Development and Intracellular Distribution of Enzymes of the Oxidative Pentose Phosphate Cycle in Radish Cotyledons

Abstract

Developmeental and compartmenitation studies were used to evaluate the relative roles of the oxidative pentose phosphate cycle, the Calvin cycle, and the glycoly-sis in cotvledons of radish (Raphanus sativus L.).Glucose-6-P dehydrogenase, 6-P-gluconate dehydrogenase, glucose-6-P isomerase, and the NAD-dependent glyceraldehyde-3-P dehydrogenase were present in high activity in ungerminated seeds, increased about 2-fold during germination in the dark, and were slightly enhanced by light. In contrast, NADP-dependent glyceraldehvde-3-P dehydrogenase was developed to only a small degree in the dark, but increased severalfold in continuous white or far red light. The activity of phosphofructokinase was low throughout germinationi.The separation of cell conmpartment-specific isoenzymes showed that, except in ungerminated seeds, the plastid enzyme accouinted for 40 to 45% of the total activity of 6-Pgluconate dehydrogenase and for 13 to 20% of glucose-6-P isomerase. The remaining activity was due to the cytosolic isoenzymes. The presence of gltucose-6-P dehydrogenase and 6-P-gluconate dehydrogeniase in plastids was also established by their presence in the isolated organelle. The NAD -dependenit glyceraldehyde-3 -P dehydrogeniase was mostly due to the cytosolic isoenzyme, whereas the NADdependent activity associated with the NADP-dependent glyceraldehyde-3-P dehydrogenase was very small.The data indicate that the en-zynmes of the oxidative penitose phosphate cycle are present in the cytosol throughotut germiniation. IIn the plastids these enzy-iies already becanie fully developed duiring early germninationi in the dark, whereas entzymes of the Calvin cycle increased onlly in the light. Glycolvsis seemned to be of m-linor importanice.Enzymes involved in the sugar phosphate metabolism of plants have been found in two subcellular compartments, the chloroplasts and the cytosol (15, 25). Only enzymes specific for the Calvin cycle were restricted to the chloroplasts (12,16,27). When enzyme activities are present in both cell compartments, they are due to isoenzymes, one located in the chloroplasts and the other in the cytosol (1, 2, 4). This distribution also applies to the two key enzymes of the oxidative pentose phosphate cycle, glucose-6-P dehydrogenase and 6-P-gluconate dehydrogenase (11,25,26).At present only little evidence is available about the intra-'This investigation was supported by the Deutsche Forschungsgemeinschaft. cellular distribution of enzymes in etiolated plant tissues, since most work has been performed using green leaves. In general, one can assume the same intracellular distribution of enzymes in etiolated tissues as in green leaves. In addition to changing levels of total enzyme activities in etiolated tissues (10, 28) it is possible that the quantitative distribution may be different in the two cellular compartments. Previous studies have measured only total enzyme activities from crude extracts, so that the extent of compartmentation of enzyme activities in the plastids and in the cytosol wa...

Enzymatic Evidence for a Complete Oxidative Pentose Phosphate Pathway in Chloroplasts and an Incomplete Pathway in the Cytosol of Spinach Leaves

“…However, it has been possible by using anion-exchange chromatography to separate a11 other chloroplast/cytosol isoenzymes of sugar phosphate metabolism investigated to date, e.g. starch phosphorylase (Steup et al, 1980;Mateyka and Schnarrenberger, 1984), phosphoglucomutase (Mühlbach and Schnarrenberger, 1978), Glc-P isomerase (Schnarrenberger and Oeser, 1974), FBPase (Latzko et al, 19741, FBP aldolase (Kriiger and Schnarrenberger, 1983;Lebherz et al, 1984;Schnarrenberger and Kriiger, 1986), hexokinase (Schnarrenberger, 1990), G6PDH and 6PGDH (Schnarrenberger et al, 1973(Schnarrenberger et al, , 1975Simcox et al, 1977;Simcox and Dennis, 1978;Nishimura and Beevers, 19811, glyceraldehyde-3-P dehydrogenase (Miernyk and Dennis, 19821, 3-phosphoglycerate kinase (Miernyk and Dennis, 1982;Kopke-Secundo et al, 1990;Morrow and Bradbeer, 1990), triose-P isomerase (Kurzok and Feierabend, 1984), pyruvate kinase (Ireland et al, 1979), and enolase and phosphoglycerate mutase (Miernyk and Dennis, 1982;Plaxton, 1989). It is conceivable, yet we consider it highly unlikely, that isoenzymes for transaldolase, transketolase, Rib-5-P isomerase, and ribulose-5-P epimerase exist but are undetectable.…”

“…It was unequivocally shown that chloroplast/cytosol isoenzymes exist for G6PDH and 6PGDH both in green leaf cells (Schnarrenberger et al, 1973) and in nongreen tissues (Schnarrenberger et al, 1975;Simcox et al, 1977;Simcox and Dennis, 1978;Nishimura and Beevers, 1981). Cytosolic G6PDH has even been cloned from potato tuber (Graeve et al, 1994).…”

“…In another instance, a widely held view was also contradicted in that plant cells, in general, possess a full complement of plastidic and cytosolic isoenzymes of sugar phosphate metabolism. In this case Cklamydomonas reinkardtii and Dunaliella bioculata appear to lack cytosolic isoforms of glycolytic and gluconeogenetic enzymes (Klein, 1986;Schnarrenberger et al, 1990Schnarrenberger et al, , 1994. Therefore, the a priori assumption that cytosolic isoenzymes generally exist for activities of sugar phosphate metabolism in plants should be viewed more critically until the presence of such isoenzymes is demonstrated beyond doubt.…”

Phosphoglucose isomerase (EC 5.3.1.9) isozymes in diploid and tetraploid Polygala species: evidence for gene duplication and diversification

“…Tables 1 and 3 show that there are clear differences between the populations studied. Gene frequencies (table 3) Distinct plastid (PGI-1) and cytosolic (PGI-2) forms of PGI, coded for at different loci, have been shown to exist in several species of angiosperm (Schnarrenberger and Oeser, 1974;Schnarrenberger et a!., 1975;Gottlieb and Weeden, 1979;1981;Weeden and Gottlieb, 1979;Jones, 1984). The most anodal PGI band was the same in all three species of Polygala, a lack of diversity typical of plastid isozymes (Gottlieb and Weeden, 1981).…”

Genetics of cytosolic phosphoglucose isomerase (PGI) variation in the Amazonian tree Pseudobombax munguba (Bombacaceae)

“…All individuals shared a single band in the most anodal zone (approximately 55 mm from the origin). This enzyme, designated PGI-1, was not well resolved on most gels, and, by analogy with the findings of studies on other plant species, was inferred to be the chloroplastic PGI (Schnarrenberger et al, 1975;Weeden & Gottlieb, 1980b;Gottlieb & Weeden, 1981;Gottlieb 1982). …”