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1.
Vitrified shoots regenerated from carnation petals (Dianthus caryophyllus L. cv. Scania) were recovered by culturing them in a medium containing 3.0 g/l Bacto-Peptone. Wax structures not found on vitrified shoots developed on the abaxial surface of leaves of recovered shoots and on those of normal leaves. Recovered shoots were rooted and successfully acclimatized while vitrified shoots could not survive the acclimatization process. The Bacto-Peptone solution was fractionated and the efficiency of each fraction for the recovery of vitrification was examined. Only basic, non high molecular fractions whose molecular weight was less than 10,000 were effective. 相似文献
2.
The effect of various culture conditions on the levels of ammonia assimilatory enzymes of Corynebacterium callunae 总被引:2,自引:0,他引:2
Haluk Ertan 《Archives of microbiology》1992,158(1):42-47
Corynebacterium callunae (NCIB 10338) grows faster on glutamate than ammonia when used as sole nitrogen sources. The levels of glutamine synthetase (GS; EC 6.3.1.2) and glutamate synthase (GOGAT; EC 1.4.1.13) of C. callunae were found to be influenced by the nitrogen source. Accordingly, the levels of GS and GOGAT activities were decreased markedly under conditions of ammonia excess and increased under low nitrogen conditions. In contrast, glutamate dehydrogenase (GDH; EC 1.4.1.4) activities were not significantly affected by the type or the concentration of the nitrogen source supplied. The carbon source in the growth medium could also affect GDH, GS and GOGAT levels. Of the carbon sources tested in the presence of 2 mM or 10 mM ammonium chloride as the nitrogen source pyruvate, acetate, fumarate and malate caused a decrease in the levels of all three enzymes as compared with glucose. GDH, GS and GOGAT levels were slightly influenced by aeration. Also, the enzyme levels varied with the growth phase. Methionine sulfoximine, an analogue of glutamine, markedly inhibited both the growth of C. callunae cells and the transferase activity of GS. The apparent K
m values of GDH for ammonia and glutamate were 17.2 mM and 69.1 mM, respectively. In the NADPH-dependent reaction of GOGAT, the apparent K
m values were 0.1 mM for -ketoglutarate and 0.22 mM for glutamine.Abbreviations GDH
glutamate dehydrogenase
- GS
glutamine synthetase
- GOGAT
glutamate synthase 相似文献
3.
Expression of bacterial gdhA (glutamate dehydrogenase; GDH; E.C. 1.4.1.1) genes in transgenic plants fundamentally alters plant growth, herbicide tolerance and metabolite profiles. The aim was to correlate gdhA expression with water potential during deficit using transgenic Nicotiana tabacum cv. ‘SR1’ (tobacco). Expression of GDH activity from the transgene was significantly correlated with high water potentials during deficit, both after 5 days of water deprivation (R = 0.91) and after 6 h after re-watering on day 6 (R = 0.72). GDH expression may provide a tool to alter the response of plants to periodic water deficit. 相似文献
4.
《Biotechnic & histochemistry》2013,88(1):32-43
An axillary proliferating clone of Prunus avium L. was subcul- tured every four weeks on solid MS medium with agar as the gelling agent. Vitrification (hyperhydricity) of shoots was induced in one four week cycle with the same medium except that agar was replaced by gel- rite. During culture on the vitrifying medium, the water content of the shoots progressively increased with a parallel decrease in chlorophyll content. Cytological differences between the leaves and stems of the vitrified and normal shoots were detected by light and electron (both transmission and scanning) microscopy. Leaves of vitrified shoots were characterized by lower number of chloroplasts in the palisade parenchyma and by a defective cuticle. The stems of vitrified shoots had a less developed and lignified xylem tissue, lacked sclerenchy- matic areas and showed hypertrophy of the cortical parenchyma. More intense vacuolar activity with evagina- tions of the chloroplast envelope into the vacuole was noted in cells of vitrified leaves. 相似文献
5.
