Inhibition of Glycine Oxidation by Carboxymethoxylamine, Methoxylamine, and Acethydrazide

Aminooxyacetate, a known inhibitor of transaminase reactions and glycine decarboxylase, promotes rapid depletion of the free pools of serine and aspartate in nitrate grown Lemna minor L. This compound markedly inhibits the methionine sulfoximine-induced accumulation of free ammonium ions and greatly restricts the methionine sulfoximine-induced depletion of amino acids such as glutamate, alanine, and asparagine. These results suggest that glutamate, alanine, and asparagine are normally catabolized to ammonia by transaminase-dependent pathways rather than via dehydrogenase or amidohydrolase reactions. Aminooxyacetate does not inhibit the methionine sulfoximine-induced irreversible deactivation of glutamine synthetase in vivo, indicating that these effects cannot be simply ascribed to inhibition of methionine sulfoximine uptake by aminooxyacetate. This transaminase inhibitor promotes extensive accumulation of several amino acids including valine, leucine, isoleucine, alanine, glycine, threonine, proline, phenylalanine, lysine, and tyrosine. Since the aminooxyacetate induced accumulations of valine, leucine, and isoleucine are not inhibited by the branched-chain amino acid biosynthesis inhibitor, chlorsulfuron, these amino acid accumulations most probably involve protein turnover. Depletions of soluble protein bound amino acids are shown to be approximately stoichiometric with the free amino acid pool accumulations induced by aminooxyacetate. Aminooxyacetate is demonstrated to inhibit the chlorsulfuron-induced accumulation of a-amino-n-butyrate in L. minor, supporting the notion that this amino acid is derived from transamination of 2-oxobutyrate.In a previous paper in this series (26), we have reported on the metabolic changes induced by the irreversible inhibitor of GS,2 MSO, in Lemna minor. It was proposed that glutamate, glutamine, asparagine, aspartate, alanine, and serine may be selectively catabolized to NH4+, either in the photorespiratory nitrogen cycle or by alternative routes (26). These alternative routes include dehydrogenase (e.g. GDH) and amidohydrolase (deamidase) (e.g. asparaginase) pathways (11,20,22,23,31).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. Although inhibition of asparagine catabolism by AOA would rule out an asparaginase pathway, this result would not discriminate between asparagine amino-N transfer to glycine or other amino acids as major pathways of asparagine catabolism (12,16,(33)(34)(35). Similarly, inhibition of alanine catabolism by AOA would not discriminate between alanine amino-N transfer directly to glycine via alanine :glyoxylate aminotransferase or alanine amino-N transfer to glutamate by glutamate:pyruvate aminotransferase (2, 9, 24). Nevertheless, the responses of L. minor to AOA...

Section: Resultssupporting

confidence: 71%

mentioning

confidence: 99%

Amino Acid Metabolism of Lemna minor L.

Brunk¹,

Rhodes²

1988

“…The lack of inhibition by urea suggests that the aminohydroxyl group of hydroxyurea and hydroxamates is critical for an effective binding to the enzyme. GR was also strongly inhibited (87%) by 5 mm AOA, an inhibitor of both glycine oxidation and transaminases (2, 10, 11, 23,26), and slightly affected (26% inhibition) by glycidate, an inhibitor of transaminase activities (16,32). Glycidate has also been reported to inhibit an NADPH-dependent activity of crude leaf GR from tobacco (32).…”

Section: Resultsmentioning

confidence: 99%

“…Hydroxamates have previously been reported to be 'specific' inhibitors of urease as well as some other metalloenzymes (8,14). Both GHA and AOA (as well as aminoacetonitrile which had no effect on GR activity) are structurally related to glycine and have been shown to inhibit glycine oxidation (2,7,9,11,17,18,23,28). AOA has also been reported to inhibit glyoxylatedependent aminotransferases (2,10,11,23,26) and an NADPHdependent activity of GR from Chiamydomonas (K, of 1 mM 126]).…”

Section: Discussionmentioning

confidence: 99%

Inhibition of Spinach Leaf NADPH(NADH)-Glyoxylate Reductase by Acetohydroxamate, Aminooxyacetate, and Glycidate

