Reductive Modification and Nonreductive Activation of Purified Spinach Chloroplast NADP-Dependent Glyceraldehyde-3-phosphate Dehydrogenase

Baalmann, Elisabeth; Backhausen, Jan E.; Rak, Christiane; Vetter, Susanne; Scheibe, Renate

doi:10.1006/abbi.1995.0031

Cited by 89 publications

(92 citation statements)

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“…PRK is strongly regulated by thioredoxins in higher plants and green algae (Porter et al, 1988;Graciet et al, 2004), but apparently is less redox sensitive in cyanobacteria and diatoms (Kobayashi et al, 2003;Michels et al, 2005). GAPDH is finely regulated by thioredoxins, NAD(P)(H), and BPGA in higher plants (Pupillo and Giuliani Piccari, 1975;Wolosiuk and Buchanan, 1978;Trost et al, 1993;Baalmann et al, 1995;Sparla et al, 2002), but it does not appear to be regulated in lower photosynthetic organisms. In fact, in green unicellular algae and in cyanobacteria, the GAPDH is composed of a single type of subunit related to subunit A of higher plants (Figge et al, 1999).…”

Section: Discussionmentioning

confidence: 99%

Reconstitution and Properties of the Recombinant Glyceraldehyde-3-Phosphate Dehydrogenase/CP12/Phosphoribulokinase Supramolecular Complex of Arabidopsis

et al. 2005

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Calvin cycle enzymes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoribulokinase (PRK) form together with the regulatory peptide CP12 a supramolecular complex in Arabidopsis (Arabidopsis thaliana) that could be reconstituted in vitro using purified recombinant proteins. Both enzyme activities were strongly influenced by complex formation, providing an effective means for regulation of the Calvin cycle in vivo. PRK and CP12, but not GapA (A4 isoform of GAPDH), are redox-sensitive proteins. PRK was reversibly inhibited by oxidation. CP12 has no enzymatic activity, but it changed conformation depending on redox conditions. GapA, a bispecific NAD(P)-dependent dehydrogenase, specifically formed a binary complex with oxidized CP12 when bound to NAD. PRK did not interact with either GapA or CP12 singly, but oxidized PRK could form with GapA/CP12 a stable ternary complex of about 640 kD (GapA/CP12/PRK). Exchanging NADP for NAD, reducing CP12, or reducing PRK were all conditions that prevented formation of the complex. Although GapA activity was little affected by CP12 alone, the NADPH-dependent activity of GapA embedded in the GapA/CP12/PRK complex was 80% inhibited in respect to the free enzyme. The NADH activity was unaffected. Upon binding to GapA/CP12, the activity of oxidized PRK dropped from 25% down to 2% the activity of the free reduced enzyme. The supramolecular complex was dissociated by reduced thioredoxins, NADP, 1,3-bisphosphoglycerate (BPGA), or ATP. The activity of GapA was only partially recovered after complex dissociation by thioredoxins, NADP, or ATP, and full GapA activation required BPGA. NADP, ATP, or BPGA partially activated PRK, but full recovery of PRK activity required thioredoxins. The reversible formation of the GapA/CP12/PRK supramolecular complex provides novel possibilities to finely regulate GapA (“non-regulatory” GAPDH isozyme) and PRK (thioredoxin sensitive) in a coordinated manner.

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Section: Discussionmentioning

confidence: 99%

Reconstitution and Properties of the Recombinant Glyceraldehyde-3-Phosphate Dehydrogenase/CP12/Phosphoribulokinase Supramolecular Complex of Arabidopsis

et al. 2005

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“…This switch region consists of 37 amino acid residues (Pro 194 -Ile 230 in case of the spinach chloroplast enzyme) containing two cysteine residues, but its structure is unknown, and there are no data available on how reduction of the disulfide bond leads to activation of the enzyme. Such redox regulation is a common property of socalled thiol enzymes in the chloroplasts like glyceraldehyde-3-phosphate dehydrogenase (17), fructose-1,6-bisphosphatase (18), sedoheptulose-1,7-bisphosphatase (19), phosphoribulokinase (20), and NADP-malate dehydrogenase (21). The chloroplast thioredoxin, which is reduced by electron flow from photosystem, plays a main role in this regulation system (21).…”

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Redox Regulation of the Rotation of F1-ATP Synthase