No active uptake of ammonium was detected in Proteus vulgaris, Bacillus pasteurii, and Sporosarcina ureae, which indicates that these bacteria depend on the passive diffusion of ammonia across the cell membrane. In P. vulgaris the glutamine synthetase-glutamate synthase (GS-GOGAT) pathway and glutamate dehydrogenase (GDH) were present, and these enzymes exhibited high affinities for ammonium. In B. pasteurii and S. ureae, however, no GS activity was detected, and GOGAT activity was only present in S. ureae. GDH enzymes were present in these two organisms, but showed only low affinity for ammonium, with apparent K
m-values of 55.2 mM in B. pasteurii and 36.7 mM in S. ureae, repectively. These observations explain why P. vulgaris is able to grow at neutral pH and low ammonium concentration (2 mM), while B. pasteurii and S. ureae require high ammonium concentration (40 mM) and alkaline pH for growth.Non-standard abbreviations GS
glutamine synthetase
- GOGAT
glutamate synthase
- GDH
glutamate dehydrogenase
- GT
glutamyl transferase
- MA
methylammonium
- NB
nutrient broth
- YE
yeast extract
- NA
nocotinic acid 相似文献
6.
Cruz C Bio AF Domínguez-Valdivia MD Aparicio-Tejo PM Lamsfus C Martins-Loução MA 《Planta》2006,223(5):1068-1080
The wide range of plant responses to ammonium nutrition can be used to study the way ammonium interferes with plant metabolism
and to assess some characteristics related with ammonium tolerance by plants. In this work we investigated the hypothesis
of plant tolerance to ammonium being related with the plants’ capacity to maintain high levels of inorganic nitrogen assimilation
in the roots. Plants of several species (Spinacia oleracea L., Lycopersicon esculentum L., Lactuca sativa L., Pisum sativum L. and Lupinus albus L.) were grown in the presence of distinct concentrations (0.5, 1.5, 3 and 6 mM) of nitrate and ammonium. The relative contributions
of the activity of the key enzymes glutamine synthetase (GS; under light and dark conditions) and glutamate dehydrogenase
(GDH) were determined. The main plant organs of nitrogen assimilation (root or shoot) to plant tolerance to ammonium were
assessed. The results show that only plants that are able to maintain high levels of GS activity in the dark (either in leaves
or in roots) and high root GDH activities accumulate equal amounts of biomass independently of the nitrogen source available
to the root medium and thus are ammonium tolerant. Plant species with high GS activities in the dark coincide with those displaying
a high capacity for nitrogen metabolism in the roots. Therefore, the main location of nitrogen metabolism (shoots or roots)
and the levels of GS activity in the dark are an important strategy for plant ammonium tolerance. The relative contribution
of each of these parameters to species tolerance to ammonium is assessed. The efficient sequestration of ammonium in roots,
presumably in the vacuoles, is considered as an additional mechanism contributing to plant tolerance to ammonium nutrition. 相似文献
7.
The development of adventitious shoots of Picea abies was affected by the agar concentration in the culture medium. Increasing the agar concentration from 0.5 to 2.0% decreased vitrification, but at the same time reduced shoot growth and rooting potential. Slightly vitrified plantlets could become acclimatized to greenhouse conditions. The mesophyll of needles developed in vitro was interspersed with large air spaces; the lower the agar concentration, the larger the air spaces. After transfer to the greenhouse, the new needles from the acclimatized plantlets had an anatomy approaching that of plants growing in field. 相似文献
8.
