Subunit structure and activity of glyceraldehyde-3-phosphate dehydrogenase from spinach chloroplasts

Ferri, Giuseppina; Comerio, Giovanna; Ladarola, Paolo; Zapponi, Maria Carla; Speranza, Maria Luisa

doi:10.1016/0005-2744(78)90318-2

Cited by 55 publications

(33 citation statements)

References 23 publications

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“…Clearly, the C-terminal extension does not affect the response of the B-subunit enzyme to pH. The enzyme from spinach chloroplasts has a similar pH optimum (see Ferri et al, 1978). The pH optima of two NADP-linked G-3-P dehydrogenases from the alga Scenedesmus obliquus are 7.5 (OBrien and Powls, 1976).…”

Section: Ph Optimamentioning

confidence: 99%

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“…It can use either NADP(H) or NAD(H) as the pyridine nucleotide substrate. In angiosperm chloroplasts there are two G-3-P dehydrogenase isozymes: one is composed of two A-subunits and two B-subunits (A,B,, isozyme I) and has a marked tendency to aggregate; the other is composed of four identical A-subunits (A4, isozyme 11) and does not aggregate Chambers, 1978, 1979;Ferri et al, 1978Ferri et al, , 1990Cerff, 1979;Pupillo and Faggiani, 1979;Scagliarini et al, 1993).The A-and B-subunits are very similar, except that the B-subunit has a highly negatively charged C-terminal extension that contains two Cys groups and is not found in other G-3-P dehydrogenases (Shih et al, 1986(Shih et al, , 1991Brinkmann et al, 1989;Liaud et al, 1990;Baalmann et al, 1996) This work was supported by National Science Foundation grant no. MCB-9513498.…”

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Expression and Characterization of Pea Chloroplastic Glyceraldehyde-3-Phosphate Dehydrogenase Composed of Only the B-Subunit

Anderson

1997

Plant Physiology

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A cDNA fragment coding for the pea (Pisum sativum 1.) chloroplastic glyceraldehyde-3-P dehydrogenase (EC 1.2.1.1 3) B-subunit and a truncated form corresponding in length to the A-subunit have been cloned into an expression vector, expressed in the absence of the A-subunit in a gap-Escherichia coli strain, purified, and studied. Like the isolated enzyme from higher plant chloroplasts, the recombinant enzymes have dual specificity for NADPH and NADH. l h e recombinant glyceraldehyde-3-P dehydrogenases have the same optimal pH as the enzyme isolated from pea chloroplasts. Like the native chloroplast enzyme, the recombinant B-subunit has a marked tendency to form large aggregates, whereas the truncated B-subunit exists as the tetramer. The recombinant 8-subunit glyceraldehyde 3-P dehydrogenase is more sensitive to dithiothreitol than its truncated form. It seems likely that a different pair of cysteines is responsible for the redox sensitivity of the activity of the enzyme composed of B-subunits than the cysteine residues implicated in the modulation of the activity of the enzyme composed of A-subunits by previous work in this laboratory.In chloroplasts G-3-P dehydrogenase (EC 1.2.1.13) catalyzes the reversible reduction and dephosphorylation of 1,3-bisphosphoglycerate to G-3-P using NADPH generated by PSI in the light. The enzyme is light-and DTT-activated (Buchanan, 1980; Anderson, 1986). It can use either NADP(H) or NAD(H) as the pyridine nucleotide substrate. In angiosperm chloroplasts there are two G-3-P dehydrogenase isozymes: one is composed of two A-subunits and two B-subunits (A,B,, isozyme I) and has a marked tendency to aggregate; the other is composed of four identical A-subunits (A4, isozyme 11) and does not aggregate Chambers, 1978, 1979;Ferri et al., 1978Ferri et al., , 1990Cerff, 1979;Pupillo and Faggiani, 1979;Scagliarini et al., 1993).The A-and B-subunits are very similar, except that the B-subunit has a highly negatively charged C-terminal extension that contains two Cys groups and is not found in other G-3-P dehydrogenases (Shih et al., 1986(Shih et al., , 1991Brinkmann et al., 1989;Liaud et al., 1990;Baalmann et al., 1996) This work was supported by National Science Foundation grant no. MCB-9513498.Present address: Department of Genetics, Research Institute, Hospital for Sick Children, 555 University Avenue, Toronto, Ontario, Canada M5G 1x8.* Corresponding author; e-mail louise@uic.edu; fax 1-312-413-2435. (Fig. 1). Both subunits are encoded in the nucleus (Cerff and Kloppstech, 1982;Shih et al., 1986). The existence of a B, tetramer in vivo has not been suggested. Baalmann et al. (1996) have developed an expression system for the A-and B-subunits of spinach (Spinacia oleracea) chloroplastic G-3-P dehydrogenase. Here we describe an expression system for the pea (Pisum sativum) chloroplastic G-3-P dehydrogenase B-subunit and for a truncated form corresponding in length to the A-subunit, and the characterization of the recombinant enzymes.

