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Mass spectrometric analysis of L-cysteine metabolism: physiological role and fate of L-cysteine in the enteric protozoan parasite Entamoeba histolytica.

Jeelani G, Sato D, Soga T, Watanabe H, Nozaki T - MBio (2014)

Bottom Line: Furthermore, T4C and MT4C significantly enhanced trophozoite growth and reduced intracellular reactive oxygen species (ROS) levels when it was added to cultures, suggesting that 2-(R)-thiazolidine-4-carboxylic acids are involved in the defense against oxidative stress.We found that L-cysteine inside the cell rapidly reacts with aldehydes to form 2-(R)-thiazolidine-4-carboxylic acid.We showed that these 2-(R)-thiazolidine-4-carboxylic derivatives serve as an L-cysteine source, promote growth, and protect cells against oxidative stress by scavenging aldehydes and reducing the ROS level.

View Article: PubMed Central - PubMed

Affiliation: Department of Parasitology, National Institute of Infectious Diseases, Shinjuku, Tokyo, Japan.

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Metabolic decomposition of 2-(R)-thiazolidine-4-carboxylic acid in E. histolytica trophozoites. (A) Schematic representation of enzymatic degradation of 2-(R)-thiazolidine-4-carboxylic acids as previously proposed for Escherichia coli by Deutch (50). (B) Time course of T4C’s metabolism. The assay was performed as described in Materials and Methods. The means and SD from three independent experiments performed in triplicate are shown. (C) Relative intracellular concentrations of various unlabeled and isotope-labeled metabolites in E. histolytica trophozoites. Trophozoites were cultured in the presence of 8 mM stable-isotope-labeled l-cysteine (U-13C3, 15N) for 24 h. Then, stable-isotope-labeled l-cysteine-containing medium was replaced with l-cysteine-deprived BI-S-33 medium, and the trophozoites were harvested at 0, 0.5, 3, 9, and 24 h of cultivation. The bottom center plot is a magnified (at the y axis) plot of labeled l-alanine, shown at the bottom left. The x axis represents time in hours, whereas the y axis represents the relative peak areas per 1 × 106 cells. Metabolite data are presented as means ± SD from 3 biological replicates.
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fig4: Metabolic decomposition of 2-(R)-thiazolidine-4-carboxylic acid in E. histolytica trophozoites. (A) Schematic representation of enzymatic degradation of 2-(R)-thiazolidine-4-carboxylic acids as previously proposed for Escherichia coli by Deutch (50). (B) Time course of T4C’s metabolism. The assay was performed as described in Materials and Methods. The means and SD from three independent experiments performed in triplicate are shown. (C) Relative intracellular concentrations of various unlabeled and isotope-labeled metabolites in E. histolytica trophozoites. Trophozoites were cultured in the presence of 8 mM stable-isotope-labeled l-cysteine (U-13C3, 15N) for 24 h. Then, stable-isotope-labeled l-cysteine-containing medium was replaced with l-cysteine-deprived BI-S-33 medium, and the trophozoites were harvested at 0, 0.5, 3, 9, and 24 h of cultivation. The bottom center plot is a magnified (at the y axis) plot of labeled l-alanine, shown at the bottom left. The x axis represents time in hours, whereas the y axis represents the relative peak areas per 1 × 106 cells. Metabolite data are presented as means ± SD from 3 biological replicates.

Mentions: It has previously been shown that T4C is oxidized by E. coli (48), rat liver mitochondria (43), and barley (49). Oxidation of T4C by purified rat liver mitochondria yielded N-formyl-cysteine as a major end product (43). T4C is first converted to 2,3-thiazoline-4-carboxylate (Fig. 4A), 2,3-thiazolidine-4-carboxylate, and then N-acetyl (or formyl or propinyl)-l-cysteine by ring opening and finally gives rise to acetate (or formate or propionate) and l-cysteine by l-proline dehydrogenase (EC 1.5.99.8) (Fig. 4A) (50). Whether an additional enzyme is required to convert N-formyl-l-cysteine to formate and l-cysteine is still not clear (50). However, it was suggested that the hydrolysis of N-formyl-l-cysteine occurs nonenzymatically (50).