An effective procedure for obtaining healthy shoots from nodal segments of Scrophularia yoshimurae is described. Nodal segments cultured on Murashige and Skoog's (MS) basal medium supplemented with 1.0 mg L(-1) benzyladenine (BA) and 0.2 mg L(-1) alpha-naphthaleneacetic acid (NAA) induced multiple shoots in conical flasks that differed in the way they were closed and sealed. Hermitically sealed culture vessels resulted in high hyperhydricity/vitrification. High concentrations of ethylene and CO2 were found to accumulate in these vessels. The hyperhydricity of the shoot cultures could be decreased by progressively ventilating the vessels. Exchange of gases was achieved by removing the Parafilm sealing without affecting sterility. This reduced the hyperhydricity rate and gave a good recovery of vitrified shoots, but resulted in decreased proliferation and a dehydration of proliferating nodal segments and the culture medium. The best number of normal shoots was observed when the parafilm was removed for gaseous exchange after four weeks of culture incubation. The results show that hyperhydricity in shoot cultures of S. yoshimurae could be prevented by sufficient gas exchange during culture. 相似文献
9.
Glutamate-dehydrogenase (GDH, EC 1.4.1.2) activity and isoenzyme patterns were investigated in Arabidopsis thaliana plantlets, and parallel studies were carried out on glutamine synthetase (GS, EC 6.3.1.2). Both NADH-GDH and NAD-GDH activities increased during plant development whereas GS activity declined. Leaves deprived of light showed a considerable enhancement of NADH-GDH activity. In roots, both GDH activities were induced by ammonia whereas in leaves nitrogen assimilation was less important. It was demonstrated that the increase in GDH activity was the result of de-novo protein synthesis. High nitrogen levels were first assimilated by NADH-GDH, while GS was actively involved in nitrogen metabolism only when the enzyme was stimulated by a supply of energy, generated by NAD-GDH or by feeding sucrose. When methionine sulfoximine, an inhibitor of GS, was added to the feeding solution, NADH-GDH activity remained unaffected in leaves whereas NAD-GDH was induced. In roots, however, there was a marked activation of GDH and no inactivation of GS. It was concluded that NADH-GDH was involved in the detoxification of high nitrogen levels while NAD-GDH was mainly responsible for the supply of energy to the cell during active assimilation. Glutamine synthetase, on the other hand was involved in the assimilation of physiological amounts of nitrogen. A study of the isoenzyme pattern of GDH indicated that a good correlation existed between the relative activity of the isoenzymes and the ratio of aminating to deaminating enzyme activities. The NADH-GDH activity corresponded to the more anodal isoenzymes while the NAD-GDH activity corresponded to the cathodal ones. The results indicate that the two genes involved in the formation of GDH control the expression of enzymes with different metabolic functions.Abbreviations GDH
glutamate dehydrogenase
- GS
glutamine synthetase
- MSO
methionine sulfoximine 相似文献
10.
Various enzymes involved in the initial metabolic pathway for ammonia assimilation by Methanobacterium ivanovii were examined. M. ivanovii showed significant activity of glutamine synthetase (GS). Glutamate synthase (GOGAT) and alanine dehydrogenase (ADH) were present, wheras, glutamate dehydrogenase (GDH) was not detected. When M. ivanovii was grown with different levels of NH
+
4
(i.e. 2, 20 or 200 mM), GS, GOGAT and ADH activities varied in response to NH
+
4
concentration. ADH was not detected at 2 mM level, but its activity increased with increased levels of NH
+
4
in the medium. Both GS and GOGAT activities increased with decreasing concentrations of NH
+
4
and were maximum when ammonia was limiting, suggesting that at low NH
+
4
levels, GS and GOGAT are responsible for ammonia assimilation and at higher NH
+
4
levels, ADH might play a role. Metabolic mutants of M. ivanovii that were auxotrophic for glutamine were obtained and analyzed for GS activity. Results indicate two categories of mutants: i) GS-deficient auxotrophic mutants and ii) GS-impaired auxotrophic mutants.Abbreviations GS
Glutamine synthetase
- GOGAT
glutamate synthase
- GDH
glutamate dehydrogenase
- ADH
alanine dehydrogenase 相似文献
11.
