mean (SEM) of at least three experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4594004&req=5

f3: H2S3 and H2S production depends on 3MST catalytic site.(a,b) The kinetics of H2S3 generation by 3MST in the presence of 3 MP. The production of H2S3 from 3 MP by purified 3MST (a), and its Lineweaver-Burk plot (b). (c,d) Production of H2S3 (c) and H2S (d) from 3 MP by various defective 3MST mutants. Lysates of COS cells expressing 3MST mutants as a source of the enzymes were incubated with 100 μM 3 MP for 15 min, and the H2S3 (c) and H2S (d) produced were measured. Control: lysates of cells transfected with an empty vector. #p < 0.05, ** and ##p < 0.01. **: the comparison with a value of an empty vector for (c,d). All data represent the mean ± standard error of the mean (SEM) of at least three experiments.
Mentions: We examined the affinity of 3MST for 3 MP using purified recombinant 3MST produced by bacteria (Fig. 3a,b). 3MST produced H2S3 from 3 MP in a concentration-dependent manner up to 1.5 mM. The Km value of 3MST for 3 MP was 4.5 ± 1.5 mM.

Bottom Line: We recently found H2Sn in the brain.Purified recombinant 3MST and lysates of COS cells expressing 3MST produced H2S3 from 3 MP, while those expressing defective 3MST mutants did not.The present study provides new insights into the physiology of H2S3 and H2S, as well as novel therapeutic targets for diseases in which these molecules are involved.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Pharmacology, National Institute of Neuroscience, National Center of Neurology and Psychiatry, 4-1-1 Ogawahigashi, Kodaira, Tokyo 187-8502, Japan.

ABSTRACT
Hydrogen polysulfides (H2Sn) have a higher number of sulfane sulfur atoms than hydrogen sulfide (H2S), which has various physiological roles. We recently found H2Sn in the brain. H2Sn induced some responses previously attributed to H2S but with much greater potency than H2S. However, the number of sulfur atoms in H2Sn and its producing enzyme were unknown. Here, we detected H2S3 and H2S, which were produced from 3-mercaptopyruvate (3 MP) by 3-mercaptopyruvate sulfurtransferase (3MST), in the brain. High performance liquid chromatography with fluorescence detection (LC-FL) and tandem mass spectrometry (LC-MS/MS) analyses showed that H2S3 and H2S were produced from 3 MP in the brain cells of wild-type mice but not 3MST knockout (3MST-KO) mice. Purified recombinant 3MST and lysates of COS cells expressing 3MST produced H2S3 from 3 MP, while those expressing defective 3MST mutants did not. H2S3 was localized in the cytosol of cells. H2S3 was also produced from H2S by 3MST and rhodanese. H2S2 was identified as a minor H2Sn, and 3 MP did not affect the H2S5 level. The present study provides new insights into the physiology of H2S3 and H2S, as well as novel therapeutic targets for diseases in which these molecules are involved.

No MeSH data available.


Related in: MedlinePlus