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Section: Ph Optimamentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Expression and Characterization of Pea Chloroplastic Glyceraldehyde-3-Phosphate Dehydrogenase Composed of Only the B-Subunit

Anderson

1997

Plant Physiology

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“…The chloroplast stromal marker NADP-GAPDH was quantified using the method of Ferri et al (1978). Catalase was assayed spectrophotometrically for use as a peroxisome marker (Luck, 1965).…”

Section: Enzymatic Marker Assaysmentioning

confidence: 99%

Light-Dependent Isoprene Emission (Characterization of a Thylakoid-Bound Isoprene Synthase in Salix discolor Chloroplasts)

1996

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lsoprene synthase is an enzyme that is responsible for the production of the volatile C5 hydrocarbon, isoprene, in plant leaves. lsoprene formation in numerous C3 plants is interesting because (a) large quantities of isoprene are emitted, 5 x IOl4 g of C annually, (b) a plant may release 1 to 8% of its fixed C as isoprene, and (c) the function of plant isoprene production is unknown. Because of the dependence of foliar isoprene emission on light, the existence of a plastidic isoprene synthase has been postulated. To pursue this idea, a method to isolate chloroplasts from Salix discolor was developed and shows a plastidic isoprene synthase that is tightly bound to the thylakoid membrane and accessible to trypsin inactivation. The thylakoid-bound isoprene synthase has catalytic properties similar to known soluble isoprene synthases; however, the relationship between these enzymes is unknown. The discovery of a thylakoidbound isoprene synthase with a stromal-facing domain places it in the chloroplast, where it may be subject to numerous direct and indirect light-mediated effects. lmplications for the light-dependent regulation of foliar isoprene production and its function are presented.

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“…The second (E.C. 1.2.1.13), closely related to the glycolytic type, is involved in the Calvin cycle and exhibits a dual-coenzyme specificity (Ferri, Comerio, Iadarola, Zapponi & Speranza, 1978). Apart from its essential role in the glycolytic pathway, various other cellular roles have been recently attributed to the strictly NAD-dependent GAPDH (Forthergill-Gilmore & Michels, 1992).…”

Section: Introductionmentioning

confidence: 99%

Crystallization and preliminary X-ray diffraction studies ofEscherichia coliglyceraldehyde-3-phosphate dehydrogenase

Olivier

Buisson²,

Fanchon³

et al. 1995

Acta Cryst D

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Phosphorylating glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key enzyme in glycolysis. Single crystals of NAD-dependent GAPDH from Escherichia coli have been obtained by vapour diffusion at room temperature using trisodium citrate as precipitant. In almost the same crystallization conditions, two kinds of crystals were found to be suitable for X-ray diffraction. The crystals with only one half of a tetramer in the asymmetric unit were chosen for highresolution analysis. They belonged to space group C2221, with cell dimensions a = 79.1, b = 189.6 and c = 122.2 A. These crystals diffracted to 1.8 A resolution.

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Subunit structure and activity of glyceraldehyde-3-phosphate dehydrogenase from spinach chloroplasts

Cited by 55 publications

References 23 publications

Expression and Characterization of Pea Chloroplastic Glyceraldehyde-3-Phosphate Dehydrogenase Composed of Only the B-Subunit

Expression and Characterization of Pea Chloroplastic Glyceraldehyde-3-Phosphate Dehydrogenase Composed of Only the B-Subunit

Light-Dependent Isoprene Emission (Characterization of a Thylakoid-Bound Isoprene Synthase in Salix discolor Chloroplasts)

Crystallization and preliminary X-ray diffraction studies ofEscherichia coliglyceraldehyde-3-phosphate dehydrogenase

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