Effect of applied and internal hormones on vitrification and apical necrosis of different plants cultured in vitro 总被引:2,自引:0,他引:2
Natalia V. Kataeva Irena G. Alexandrova Raisa G. Butenko Elena V. Dragavtceva 《Plant Cell, Tissue and Organ Culture》1991,27(2):149-154
Development of vitrification and apical necrosis was followed in Camellia sinensis, Gerbera jamesonii, Malus domestica and hybrid Populus tremula x P. alba shoots cultured in vitro on Murashige & Skoog (MS) medium with different concentrations of growth regulators. High humidity in the culture vessels and excess of BA in the medium were found to be the major factors influencing vitrification. Lack of exogenous cytokinin in the medium during successive subcultures induced apical necrosis in poor-rooting species (Malus domestica, Camellia sinensis). The level of internal phytohormones (ABA, IAA, IPA, 2iP, Z, ZR) was determined in the apple shoots by means of ELISA. The content of internal cytokinins in the vitrified apple shoots was several times greater than in normal ones, which supports the hypothesis that excess of cytokinins, inducing rapid divisions of cells in meristems in the atmosphere with high humidity, is responsible for vitrification. Apical necrosis of the plantlets that appeared after cultivation on cytokinin-free medium is the result of deficiency in endogenous hormones in apple shoots and this being confirmed by analysis of endogenous hormones in apple shoots.Abbreviations BA
benzyladenine
- BHT
butylated hydroxy-toluene
- ABA
abscisic acid
- IAA
indole-3-acetic acid
- ELISA
enzyme-linked immunosorbent assay
- IPA
isopentenyladenosine
- 2iP
isopentenyladenine
- NAA
naphthyl-3-acetic acid
- TBS
trishydroxymethylaminomethane buffered saline
- TLC
thin layer chromatography
- Z
zeatin
- ZR
zeatin riboside 相似文献
12.
The activities of nitrate reductase (EC1.6.6.1), nitrite reductase (EC 1.6.6.4), glutamine synthetase (EC6.3.1.2), glutamate synthase (EC1.4.7.1) and NAD(P)H-dependent glutamate dehydrogenase (EC 1.4.1.3) were investigated in mesophyll and bundle sheath cells of maize leaves (Zea mays L.). Whereas nitrate and nitrite reductase appear to be restricted to the mesophyll and GDH to the bundle sheath, glutamine synthetase and glutamate synthase are active in both tissues.During the greening process, the activities of nitrate and nitrite reductase increased markedly, but glutamine synthetase, glutamate synthase and glutamate dehydrogenase changed little.Abbreviations BDH
British Drug Houses
- EDTA
Ethylene diamine tetra-acetic acid
- GDH
Glutamate dehydrogenase
- NADH
Nicotinamide-adenine dinucleotide reduced form
- NADPH
Nicotnamide-adenine dinucleotide phosphate reduced form
- PMSF
Phenylmethyl sulphonyl fluoride 相似文献
13.
Alanine dehydrogenase in Arthrobacter fluorescens exhibited an allosteric behaviour and two K
m values for ammonium were estimated. In batch cultures at different ammonium concentrations and in continuous culture following an NH4
+ pulse, the level of ADH activity seems to be regulated by the ammonium concentration, high activities being observed when extracellular ammonium was in excess. The response to the growth rate of an ammonium-limited chemostat culture of A. fluorescens seems to indicate that alanine dehydrogenase and glutamine synthetase activities were inversely related. High activities of glutamate oxaloacetate transaminase and glutamate pyruvate transaminase have been found in crude extract of ammonium-limited cultures. From the results obtained in batch cultures grown at different glucose concentrations and in carbon-limited chemostat culture it appeared that the limitation by glucose influenced alanine dehydrogenase activity negatively. No glutamate dehydrogenase activity and no glutamate synthase activity could be detected with either NADH or NADPH as coenzymes.Abbreviations ADH
alanine dehydrogenase
- GS
glutamine synthetase
- GDH
glutamate dehydrogenase
- GOGAT
glutamine oxoglutarate aminotransferase
- GOT
glutamate oxaloacetate transaminase
- GPT
glutamate pyruvate transaminase 相似文献
14.