Bald¹,

Noji²,

Yoshida³

et al. 2001

Journal of Biological Chemistry

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In F 1 -ATPase, the smallest known motor enzyme, unidirectional rotation of the central axis subunit ␥ is coupled to ATP hydrolysis. In the present study, we report the redox switching of the rotation of this enzyme. For this purpose, the switch region from the ␥ subunit of the redox-sensitive chloroplast F O F 1 -ATP synthase, which is located on the mitochondrial inner membranes, chloroplast thyalkoid membranes, and bacterial plasma membranes, synthesizes ATP, which is known as the universal energy "coin" in the cell, from ADP and phosphate at the expense of the proton gradient across the membranes (1). The water-soluble part of ATP synthase, the F 1 region, consists of five subunits with a composition of ␣ 3 ␤ 3 ␥ 1 ␦ 1 ⑀ 1 (2). The hydrophobic F O part is composed of a 1 b 2 c 10 -14 (3-5). The subcomplex of F 1 part, ␣ 3 ␤ 3 ␥, is the minimum complex that is capable of ATP hydrolysis (6). Rotation of the ␥ subunit in the ␣ 3 ␤ 3 hexagon was first postulated by P. D. Boyer and co-workers (8) from the analysis of the cooperative kinetics of F 1 -ATPase. A high resolution x-ray structure of a major part of the bovine heart mitochondrial F 1 (7) revealed an alternating hexagonal arrangement of the ␣ 3 ␤ 3 ring with the ␥ subunit as a central axis. Then the rotation of ␥ in the central cavity of ␣ 3 ␤ 3 was suggested by biochemical experiments (10), and finally, direct visualization of the rotation of an actin filament attached to the ␥ subunit of F 1 showed unequivocally that ␥ rotates unidirectionally during ATP hydrolysis (11)(12)(13)(14).The chloroplast ATP synthase has basically the same structure as those from other sources. However the enzyme activity of chloroplast ATP synthase (CF O CF 1 ) from higher plants is modulated in a unique way, reduction of a disulfide bridge in the ␥ subunit (15, 16). In the solubilized CF 1 part, the reduction of this disulfide bond, which is located in a switch region on ␥ unique to chloroplast ATP synthase, strongly enhances the ATP hydrolysis activity (ATPase activity). This switch region consists of 37 amino acid residues (Pro 194 -Ile 230 in case of the spinach chloroplast enzyme) containing two cysteine residues, but its structure is unknown, and there are no data available on how reduction of the disulfide bond leads to activation of the enzyme. Such redox regulation is a common property of socalled thiol enzymes in the chloroplasts like glyceraldehyde-3-phosphate dehydrogenase (17), fructose-1,6-bisphosphatase (18), sedoheptulose-1,7-bisphosphatase (19), phosphoribulokinase (20), and NADP-malate dehydrogenase (21). The chloroplast thioredoxin, which is reduced by electron flow from photosystem, plays a main role in this regulation system (21). However, to our knowledge there are no data available, neither for chloroplast enzymes nor for enzymes from other organelles, on whether regulation affects the reaction rate of a single enzyme molecule or influences the number of active enzyme molecules. To address this question, observation of the rotation of F 1 -ATPase is th...

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“…[1][2][3][4][5]. Another mode of regulating enzyme activity in the cycle was shown for GAPDH, which, depending on its substrate, 1,3-bisphosphoglycerate, or NADP(H) and ATP, can reversibly dissociate from the less active hexadecameric A8B8 form to the highly active A2B2 tetramer (6,7). GAPDH was also shown to be associated with PRK in a 600-kDa protein complex, made up by the A2B2 and the A4 forms of GAPDH, together with one PRK dimer (8).…”

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CP12 provides a new mode of light regulation of Calvin cycle activity in higher plants

Wedel

Soll

Paap

1997

Proc. Natl. Acad. Sci. U.S.A.

147

220

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CP12 is a small nuclear encoded chloroplast protein of higher plants, which was recently shown to interact with NAD(P)H-glyceraldehyde-3-phosphate dehydrogenase (GAPDH; EC 1.2.1.13), one of the key enzymes of the reductive pentosephosphate cycle (Calvin cycle). Screening of a pea cDNA library in the yeast two-hybrid system for proteins that interact with CP12, led to the identification of a second member of the Calvin cycle, phosphoribulokinase (PRK; EC 2.7.1.19), as a further specific binding partner for CP12. The exchange of cysteines for serines in CP12 demonstrate that interaction with PRK occurs at the N-terminal peptide loop of CP12. Size exclusion chromatography and immunoprecipitation assays reveal the existence of a stable 600-kDa PRK͞ CP12͞GAPDH complex in the stroma of higher plant chloroplasts. Its stoichiometry is proposed to be of two Nterminally dimerized CP12 molecules, each carrying one PRK dimer on its N terminus and one A2B2 complex of GAPDH subunits on the C-terminal peptide loop. Incubation of the complex with NADP or NADPH, in contrast to NAD or NADH, causes its dissociation. Assays with the stromal 600-kDa fractions in the presence of the four different nicotinamideadenine dinucleotides indicate that PRK activity depends on complex dissociation and might be further regulated by the accessible ratio of NADP͞NADPH. From these results, we conclude that light regulation of the Calvin cycle in higher plants is not only via reductive activation of different proteins by the well-established ferredoxin͞thioredoxin system, but in addition, by reversible dissociation of the PRK͞CP12͞

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Reductive Modification and Nonreductive Activation of Purified Spinach Chloroplast NADP-Dependent Glyceraldehyde-3-phosphate Dehydrogenase

Cited by 89 publications

References 0 publications

Reconstitution and Properties of the Recombinant Glyceraldehyde-3-Phosphate Dehydrogenase/CP12/Phosphoribulokinase Supramolecular Complex of Arabidopsis

Reconstitution and Properties of the Recombinant Glyceraldehyde-3-Phosphate Dehydrogenase/CP12/Phosphoribulokinase Supramolecular Complex of Arabidopsis

Redox Regulation of the Rotation of F1-ATP Synthase

CP12 provides a new mode of light regulation of Calvin cycle activity in higher plants

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