Isoenzymes of Pyruvate Kinase in Etioplasts and Chloroplasts

Ireland, Robert J.; Deluca, Vincenzo; Dennis, David T.

doi:10.1104/pp.63.5.903

Cited by 56 publications

(40 citation statements)

References 13 publications

(12 reference statements)

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“…Although the evidence is as yet fragmentary, it seems that the situation is similar in other developing oilseeds (at least sunflower, safflower, soybean, and cotton seeds) and in some chloroplasts (10) and chromoplasts (15) as well. Nishimura and Beevers (21) recently reported that germinating castor oil seeds contain plastid and cytosolic isozymes of the enzymes of sugar-phosphate metabolism.…”

Section: Resultsmentioning

confidence: 99%

Isozymes of the Glycolytic Enzymes in Endosperm from Developing Castor Oil Seeds

Miernyk¹,

Dennis²

1982

Plant Physiol.

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Ion filtration chromatography on diethylaminoethyl-Sephadex A-25 (24). The activities of pyruvate kinase (2), gluconate-6-P dehydrogenase, glucose phosphate isomerase and aldolase (25), and phosphofructokinase (7, 8) associated with the plastids are due to distinct isozymes.Several isotope incorporation studies support the concept of plastid glycolysis and subsequent fatty acid synthesis from pyruvate (17,18,29). However, this does not appear to be universal. Insignificant phosphoglycerate mutase or enolase activities were found in particulate preparations from germinating Cucurbitapepo cotyledons (1), and phosphoglycerate mutase activity was not detected in isolated pea chloroplasts (27). It has been suggested that hexokinase activity is located only on the outer surface of pea chloroplasts and might represent an artifact associated with tissue disruption (28). In some cases, plastid-cytosol shuttles have been suggested to bypass missing enzymic steps (28). Reports that various plastids might lack specific enzymes of the complete glycolytic pathway prompted us to examine further the plastids from developing castor oil seeds. We now report that the association of triose-P isomerase, glyceraldehyde-3-P dehydrogenase, glycerate-3-P kinase, phosphoglycerate mutase, and enolase activities with plastids from castor oil seeds is due to distinct isozymes. ' Supported by the Natural Sciences and Engineering Research Council of Canada. MATERIALS AND METHODSCastor oil plants, Ricinus communis L. cv. Baker 296, were grown in a greenhouse as described previously (26). To obtain the total enzyme complement, developing seeds were homogenized for 60 s with a Polytron (Brinkman Instruments) at a setting of 6, in two volumes of 100 mm K-phosphate (pH 6.9) containing 5 mm MgCl2, and centrifuged at 27,000g for 20 min. The supernatant, after removal of the floating lipid layer, was then used directly. Crude plastid fractions were obtained by differential centrifugation, as described by Simcox et al. (24), using the homogenization medium of Miernyk and Trelease (16). The 10,000g supernatant fraction from plastid isolation was used as the crude cytosolic fraction.Ion filtration chromatography (14) on Sephadex A-25 was done essentially as described by Simcox and Dennis (25). Columns were equilibrated with 20 mm imidazole buffer containing 5 mM MgC12 and, in some cases, 7 mm mercaptoethanol. The pH of the buffer was different for each enzyme separation and is given in the figure legends. After sample application (15 ml), the activities were eluted by application of 500 ml 0.5 M KCI in the equilibration buffer. Columns were run at 90 ml h-1; 4.5 ml fractions were collected. In some cases, where enzyme activity was low, large samples were concentrated by precipitation with 80%1o (w/w) (NH4)2SO4 before loading onto the column.All A readings and enzyme assays were conducted at room temperature (approximately 21°C) using a Cary 210 spectrophotometer. All assays were linear with respect to time and sample concentration. Glycerald...

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Section: Resultsmentioning

confidence: 99%

Isozymes of the Glycolytic Enzymes in Endosperm from Developing Castor Oil Seeds

Miernyk¹,

Dennis²

1982

Plant Physiol.

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“…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.…”

Section: Resultsmentioning

confidence: 99%

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

1995

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l h e intracellular localization of transaldolase, transketolase, ribose-5-phosphate isomerase, and ribulose-5-phosphate epimerase was reexamined in spinach (Spinacia oleracea 1,) leaves. We found a highly predominant if not exclusive localization of these enzyme activities in chloroplasts isolated by isopyknic centrifugation in sucrose gradients. Clucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, glucose phosphate isomerase, and triose phosphate isomerase activity was present in the chloroplast fraction but showed additional activity in the cytosol (supernatant) fraction attributable to the cytosol-specific isoforms known to exist for these enzymes. Anion-exchange chromatography of proteins of crude extracts on diethylaminoethyl-Fractogel revealed only a single enzyme each for transaldolase, transketolase, ribose-5-phosphate isomerase, and ribulose-5-phosphate epimerase. The data indicate that chloroplasts of spinach leaf cells possess the complete complement of enzymes of the oxidative pentose phosphate pathway (OPPP), whereas the cytosol contains only the first two reactions, contrary to the widely held view that plants generally possess a cytosolic OPPP capable of cyclic function. l h e chloroplast enzymes transketolase, ribose-5-phosphate isomerase, and ribulose-5-phosphate epimerase appear to be amphibolic for the Calvin cycle and OPPP.

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“…A possible explication is related with BAP action on the biosynthesis of fatty acids 20 , it could be a factor that influence in the synthesis of enzymatic proteins such as piruvate kinases related with the formation of carbonated chains in the biosynthesis of fatty acids in plants 21 .…”

Section: Discussionmentioning

confidence: 99%

Effect of Plant Growth Regulators on Fatty Acids Composition in <i>Jatropha curcas</i> L. Callus Culture

Hernandez¹,

Mendiola²,

Castro³

et al. 2015

J. Oleo Sci.

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Isoenzymes of Pyruvate Kinase in Etioplasts and Chloroplasts

Cited by 56 publications

References 13 publications

Isozymes of the Glycolytic Enzymes in Endosperm from Developing Castor Oil Seeds

Isozymes of the Glycolytic Enzymes in Endosperm from Developing Castor Oil Seeds

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

Effect of Plant Growth Regulators on Fatty Acids Composition in <i>Jatropha curcas</i> L. Callus Culture

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