Mass spectrometric analysis of L-cysteine metabolism: physiological role and fate of L-cysteine in the enteric protozoan parasite Entamoeba histolytica.

Jeelani G, Sato D, Soga T, Watanabe H, Nozaki T - MBio (2014)

Metabolic decomposition of 2-(R)-thiazolidine-4-carboxylic acid in E. histolytica trophozoites. (A) Schematic representation of enzymatic degradation of 2-(R)-thiazolidine-4-carboxylic acids as previously proposed for Escherichia coli by Deutch (50). (B) Time course of T4C’s metabolism. The assay was performed as described in Materials and Methods. The means and SD from three independent experiments performed in triplicate are shown. (C) Relative intracellular concentrations of various unlabeled and isotope-labeled metabolites in E. histolytica trophozoites. Trophozoites were cultured in the presence of 8 mM stable-isotope-labeled l-cysteine (U-13C3, 15N) for 24 h. Then, stable-isotope-labeled l-cysteine-containing medium was replaced with l-cysteine-deprived BI-S-33 medium, and the trophozoites were harvested at 0, 0.5, 3, 9, and 24 h of cultivation. The bottom center plot is a magnified (at the y axis) plot of labeled l-alanine, shown at the bottom left. The x axis represents time in hours, whereas the y axis represents the relative peak areas per 1 × 106 cells. Metabolite data are presented as means ± SD from 3 biological replicates.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4222106&req=5

fig4: Metabolic decomposition of 2-(R)-thiazolidine-4-carboxylic acid in E. histolytica trophozoites. (A) Schematic representation of enzymatic degradation of 2-(R)-thiazolidine-4-carboxylic acids as previously proposed for Escherichia coli by Deutch (50). (B) Time course of T4C’s metabolism. The assay was performed as described in Materials and Methods. The means and SD from three independent experiments performed in triplicate are shown. (C) Relative intracellular concentrations of various unlabeled and isotope-labeled metabolites in E. histolytica trophozoites. Trophozoites were cultured in the presence of 8 mM stable-isotope-labeled l-cysteine (U-13C3, 15N) for 24 h. Then, stable-isotope-labeled l-cysteine-containing medium was replaced with l-cysteine-deprived BI-S-33 medium, and the trophozoites were harvested at 0, 0.5, 3, 9, and 24 h of cultivation. The bottom center plot is a magnified (at the y axis) plot of labeled l-alanine, shown at the bottom left. The x axis represents time in hours, whereas the y axis represents the relative peak areas per 1 × 106 cells. Metabolite data are presented as means ± SD from 3 biological replicates.
Mentions: It has previously been shown that T4C is oxidized by E. coli (48), rat liver mitochondria (43), and barley (49). Oxidation of T4C by purified rat liver mitochondria yielded N-formyl-cysteine as a major end product (43). T4C is first converted to 2,3-thiazoline-4-carboxylate (Fig. 4A), 2,3-thiazolidine-4-carboxylate, and then N-acetyl (or formyl or propinyl)-l-cysteine by ring opening and finally gives rise to acetate (or formate or propionate) and l-cysteine by l-proline dehydrogenase (EC 1.5.99.8) (Fig. 4A) (50). Whether an additional enzyme is required to convert N-formyl-l-cysteine to formate and l-cysteine is still not clear (50). However, it was suggested that the hydrolysis of N-formyl-l-cysteine occurs nonenzymatically (50).

Bottom Line: Furthermore, T4C and MT4C significantly enhanced trophozoite growth and reduced intracellular reactive oxygen species (ROS) levels when it was added to cultures, suggesting that 2-(R)-thiazolidine-4-carboxylic acids are involved in the defense against oxidative stress.We found that L-cysteine inside the cell rapidly reacts with aldehydes to form 2-(R)-thiazolidine-4-carboxylic acid.We showed that these 2-(R)-thiazolidine-4-carboxylic derivatives serve as an L-cysteine source, promote growth, and protect cells against oxidative stress by scavenging aldehydes and reducing the ROS level.

View Article: PubMed Central - PubMed

Affiliation: Department of Parasitology, National Institute of Infectious Diseases, Shinjuku, Tokyo, Japan.

Show MeSH
Related in: MedlinePlus