Regulation of protoporphyrin IX biosynthesis by intraplastidic compartmentalization and adenosine triphosphate

Manohara, M S; Tripathy, Baishnab C.

doi:10.1007/s004250000363

Cited by 11 publications

(11 citation statements)

References 32 publications

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“…For example, the activity of glutamate-1-semialdehyde aminomutase in etiolated barley seedlings increases dramatically in the light whereas the level of the corresponding mRNA remains constant (21). Further, it has been observed that DTT alleviates the inhibition of the synthesis of an intermediate of the pathway, protoporphyrin IX under oxidizing conditions (22). Finally, the second potential target, uroporphyrinogen decarboxylase, catalyzes a reaction in common with heme biosynthesis, and the third, magnesium chelatase, catalyzes the insertion of magnesium, the first dedicated step of chlorophyll synthesis (21) that is considered pivotal to plastid-tonucleus signaling (23,24).…”

Section: Resultsmentioning

confidence: 93%

Proteomics gives insight into the regulatory function of chloroplast thioredoxins

Balmer

Koller

Val

et al. 2002

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

399

370

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Thioredoxins are small multifunctional redox active proteins widely if not universally distributed among living organisms. In chloroplasts, two types of thioredoxins ( f and m) coexist and play central roles in regulating enzyme activity. Reduction of thioredoxins in chloroplasts is catalyzed by an iron-sulfur disulfide enzyme, ferredoxin-thioredoxin reductase, that receives photosynthetic electrons from ferredoxin, thereby providing a link between light and enzyme activity. Chloroplast thioredoxins function in the regulation of the Calvin cycle and associated processes. However, the relatively small number of known thioredoxin-linked proteins (about 16) raised the possibility that others remain to be identified. To pursue this opportunity, we have mutated thioredoxins f and m, such that the buried cysteine of the active disulfide has been replaced by serine or alanine, and bound them to affinity columns to trap target proteins of chloroplast stroma. The covalently linked proteins were eluted with DTT, separated on gels, and identified by mass spectrometry. This approach led to the identification of 15 potential targets that function in 10 chloroplast processes not known to be thioredoxin linked. Included are proteins that seem to function in plastid-to-nucleus signaling and in a previously unrecognized type of oxidative regulation. Approximately two-thirds of these targets contained conserved cysteines. We also identified 11 previously unknown and 9 confirmed target proteins that are members of pathways known to be regulated by thioredoxin. In contrast to results with individual enzyme assays, specificity for thioredoxin f or m was not observed on affinity chromatography.

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Section: Resultsmentioning

confidence: 93%

Proteomics gives insight into the regulatory function of chloroplast thioredoxins

Balmer

Koller

Val

et al. 2002

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

399

370

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“…Tetrapyrrole biosynthesis. It has been demonstrated that synthesis of protoporphyrin IX from 5-aminolevulinic acid is stimulated by DTT in chloroplast extracts (36). Three enzymes from the chloroplast heme and chlorophyll synthesis pathway recently were found to interact with Trxs (6).…”

Section: Resultsmentioning

confidence: 99%

Thioredoxin-linked processes in cyanobacteria are as numerous as in chloroplasts, but targets are different

