Serine Transhydroxymethylase of Cauliflower (<i>Brassica oleracea</i> var. <i>botrytis</i> L.): Partial Purification and Properties

Mazelis, Mendel; Liu, Elizabeth S.

doi:10.1104/pp.42.12.1763

Cited by 21 publications

(5 citation statements)

References 21 publications

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“…Although we were not able to ascertain the location of the residual seine transhydroxymethylase activity, it was clearly extramitochondrial. The existence of two isozymes of serine transhydroxymethylase in leaves has been suggested previously on the basis of cell-fractionation studies (10,23,30) and enzymological analysis (17). Also, mutants of yeast have been described which lack mitochondrial serine transhydroxymethylase but contain a second, distinguishable enzyme which is controlled or specified by a separate gene (33 Gas-exchange Analysis.…”

Section: Methodsmentioning

confidence: 93%

Photorespiration-deficient Mutants of Arabidopsis thaliana Lacking Mitochondrial Serine Transhydroxymethylase Activity

1981

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Three allelic mutants of Arabidopsis thalma which lack mitochondrial serine transhydroxymethylase activity due to a recessive nuclear mutation have been characterized. The mutants were shown to be deficient both in glycine decarboxylation and in the conversion of glycine to serine. Glycine accumulated as an end product of photosynthesis in the mutants, largely at the expense of serine, starch, and sucrose formation. The mutants photorespired CO2 at low rates in the light, but this evolution of photorespiratory CO2 was abolished by provision of exogenous NH3. Exogenous NH3 was required by the mutants for continued synthesis of glycine under photorespiratory conditions. These and related results with wild-type Arabidopsis suggested that glycine decarboxylation is the sole site of photorespiratory CO2 release in wild-type plants but that depletion of the amino donors required for glyoxylate amination may lead to CO2 release from direct decarboxylation of glyoxylate. Photosynthetic CO2 fixation was inhibited in the mutants under atmospheric conditions which promote photorespiration but could be partialiy restored by exogenous NH3. The magnitude of the NH3 stimulation of photosynthesis indicated that the increase was due to the suppression of glyoxylate decarboxylation. The normal growth of the mutants under nonphotorespiratory atmospheric conditions indicates that mitochondrial serine transhydroxymethylase is not required in C3 plants for any function unrelated to photorespiration. (3,12,18,31), by comparing the specific activity of photorespiratory CO2 to that of photorespiratory cycle intermediates (16), and by demonstrating, in the presence of chemical inhibitors, that the glycine to serine conversion is tightly coupled to photorespiratory CO2 evolution (11,14,15). These experiments cannot unequivocally exclude the possibility that there are other sites of photorespiratory CO2 release (4). Inhibitor studies are also plagued by nonspecific effects which preclude an appraisal of the long-term physiological effects of blocking the decarboxylation reaction.It has been suggested (4, 7, 8, 15, 34) that, under certain circumstances, glyoxylate, the immediate precursor of photorespiratory glycine, may undergo decarboxylation to produce CO2 and formate. This reaction has been demonstrated in vitro with isolated peroxisomes (7), but evidence substantiating the occurrence of this reaction in vivo is totally lacking. Here, we provide evidence that, under normal physiological conditions, glycine decarboxylation is the sole site of photorespiratory CO2 release but that, under conditions of severe amino depletion, photorespiratory CO2 may arise from direct decarboxylation of glyoxylate. This conclusion is based upon an analysis of mutants of Arabidopsis that lack mitochondrial serine transhydroxymethylase activity and are, therefore, defective in both glycine decarboxylation and the glycine to serine conversion. Photorespiratory CO2 is the product ofa complex mitochondrial reaction in which glycine is converted to stoi...

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Section: Methodsmentioning

confidence: 93%

Photorespiration-deficient Mutants of Arabidopsis thaliana Lacking Mitochondrial Serine Transhydroxymethylase Activity

1981

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“…this enzyme from a plant source. Previous attempts to isolate this enzyme had resulted in only partial purification due to the reported instability of this enzyme from cauliflower buds (13) and tobacco roots (15). The use of pyridoxal 5'-phosphate, 2-mercaptoethanol, and EDTA prevented the inactivation of this enzyme during purification (16).…”

Section: Discussionmentioning

confidence: 99%

“…On the other hand, there is some doubt on the requirement of pyridoxal 5'-phosphate for catalysis for some of the plant serine hydroxymethyltransferases (13). As most of the early work was carried out with partially purified preparations, it was difficult to draw unequivocal conclusions.…”

Section: Discussionmentioning

confidence: 99%

Purification and Regulatory Properties of Mung Bean (Vigna Radiata L.) Serine Hydroxymethyltransferase

Rao¹,

Rao²

1982

Plant Physiol.