Activities of nitrate reductase (NR; EC 1.6.6.1), nitrite reductase (NiR; EC 1.7.7.1), glutamine synthetase (GS; EC 6.3.1.2) and glutamate dehydrogenase (GDH; EC 1.4.1.3) were measured in cotyledons of sunflower (Helianthus annuus L. cv Peredovic) seedlings during germination and early growth under various external nitrogen sources. The presence of NO
3
-
in the medium promoted a gradual increase in the levels of NR and NiR activities during the first 7 d of germination. Neither NR nor NiR activities were increased in a nitrogen-free medium or in media with either NH
4
+
or urea as nitrogen sources. Moreover, the presence of NH
4
+
did not abolish the NO
3
-
-dependent appearance of NR and NiR activities. The increase of NR activity was impaired both by cycloheximide and chloramphenicol, which indicates that both cytoplasmic 80S and plastidic 70S ribosomes are involved in the synthesis of the NR molecule. By contrast, the appearance of NiR activity was only inhibited by cycloheximide, indicating that NiR seems to be exclusively synthesized on the cytoplasmic 80S ribosomes. Glutamine-synthetase activity was also strongly increased by external NO
3
-
but not by NH
4
+
or urea. The appearance of GS activity was more efficiently suppressed by cycloheximide than chloramphenicol. This indicates that GS is mostly synthesized in the cytoplasm. The cotyledons of the dry seed contain high levels of GDH activity which decline during germination independently of the presence or absence of a nitrogen source. Cycloheximide, but not chloramphenicol, greatly prevented the decrease of GDH activity.Abbreviations GDH
glutamate dehydrogenase
- GS
glutamine synthetase
- NiR
nitrite reductase
- NR
nitrate reductase 相似文献
15.
A 2–8-fold increase in the activity of glutamate dehydrogenase (GDH), accompanied by an alteration of the GDH isoenzyme pattern, was observed in detached pea shoots floated on tap water (preincubated shoots). Sugars supressed the process, whereas NH
+
4
and various metabolites as well as inhibitors of energy metabolism and protein synthesis were ineffective. The subcellular distribution pattern revealed evidence that the GDH isoenzymes are exclusively located in the mitochondrial matrix. The alterations in GDH activity occurring in preincubated shoots are restricted to the mitochondria.An experimental device suitable for studying the GDH function in isolated intact mitochondria has been established. Using [14C] citrate as the carbon source and hydrogen donor, the mitochondria synthesized considerable amounts of glutamate upon addition of NH
+
4
. The rates of glutamate formation in dependency of increasing NH
+
4
levels follow simple Michaelis-Menten kinetics. Half-saturation concentrations of NH
+
4
of 3.6±1.2 mM; 1.9±0.06 mM and 1.6±0.1 mM were calculated for the mitochondria isolated from pea shoots, roots, and preincubated shoots, respectively. The results are discussed in relation to the possible role of GDH in NH+/4 assimilation at elevated intracellular NH+/4 levels.Abbreviations GDH
Glutamate dehydrogenase
- MDH
malate dehydrogenase
- GOT
aspartate aminotransferase
- SDH
succinate dehydrogenase
- HEPES
4-(2-hydroxyethyl)-1-piperazineethan-sulfonic acid
- BSA
bovine serum albumin
- TPP
thiamine pyrophosphate
- DNP
2,4-dinitrophenol
- CCCP
carbonyl cyanide m-chlorophenylhydrazone
- DCPIP
2,6-dichlorophenolindophenol
Dedicated to Professor Dr. Maximilian Steiner on the occasion of his 75th birthday 相似文献
16.
