Biosynthesis of S-Methylcysteine in Radish Leaves¹

Abstract

Summary. Investigationl on the biosynthesis of S-methyl-L-cysteine in radish leaves has shown that it is formed by the methylation of cysteine. This conclusioii is based on: A) the relatively high recovery of radioactivity in methylcysteine sulfoxide after the administration of cysteine or methyl-labeled methionine to radish leaves; B) the nearly complete recovery of label from methyl-labeled methionine in the methyl group of methylcysteine sulfoxide; and C) the similarity in the ratio of tritium to 14C in methylcysteine sulfoxide and in its methyl group to this ratio in the methyl group of methionine given to radish leaves. Direct evidence for the synthesis of methylcysteine in radishes was obtained for the first time.Conclusive evidence against the formation of methylcysteine from serine and a thiomethyl group from methionine as stuggested for garlic was the more efficient incorporation of the methyl group of methionine as compared to the sulfur atom into methylcysteine suilfoxide.S-Methyl-L-cysteine has been shown to be a nattural constituent of kidney bean seed (19) and . It is the putative precursor of methylcysteine sulfoxide (1) in crucifers. Wolff, Black and Downey (22) showed that methylmercaptan and serine were combined in the presence of yeast enzymes to form S-methylcysteine (equation I). CH3SH + CH2OHCHNH.,COOH -CH,SCH2CHNHXCOOH + H2O (I).More recently, Sugii and coworkers (15) have observed a high recovery of radioactivity in methylcysteine sulfoxide after feeding sulfuir labeled methionine to a garlic plant. The interpretation of this finding was that methylmercaptan was formed from methionine in garlic as in the rat (4) and that the methylmercaptan was then combined with serine as in the yeast system to form methylcysteine.In this paper, evidence is presented which indicates that S-methylcysteine is formed in radishes by methylation of cysteine. Materials and MethodsMethyl labeled methionine and unlabeled S-adenosylmethionine were obtained from Cal Biochem2. Method of Administrationt of Labeled and Unlabeled MIaterials to Radish Leaves. In order to carry out the studies described in this paper, it was necessary to have a method of getting quantitative uptake of radioactive compounds in a relatively short time, while simultaneously stupplying unlabeled compoulnds and preventing the leaf from wilting. The latter 2 problems were solved by the tuse of a collar around the petiole (fig 1). The handling of leaves and administration of labeled and unlabeled compouinds were carried out by the following procedures.An intact radish plant was dtug uip and immersed in water. The leaves were excised with a razor blade under water, rinsed, and transferred to distilled water. An alternative method which was not as reliable but saves plants was also employed. In the latter case, the leaves were excised withott tunearthing the plant and the entire petiole immediately immersed in distilled water. Any leaves which remained turgid after 4 to 6 hours at room temperature were used.The petiole was cleanied of any small ...

In Vivo and In Vitro Studies on γ-Aminobutyric Acid Metabolism with the Radish Plant (Raphanus sativus, L.)

“…Radioactive solutions were administered to the leaves via the petiole (30). After the radioactive solution was absorbed (15 + 5 min), distilled water was supplied.…”

“…An aliquot of the aqueous phase was put through tandem columns (0.9 X 7 cm) of Dowex 50-X8 (200-400 mesh) in the hydrogen form and of Dowex 1-X8 (200-400 mesh) in the formate form at 2 to 4 C. After washing the columns with water, we separated the columns and amino acids were eluted from the Dowex 50 with 2 N NH4OH and 40 ml of water. A partial elution of organic acids from Dowex 1 was achieved with 60 ml of 1.5 N (30). Organic acids were separated by unidirectional, ascending paper chromatography (7) and detected by fluorescence after treatment with acridine (25).…”

“…Organic acids were separated by unidirectional, ascending paper chromatography (7) and detected by fluorescence after treatment with acridine (25). Radioactive spots were cut from chromatograms, eluted, and counted (30 (3). After purification of the diethylformylsuccinate by fractional distillation at 15 mm pressure, the purified product was hydrolyzed with 0.1 N HCI to succinic semialdehyde which was stored at 4 C and titrated with KOH to pH 6.0 just before use (8).…”

Anaerobic Accumulation of γ-Aminobutyric Acid and Alanine in Radish Leaves (Raphanus sativus, L.)

“…23) to trap GAB-"C formed from glutamate. Unlabeled GAB feeding was continued during uptake of acetate-"C (15 min) and a subsequent 15-min period.…”

“…An ascending run over 10 to 12 hr separated a-ketoglutarate hydrazone from other hydrazones. The hydrazone spots were cut up into scintillation vials, and hydrazones were eluted from the paper with 1 ml of ethanol followed by 4 ml of distilled water and counted by liquid scintillation (23).…”