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Serine hydroxymethyltransferase, the rirst enzyme in the pathway for interconversion of C1 fragments, was purified to homogeneity for the first time from any plant source. The enzyme from 72-h mung bean (Vigna radiata L.) seedlings was isolated using Blue Sepharose CL-6B and folate-AH-Sepharose4B affinity matrices and had the highest specific activity (1. The sigmoid saturation pattern of H4folate (nH = 2.0), one of the substrates, the abolition of sigmoidicity by NADH, an allosteric positive effector (nH = 1.0) and the increase in sigmoidicity by NAD+ and adenine nucleotides, negative allosteric effectors (nH = 2.4) clearly established that this key enzyme in the folate metabolism was an allosteric protein. Further support for this conclusion were the observations that (a) seine saturation exhibited an intermediary plateau region; (b) partial inhibition by methotrexate, aminopterin, O-phosphoserine, DL-a-methylserine and DL-O-methylserine; (c) subunit nature of the enzyme; and (d) decrease in the nH value from 2.0 for Hfolate to 1.5 in presence of L-serine.These results highlight the regulatory nature of mung bean serine hydroxymethyltransferase and its possible involvement in the modulation of the interconversion of folate coenzymes.The major storage form of folate coenzymes in plants is N5-methyltetrahydrofolate (21). Although this compound is present in abundant quantities the metabolism of C1 compounds and the 'This investigation was supported from a research

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“…Relatively few investigations have been described on the isolation and characterization of SHMT from plants. Although SHMT from spinach (28), maize (13), soybean nodules (15), tobacco (16), castor endosperm (5), carrot, pea leaves, wheat (4), cauliflower (14), and Lemna minor (27) have been studied in some detail, the requirement of PLP for catalysis has not been clearly established. Earlier work from our laboratory on the homogeneous mung bean SHMT (18) had suggested that this enzyme may not require PLP for catalysis.…”

mentioning

confidence: 99%

Serine Hydroxymethyltransferase from Mung Bean (Vigna radiata) Is Not a Pyridoxal-5′-Phosphate-Dependent Enzyme

Sukanya

Vijaya²,

Savithri³

et al. 1991

Plant Physiol.

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Serine hydroxymethyltransferase from mammalian and bacterial sources is a pyridoxal-5'-phosphate-containing enzyme, but the requirement of pyridoxal-5'-phosphate for the activity of the enzyme from plant sources is not clear. The specific activity of senne hydroxymethyltransferase isolated from mung bean (Vigna radiata) seedlings in the presence and absence of pyridoxal-5'-phosphate was comparable at every step of the purification procedure. The mung bean enzyme did not show the characteristic visible absorbance spectrum of a pyridoxal-5'-phosphate protein. Unlike the enzymes from sheep, monkey, and human liver, which were converted to the apoenzyme upon treatment with L-cysteine and dialysis, the mung bean enzyme similarly treated was fully active. Additional evidence in support of the suggestion that pyridoxal-5'-phosphate may not be required for the mung bean enzyme was the observation that pencillamine, a well-known inhibitor of pyridoxal-5'-phosphate enzymes, did not perturb the enzyme spectrum or inhibit the activity of mung bean serine hydroxymethyltransferase. The sheep liver enzyme upon interaction with O-amino-D-serine gave a fluorescence spectrum with an emission maximum at 455 nm when excited at 360 nm. A 100-fold higher concentration of mung bean enzyme-O-amino-Dseine complex did not yield a fluorescence spectrum. The following observations suggest that pyridoxal-5'-phosphate normally present as a coenzyme in serine hydroxymethyltransferase was probably replaced in mung bean seine hydroxymethyltransferase by a covalently bound carbonyl group: (a) inhibition by phenylhydrazine and hydroxylamine, which could not be reversed by dialysis and or addition of pyndoxal-5'phosphate; (b) irreversible inactivation by sodium borohydride; (c) a spectrum characteristic of a phenylhydrazone upon interaction with phenylhydrazine; and (d) the covalent labeling of the enzyme with substrate/product serine and glycine upon reduction with sodium borohydride. These results indicate that in mung bean seine hydroxymethyltransferase, a covalently bound carbonyl group has probably replaced the pyridoxal-5'-phosphate that is present in the mammalian and bacterial enzymes.

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Serine Transhydroxymethylase of Cauliflower (Brassica oleracea var. botrytis L.): Partial Purification and Properties

Cited by 21 publications

References 21 publications

Photorespiration-deficient Mutants of Arabidopsis thaliana Lacking Mitochondrial Serine Transhydroxymethylase Activity

Photorespiration-deficient Mutants of Arabidopsis thaliana Lacking Mitochondrial Serine Transhydroxymethylase Activity

Purification and Regulatory Properties of Mung Bean (Vigna Radiata L.) Serine Hydroxymethyltransferase

Serine Hydroxymethyltransferase from Mung Bean (Vigna radiata) Is Not a Pyridoxal-5′-Phosphate-Dependent Enzyme

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