The studies were performed on young triticale seedlings grown on a mineral medium containing 5 mM NO
3
−
as the nitrogen source, with the addition of 0.5 mM CdCl2. It was determined that cadmium ions accumulated mainly in the plant roots. Decreases in nitrate concentrations both in the
roots and shoots of seedlings, as well as decreases in soluble protein contents with simultaneous increases in endopeptidase
activity were also observed. Both in roots and shoots significant decreases in glutamic acid were noted. Toxic cadmium ion
accumulation in seedlings significantly modified activity of primary nitrogen assimilating enzymes, i.e. glutamine synthetase (GS, EC 6.3.1.2) and glutamate dehydrogenase (GDH, EC 1.4.1.2). There was a significant decrease in
GS activity both in roots and in shoots of the stressed plants, in comparison to plants grown without cadmium. In shoots of
the control plants and plants subjected to stress two GS isoforms were discovered: cytoplasmatic (GS1) and chloroplastic (GS2). Substantial decreases in total glutamine synthetase activity in green parts of seedlings, occurring under stress conditions,
result from dramatic decrease in GS2 activity (by 60 % in relation to the control plants); despite simultaneous increases in the cytoplasmatic isoform (GS1) activity by approx. 96 %.
Cadmium ions accumulating in roots and shoots of seedlings not only increased GDH activity, but also modified its coenzymatic
specificity. 相似文献
17.
Isabella Cacciari Daniela Lippi Tito Pietrosanti Walter Pietrosanti 《Archives of microbiology》1986,145(2):113-115
Ammonium assimilation was studied in a nitrogenfixing Arthrobacter strain grown in both batch and ammonium-limited continuous cultures. Arthrobacter sp. fluorescens grown in nitrogen-free medium or at low ammonium levels assimilated NH
4
+
via the glutamine synthetase/glutamate synthase pathway. When ammonium was in excess it was assimilated via the alanine dehydrogenase pathway. Very low levels of glutamate dehydrogenase were found, irrespective of growth conditions.Abbreviations GS
glutamine synthetase
- GOGAT
glutamine oxoglutarate aminotransferase
- GDH
glutamate dehydrogenase
- ADH
alanine dehydrogenase
- GOT
glutamate oxaloacetate transaminase
- GPT
glutamate pyruvate transaminase 相似文献
18.
An efficient in vitro method for mass propagation of a woody ornamentalIxora coccinea L. 总被引:1,自引:0,他引:1
Various factors that affect culture establishment, shoot growth, proliferation and rooting ofIxora coccinea L., a woody shrub, were studied. Stem cuttings (decapitated shoot, three nodes) were the most suitable explants for multiple-shoot proliferation, and when cultured on a woody plant medium (WPM) containing 2.5 M BA produced axillary shoots which branched repeatedly, yielding an average of 27 shoots per explant after 6 weeks in culture. Kinetin, 2-iP, zeatin and thidiazuron all induced multiple-shoot formation, but were less effective than BA. While the presence of IAA in the multiplication medium was detrimental to shoot proliferation, shoot growth was not affected by IAA. The production of large amounts of basal callus and vitrification of shoots were the major problems to be avoided in proliferating shoot cultures. Addition of TIBA to the multiplication medium markedly reduced basal callusing, while sealing the culture vessels with a fluorocarbon polymer (tetrafluoroethyleneperfluoroalkyl vinyl ether) film (Neoflon PFA film) almost completely eliminated vitrification. A reduction in the number of vitrified shoots was also achieved with AVG treatment. Following this protocol of using BA-supplemented WPM and Neoflon film, it would be possible to produce more than 100,000 plants from a single stem cutting in 1 year.Abbreviations
AVG
Aminoethoxyvinylglycine
-
BA
N6-benzyladenine
-
BM
basal medium
-
IAA
indole-3-acetic acid
-
IBA
indole-3-butyric acid
-
2-tiP
N6-(2-isopentenyl)adenine
-
KIN
kinetin
-
MS
Murashige and Skoog medium
-
NAA
-naphthaleneacetic acid
-
SRM
shoot regeneration medium
-
TDZ
thidiazuron
-
TIBA
2,3,5-triiodobenzoic acid
-
WPM
woody plant medium
-
ZEA
zeatin 相似文献
19.