Kleczkowski

Randall²,

Blevins³

1987

Glycolate formation during photosynthesis is characteristic of organisms which fix CO2 via RuBP3 carboxylase/oxygenase, particularly when CO2 concentration in chloroplasts is limiting (25,31). An oxygenolytic cleavage of RuBP, catalyzed by the oxygenase activity, followed by hydrolysis of phosphoglycolate by phosphoglycolate phosphatase are considered to be the major, if not the only, reactions leading to glycolate formation in leaves (6,22,25). Spinach chloroplasts also contain low activities (3-6 itmol/h-mg Chl [19,20]) of an NADPH-dependent glyoxylate reductase (GR) capable of producing glycolate from glyoxylate.

“…4A). AOA is a nonspecific inhibitor of ACC synthase (10,17) and can cross the mitochondrial membrane (17). Moreover, the addition of AOA did not enhance the ACC synthase activity under low 02 conditions in the roots (Fig.…”

Section: Discussionmentioning

confidence: 99%

Effects of Low O₂ Root Stress on Ethylene Biosynthesis in Tomato Plants (Lycopersicon esculentum Mill cv Heinz 1350)

Wang¹,

Arteca²

1992

Low O2 conditions were obtained by flowing N2 through the solution in which the tomato plants (Lycopersicon esculentum Mill cv Heinz 1350) were growing. Time course experiments revealed that low O2 treatments stimulated 1-aminocyclopropane-1-carboxylate (ACC) synthase production in the roots and leaves. After the initiation of low 02 conditions, ACC synthase activity and ACC content in the roots increased and reached a peak after 12 and 20 hours, respectively. The conversion of ACC to ethylene in the roots was inhibited by low levels of 02, and ACC was apparently transported to the leaves where it was converted to ethylene. ACC synthase activity in the leaves was also stimulated by low 02 treatment to the roots, reaching a peak after 24 hours. ACC synthase levels were enhanced by cobalt chloride and aminooxyacetic acid (AOA), although they inhibited ethylene production. Cobalt chloride enhanced ACC synthase only in combination with IOW 02 conditions in the roots. Under aeration, AOA stimulated ACC synthase activity in both the roots and leaves. However, in combination with low 02 conditions, AOA caused a stimulation in ACC synthase activity in the leaves and no effect in the roots.Ethylene production is observed in all higher plants, where it is involved in numerous aspects of plant growth and development but most notably as a response to stress conditions (1).Waterlogging been reported to cause elevated ethylene levels and epinasty in the shoot regardless of whether the condition is imposed by waterlogging or by flushing with N2 (4,12,14). It has been shown that ethylene production by maize (Zea mays L.) roots exposed to 5 kPa oxygen was inhibited by AVG supplemented to the nutrient solution and this also reduced aerenchyma formation (13). This work provided evidence that the stimulation in ethylene production was probably due to an enhancement in ACC synthase and that ethylene was involved the promotion of aerenchyma in adventitious roots of maize. Cohen and Kende, utilizing an in vivo assay for this enzyme (8), showed that ACC synthase levels can be enhanced in deep-water rice internodes (Oryza sativa L.). Because they were unable to detect ACC synthase in homogenates of deepwater rice internodes, they utilized in vivo methods that assayed ACC accumulation in tissue under N2. Their conclusion was that the stimulation of ACC synthase activity by low 02 pressures was one of the first biochemical events leading to internodal growth in deep-water rice. The activity of ACC synthase in homogenates of vegetative tissues is often either much lower than expected or not measurable at all in vitro (8,20). To overcome this problem, we used the method described by Tsai et al. (18) to assay for ACC synthase. In the present study, we surveyed the changes in ACC synthase, ACC, and ethylene in whole tomato plants by flushing the roots with N2 and also evaluated the effects of ethylene biosynthesis inhibitors. MATERIALS AND METHODS Plant PreparationTomato (Lycopersicon esculentum Mill cv Heinz 1350) plants were grown f...