No abstract

Individual rates of metabolism of the sulfur, methyl, and 4-carbon moieties of methionine were estimated in Lemna paucicostata Hegelm. 6746 The known major pathways for metabolism of methionine in the great majority of plant tissues not evolving large amounts of ethylene are summarized in Figure 1 (reactions a through k). Studies focused on the pathway for net methionine formation (Fig. 1, ---) have clearly demonstrated that this process occurs via transsulfuration ( 12), is subject to feedback control at cystathionine y-synthase (Fig. 1, reaction b) (14, 18), and that methionine accumulates predominantly in protein (12). By contrast, the quantitative significance and regulatory patterns of the other pathways of methionine metabolism illustrated in Figure 1

The metabolism of S-methylmethionine has been studied in cultures of plants of Lemna paucicostata and of cells of carrot (Daucus carota) and soybean (Glycine max). In each system, radiolabeled S-methylmethionine was rapidly formed from labeled L-methionine, consistent with the action of S-adenosyl-L-methionine:methionine S-methyltransferase, an enzyme which was demonstrated during these studies in Lemna homogenates. In Lemna plants and carrot cells radiolabel disappeared rapidly from Smethylmethionine during chase incubations in nonradioactive media. The results of pulse-chase experiments with Lemna strongly suggest that administered radiolabeled S-methylmethionine is metabolized initially to soluble methionine, then to the variety of compounds formed from soluble methionine. An enzyme catalyzing the transfer of a methyl group from S-methylmethionine to homocysteine to form methionine was demonstrated in homogenates of Lemna. The net result of these reactions, together with the hydrolysis of S-adenosylhomocysteine to homocysteine and adenosine, is to convert S-adenosylmethionine to methionine and adenosine. A physiological advantage is postulated for this sequence in that it provides the plant with a means of sustaining the pool of soluble methionine even when overshoot occurs in the conversion of soluble methionine to S-adenosylmethionine. The facts that the pool of soluble methionine is normally very small relative to the flux into S-adenosylmethionine and that the demand for the latter compound may change very markedly under different growth conditions make it plausible that such overshoot may occur unless the rate of synthesis of S-adenosylmethionine is regulated with exquisite precision. The metabolic cost of this apparent safeguard is the consumption of ATP. This S-methylmethionine cycle may well function in plants other than Lemna, but further substantiating evidence is neeeded. [6,25,26,35]). After administration of radiolabeled Met to a variety of intact plant preparations, substantial amounts of radiolabel have usually been found in SMM (4,18,19,21,23,29,34,37

ABSTRACICell-free extracts from Lemna and suspension cultured carrot (Daucus Carota L.) catalyze S-adenosylmethionine-dependent N-methylations of phosphoethanolamine, phosphomethylethanolamine, and phosphodimethylethanolamine; extracts of suspension cultured soybean (Glycine max), of phosphoethanolamine only. Material pelleted from each tissue between 15,000 and 100,000g catalyzes S-adenosylmethionine-dependent N-methylations of phosphatidylmethylethanolamine and phosphatidyldimethylethanolamine, but not phosphatidylethanolamine. Extracts from each tissue catalyze CTP-dependent cytidylyltransfers to each of the three methylated phosphoethanolamine derivatives, forming the corresponding CDP derivatives. Some of the properties of the activities investigated are reported. On the basis of in vivo labeling experiments, we have proposed (AH Datko, SH Mudd 1988 Plant Physiol 88: 854-861) differing pathways for phosphatidylcholine synthesis in which, after a common committing step, N-methylation of phosphoethanolamine, subsequent methylations occur in Lemna almost exclusively at the phospho-base level; in soybean, at the phosphatidyl-base level; and in carrot, at both levels. Thus, among the activities investigated, at least those required for the operation of the proposed pathways have been positively demonstrated. The extent to which the present results explain the differences between these pathways is discussed, and a speculation offered as to how these differences may have arisen phylogenetically.We have recently reported the results of studies of the kinetics of entry of the radiolabeled methyl group of L-methionine into the network of methylated derivatives of EA2 which lead ultimately to PtdCho. The patterns for each of the three plants studied, Lemna (19), soybean and carrot (4), were qualitatively different. The working models we suggested based on the results ' Reprint requests should be addressed to the authors at Building 36, Room 3D06, National Institute of Mental Health, Bethesda, MD 20892.2 Abbreviations: EA, ethanolamine; MEA, N-methylethanolamine; DMEA, N, N-dimethylethanolamine; Cho, choline. The phosphate esters of these compounds are designated by the prefix, P-(e.g. P-EA; P-Cho); the corresponding phosphatidyl, glycerylphospho-base, or CDP derivatives, by the prefixes, Ptd, GP-, or CDP-(e.g. PtdEA and PtdCho; GP-EA and GP-Cho; or CDP-EA and CDP-Cho). The methylations involved in PtdCho biosynthesis are all N-methylations. For brevity in the text Nmethylation has generally been replaced by methylation (unless special emphasis is desired). The of these studies include a committing step common to the three plants: namely an initial N-methylation of P-EA to form P-MEA. Subsequent methylations occur either virtually exclusively at the phospho-base level (Lemna) (19), at the phosphatidyl-base level (soybean), or at both of these levels (carrot) (4). These suggested pathways require the activity of certain phospho-base N-methyltransferases which have not hitherto been described, and of additional cytidy...

Regulation of enzymes of methionine bkosynthesis was investigated by measuring the specific activities of 0-p ot cys tathonine y-synthse, rim sulfhydrylase, and 0-acetylserine sulfhydrylase in Lemma paaeikesta Hegelm 6746 grown under various conditions. For cystathionine y-syntase, it was observed that (a) adding external methionlne (2 pM) decreased specific actvity to 15% of control, (b) Methionine is synthesized in higher plants from (a) the fourcarbon moiety of aspartic acid via O-phoshohomoserine, (b) the sulfur of inorganic sulfate via cysteine, and (c) a methyl group from N5-methyltetrahydrofolate (triglutamyl derivative) (14) (Fig. 1). Datko and Mudd (7) have reported upon several compounds (or combinations of compounds) each of which inhibits a specific and different step in this pathway and which therefore impairs the endogenous synthesis of methionine by Lemna paucicostata. Such inhibitors are especially valuable because they can be used as tools to investigate whether or how plants adapt to conditions of limiting methionine. Specifically, one can determine whether the activities of certain key enzymes in the methionine pathway 14 are sensitive to changes in the physiological concentration of methionine or one of its products. In this regard, cystathionine y-synthase is of primary interest. The reaction it catalyzes (reaction 1) O-phosphohomoserine + cysteine --cystathionine + Pi (1) is the central step joining branches of two separate pathways (Fig. 1). Both O-phosphohomoserine and cysteine are committed at this step to the formation of methionine rather than to their respective alternative fates of conversion to threonine or incorporation into protein and glutathione. Because regulation of biosynthetic pathways frequently occurs at committing steps, the cystathionine ysynthase reaction is a likely step at which to observe control by the end-product methionine.Another enzyme activity of interest is that which catalyzes direct sulfhiydration of O-phosphohomoserine (reaction 2): 0-phosphohomoserine + sulfide -. homocysteine + Pi (2) This reaction has been demonstrated in Lemna, Chlorella, and a number of other higher plants (8) and is a possible alternative to transsulfuration as a pathway for the formation of homocysteine (Fig. 1). However, studies of Lemna (15) and Chlorella (13) have failed to detect any homocysteine formation via this shunt and have shown that at least 90 to 95% of the homocysteine which is formed comes from cystathionine.