N. S. Shekhawat T. S. Rathore R. P. Singh N. S. Deora S. R. Rao 《Plant Growth Regulation》1993,12(3):273-280
Genotype, age of tree, nature of explant and size (length and diameter), season of explant collection, explant position on medium, plant growth regulators and certain additives (ascorbic and citric acids, adenine sulphate, L-arginine, glutamine and ammonium citrate), incubation conditions, and subculturing period greatly influenced the in vitro clonal propagation of P. cineraria. The maximum number of 10–12 shoots were induced from the nodal shoot segment from pruned thorny adult trees on Murashige and Skoog's (MS) medium containing 0.1 mgl-1 indole- 3-acetic acid (IAA)+2.5 mgl-1 benzylaminopurine (BAP)+additives. Higher temperature (31+-2°C) and mixed (fluorescent and incandescent) light of 50 mol m-2 s-1 photon flux density for 12 h per day photoperiod favoured shoot induction and subsequent growth. Explants from thornless trees produced 6–8 shoots per explant on MS medium containing 0.1 mgl-1 IAA+5.0 mgl-1 BAP + additives. Nodal shoot segments obtained from root and stump sprouts produced multiple shoots. Root segments differentiated into multiple shoots on MS medium containing 0.5 mgl-1 indolebutyric acid (IBA)+2.5 mgl-1 BAP.Differentiated shoots multiplied best on MS medium containing 0.1 mgl-1 naphthalene acetic acid (NAA)+1.0 mgl-1 BAP + additives. To yield multiple shoots the original explant was transferred 6 times on fresh medium after harvesting the differentiated shoots. Shoots were rooted by pulsing with 100 mgl-1 IBA for 4 h and then culturing on hormone-free half strength MS medium. Initial dark incubation for 5 days at high temperature (33±2°C) was found essential for root induction from shoots which was 63% within two weeks. The rooted plantlets contained a consistent number of chromosomes (2n=28). It is suggested that the protocol developed could be useful for cloning of mature and tested trees of P. cineraria. 相似文献
20.
The relationships between the metabolism of malate, nitrogen assimilation and biosynthesis of amino acids in response to different nitrogen sources (nitrate and ammonium) have been examined in cotyledons of radish (Raphanus sativus L.). Measurements of the activities of some key enzymes and pulse-chase experiments with [14C]malate indicate the operation of an anaplerotic pathway for malate, which is involved in the synthesis of glutamine during increased ammonia assimilation. It is most likely that the tricarboxylicacid cycle is supplied with carbon through entry of malate, formed via the phosphoenolpyruvate (PEP)-carboxylation pathway, when 2-oxoglutarate leaves the cycle to serve as precursor for an increased synthesis of glutamine via glutamate. This might occur predominantly in the cytosol via the activity of the glutamine synthetase/glutamate synthase (GS/GOGAT) cycle, the NADH-dependent GOGAT being the rate-limiting activity.Abbreviations DTT
dithiothreitol
- EDTA
ethylenediamine-tetraacetic acid
- GDH
glutamate dehydrogenase
- GOGAT
glutamate synthase (glutamine: 2-oxoglutarate aminotransferase)
- GOT
aspartate aminotransferase (glutamate: oxaloacetate transaminase)
- GS
glutamine synthetase
- HPLC
high-performance liquid chromatography
- MCF
extraction medium of methanol: chloroform: 7M formic acid, 12:5:3, by vol.
- MDH
malate dehydrogenase
- MSO
L-methionine, sulfoximine
- PEPCase
phosphoenolpyruvate carboxylase
- TLC
thin-layer chromatography 相似文献