Lindahl

Florencio

2003

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

160

152

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Light-dependent regulation of a growing number of chloroplast enzymatic activities has been found to occur through the reversible reduction of intra-or intermolecular disulphides by thioredoxins. In cyanobacteria, despite their similarity to chloroplasts, no proteins have hitherto been shown to interact with thioredoxins, and the role of the cyanobacterial ferredoxin͞thioredoxin system has remained obscure. By using an immobilized cysteine 35-to-serine site-directed mutant of the Synechocystis sp. PCC 6803 thioredoxin TrxA as bait, we screened the Synechocystis cytosolic and peripheral membrane protein complements for proteins interacting with TrxA. The covalent bond between the isolated target proteins and mutated TrxA was confirmed by nonreducing͞reducing twodimensional SDS͞PAGE. Thus, we have identified 18 cytosolic proteins and 8 membrane-associated proteins as candidate thioredoxin substrates. Twenty of these proteins have not previously been associated with thioredoxin-mediated regulation. Phosphoglucomutase, one of the previously uncharacterized thioredoxinlinked enzymes, has not earlier been considered a target for metabolic control through disulphide reduction. In this article, we show that phosphoglucomutase is inhibited under oxidizing conditions and activated by DTT and reduced wild-type TrxA in vitro. The results imply that thioredoxin-mediated redox regulation is as extensive in cyanobacteria as in chloroplasts but that the subjects of regulation are largely different. Dithiol͞disulphide exchange catalyzed by thioredoxin (Trx) forms the molecular basis for light-dependent regulation of many enzymes in the chloroplasts of higher plants and algae (1-3). Ferredoxin receives reducing equivalents from the photosynthetic electron transport in the light and reduces Trx m and f by means of ferredoxin-Trx reductase. The Trxs, in turn, convert disulphides to dithiols in their respective target enzymes, thereby modulating their activities. The earliest discovered targets for Trx-mediated regulation belong to the Calvin cycle of CO 2 assimilation (1). Since the initial discoveries, several more chloroplast enzymes have been recognized as substrates for Trx (3). A breakthrough in the investigation of chloroplast redox regulation came with the introduction of a new method to isolate Trx target proteins (4-6). The method involves mutation of the buried redox-active cysteine of Trx, which favors the formation of stable mixed disulphides with its target proteins. This experimental approach confirmed the interaction between Trx and its known targets and more than doubled the number of potential Trx-regulated proteins (6).Cyanobacteria are oxygenic photosynthetic prokaryotes that probably share a common ancestor with the chloroplast. The complete sequence of the cyanobacterium Synechocystis sp. PCC 6803 genome (Cyanobase, www.kazusa.or.jp͞cyano͞cyano. html) reveals that this organism, hereafter referred to as Synechocystis, contains ferredoxin-Trx reductase and at least four different Trxs. Nevertheless, attempts to ...

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“…They were resuspended in 0.3 ml of TE buffer and were subsequently layered on top of the discontinuous sucrose gradient and centrifuged as above. After centrifugation, envelope membranes at the interface of the 0.46 M and 1.0 M sucrose gradient were collected and washed once with 10 ml of TE buffer (Manohara and Tripathy 2000). The envelope membranes were resuspended in a minimal amount of TE buffer.…”

Section: Isolation Of Purified Intact Chloroplastsmentioning

confidence: 99%

“…Lysed chloroplasts were centrifuged in a sucrose step gradient (Manohara and Tripathy 2000). All operations were done in diffuse green light to minimize phototransformation of Pchlide to Chlide.…”

Section: Isolation Of Purified Intact Chloroplastsmentioning

confidence: 99%

“…5-Aminolevulinic acid (ALA) biosynthetic enzymes are located in the stroma (Kannangara et al 1994). Interplay of envelope, stroma and thylakoids is shown for protoporphyrin IX (Proto IX) synthesis and enzymes responsible for conversion of ALA to Proto IX, i.e., ALA dehydratase, porphobilinogen deaminase, uroporphyrinogen decarboxylase and coproporphyrinogen oxidase are mostly located in the stromal phase (Manohara and Tripathy 2000). All subsequent steps of Chl biosynthesis are catalyzed by membrane-bound or membrane-associated enzymes ; Lee et al 1992;Matringe et al 1992).…”

Section: Introductionmentioning

confidence: 99%

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Differential distribution of chlorophyll biosynthetic intermediates in stroma, envelope and thylakoid membranes in Beta vulgaris

Mohapatra

Tripathy

2007

Photosynth Res

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Stroma, envelope and thylakoid membranes were prepared from chloroplasts isolated from leaves of Beta vulgaris. Out of total plastidic protochlorophyllide, envelope membranes contained 1.5%, thylakoids had the maximum 98.48% and stroma had a trace fraction of 0.02%. Distribution of the Mg-protoporphyrin IX and its monoester was 89.0% in thylakoids, 10.0% in stroma and 1.0% in envelope. A substantial fraction (33.77%) of plastidic protoporphyrin IX was partitioned into stroma. Envelope contained 0.66% and thylakoids had 65.57% of the total plastidic protoporphyrin IX pool. The proportion of monovinyl and divinyl forms of protochlorophyllide was almost similar in intact plastid, thylakoids, and outer and inner envelope membranes suggesting a tight regulation of vinyl reductase enzyme. The significance of differential distribution of chlorophyll biosynthetic intermediates among thylakoids, envelope and stroma is discussed.

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Regulation of protoporphyrin IX biosynthesis by intraplastidic compartmentalization and adenosine triphosphate

Cited by 11 publications

References 32 publications

Proteomics gives insight into the regulatory function of chloroplast thioredoxins

Proteomics gives insight into the regulatory function of chloroplast thioredoxins

Thioredoxin-linked processes in cyanobacteria are as numerous as in chloroplasts, but targets are different

Differential distribution of chlorophyll biosynthetic intermediates in stroma, envelope and thylakoid membranes in Beta vulgaris

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