The Role of Transamination in the Synthesis of Homoserine in Peas

Abstract

ABSTRACIÌ ncubation of intact pea plants (Pisum sativum), or detached shoots, in continuous light caused a substantial increase (up to 4-fold in 2 days) in levels of homoserine. Amino acids supplied to leaves in the transpiration stream enhanced the accumulation, with glutamate, aspartate, and asparagine causing similar enhancement. Aminooxyacetate (AOA), a transamination inhibitor, at 1 millimolar prevented the accumulation. '4C-labeling experiments showed that succinate was a good source of carbon for homoserine synthesis; carbon from aspartate or asparagine was also incorporated into homoserine. For each precursor, the transfer of label was prevented by AOA. The keto acid analog of homoserine was rapidly transaminated in leaves to give homoserine. The results suggest that accumulating homoserine is synthesised by transamination rather than being derived from aspartate via the aspartate kinase/homoserine dehydrogenase pathway. The latter pathway was shown to be operating in the chloroplasts, and was sensitive to threonine (but was not inhibited by AOA), suggesting that this path has a role in synthesis of aspartatederived amino acids but is not involved in the accumulation of excess homoserine in the pea.Although homoserine is undetectable in most plants, it is thought to be an essential intermediate in the synthesis of threonine and methionine, and is considered to be derived from aspartate by the action of aspartate kinase, aspartyl semi-aldehyde dehydrogenase and homoserine dehydrogenase (9). In the leaf, this activity is located in the chloroplasts, including those of pea (1 1, 18). The pathway appears to be tightly regulated by threonine and lysine (9). In some plants such as the pea and Lathyrus (5), homoserine is present at quite high levels. It is particularly prominent during germination (4, 6), and may have a role as a transport compound. Kinetic properties of enzymes of homoserine metabolism in the pea and nonaccumulating species have been compared, but results did not provide a convincing explanation for homoserine accumulation (17).In our investigations of asparagine metabolism in the pea, several observations have suggested that homoserine was not directly derived from aspartate. There was a flow of nitrogen from the amino group of asparagine to homoserine, and this transfer was sensitive to the transamination inhibitor aminooxyacetate (15). In double labeling experiments, aspartate received considerable amounts of carbon and nitrogen from asparagine, whereas homoserine received mainly nitrogen (15). The deamidation inhibitor 5-diazo-4-oxo L-norvaline drastically inhibited production ofaspartate from asparagine, but had much less effect on transfer of nitrogen to homoserine (16). '5N-labeled alanine, asparagine, aspartate and glutamate were all effective as a source of nitrogen for homoserine synthesis (TC Ta, KW Joy, unpublished data). These results suggest a transfer of nitrogen to homoserine by transamination. Labeling experiments by other workers are also consistent with derivation of carbon,...

Lipogenesis and Redox Balance in Nitrogen-Fixing Pea Bacteroids

“…2). Levels of both were also high in the nodule cytosolic fraction relative to bacteroids (33-and 11-fold increases; Table S4 in the supplemental material), in accordance with previous observations (41,42) and consistent with their known plant origin. Asparagine is made in the plant cytosol as the primary nitrogen export product from nodules (40).…”

Downy Mildew Resistance in Arabidopsis by Mutation of HOMOSERINE KINASE

“…As homoserine is abundant in the phloem sap of these plants, it is thought to be a transport molecule for nitrogen and carbon allocation. Interestingly, in pea, an alternative biosynthetic route exists, by transamination of a keto acid precursor, to produce high amounts of homoserine (Joy and Prabha, 1986). Aminooxyacetate, a transamination inhibitor, can inhibit this reaction, whereas homoserine synthesis through the Asp pathway remains unaffected by this drug.…”

Amino Acid Metabolism of Lemna minor L.

“…We have reasoned that if such alternative pathways of amino acid catabolism to NH4+ are in operation, then these should not be inhibited by the transaminase inhibitor, AOA (9,15,16,36 (20,23,31). In either case, asparagine catabolism should be inhibited by AOA since the initial step in asparagine catabolism entails transamination.…”