The 64 kDa polypeptide of spinach may not be the LHCII kinase, but a lumen‐located polyphenol oxidase

Sokolenko, Anna; Fulgosi, Hrvoje; Gal, Adiv; Altschmied, Lothar; Ohad, Itzhak; Herrmann, Reinhold G.

doi:10.1016/0014-5793(95)00892-d

Cited by 49 publications

(29 citation statements)

References 43 publications

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“…They can also create a considerable amount of confusion and ambiguity as to the real identity of kinase substrates (25). Therefore, we have employed functional renaturation assays in a search for outer envelope kinases.…”

Section: Discussionmentioning

confidence: 99%

“…Therefore, we have employed functional renaturation assays in a search for outer envelope kinases. This approach has so far been successfully used in several investigations of other chloroplast protein kinases (25,26). Renaturations in the presence of unspecific substrates failed to show kinase activity in outer envelopes, but simultaneously demonstrated that, at least in vitro, no protein from this chloroplast subfraction is able to autophosphorylate.…”

Section: Discussionmentioning

confidence: 99%

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The Chloroplast Protein Import Receptors Toc34 and Toc159 Are Phosphorylated by Distinct Protein Kinases

Fulgosi¹,

Soll²

2002

Journal of Biological Chemistry

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The molecular composition of chloroplast outer and inner envelope translocons is fairly well established, but little is known about mechanisms and elements involved in import regulation. After synthesis in the cytosol, chloroplast targeted precursor proteins are recognized by outer envelope receptors Toc34 and Toc159. Phosphorylation plays an important role in regulation of Toc34 activity and preprotein binding. Using kinase renaturation assays, we have identified an ATP-dependent 98-kDa outer envelope kinase which is able to selectively phosphorylate Toc34 at a specific site. A 70-kDa outer envelope polypeptide phosphorylating Toc159 was identified by the same strategy. Antiserum against the 98-kDa kinase inhibits phosphorylation of Toc34, whereas labeling of Toc159 remains unaffected. Both kinases do not autophosphorylate in vitro and are unable to utilize myelin basic protein as substrate. We propose that distinct kinases are involved in regulation of chloroplast import via desensitization of preprotein receptors.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

The Chloroplast Protein Import Receptors Toc34 and Toc159 Are Phosphorylated by Distinct Protein Kinases

Fulgosi¹,

Soll²

2002

Journal of Biological Chemistry

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“…As part of a project aimed at the identification of the thylakoidbound protein kinase(s) (19,22), we have partially purified a solubilized protein kinase preparation and assayed the kinasesubstrate interaction in an in vitro reconstituted system. Experimental results obtained with this system disclosed that light may contribute to the regulation of LHCII phosphorylation by affecting the conformation of its N-terminal domain.…”

Section: Discussionmentioning

confidence: 99%

“…Kinase activity of protein bands in the various fractions was detected by electrotransfer of the proteins after denaturing SDS͞PAGE to poly-(vinylidene difluoride) membranes followed by renaturation of the kinase as in ref. 22. The renatured kinase bands did not exhibit self-phosphorylation and were detected by using 32 P-␥-ATP-Mg and histone S-III as a substrate.…”

Section: Methodsmentioning

confidence: 99%

Regulation of thylakoid protein phosphorylation at the substrate level: Reversible light-induced conformational changes expose the phosphorylation site of the light-harvesting complex II

Zer

Vink

Keren

et al. 1999

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

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Light-dependent activation of thylakoid protein phosphorylation regulates the energy distribution between photosystems I and II of oxygen-evolving photosynthetic eukaryotes as well as the turnover of photosystem II proteins. So far the only known effect of light on the phosphorylation process is the redox-dependent regulation of the membranebound protein kinase(s) activity via plastoquinol bound to the cytochrome bf complex and the redox state of thylakoid dithiols. By using a partially purified thylakoid protein kinase and isolated native chlorophyll (chl) a͞b light-harvesting complex II (LHCII), as well as recombinant LHCII, we find that illumination of the chl-protein substrate exposes the phosphorylation site to the kinase. Light does not activate the phosphorylation of the LHCII apoprotein nor the recombinant pigment-reconstituted complex lacking the N-terminal domain that contains the phosphothreonine site. The suggested light-induced conformational change exposing the Nterminal domain of LHCII to the kinase is evidenced also by an increase in its accessibility to tryptic cleavage after light exposure. Light activates preferentially the trimeric form of LHCII, and the process is paralleled by chl f luorescence quenching. Both phenomena are slowly reversible in darkness. Light-induced exposure of the LHCII N-terminal domain to the endogenous protein kinase(s) and tryptic cleavage occurs also in thylakoid membranes. These results demonstrate that light may regulate thylakoid protein phosphorylation not only via the signal transduction chain connecting redox reactions to the protein kinase activation, but also by affecting the conformation of the chl-protein substrate.Redox-controlled thylakoid protein phosphorylation in photosynthetic eukaryotes plays an important role in the regulation of the light energy distribution between the two photosystems (PSs) and controls the light-induced turnover of PSII reaction center subunits. The reversible association͞ dissociation of a mobile pool of the light-harvesting chlorophyll (chl) a͞b protein complex (LHCII) with PSII (state I to state II transition) is ascribed to the redox-controlled activation͞deactivation of thylakoid-bound protein kinase(s) and phosphorylation of the LHCII proteins (1, 2). Phospho-LHCII dissociates from PSII and transfers energy to PSI (state II, reviewed in ref.2). Phospho-LHCII is dephosphorylated by a thylakoid-bound phosphatase (3-5) regulated by a recently discovered cyclophilin-like protein, TLP40, localized in the thylakoid lumen (6, 7). After dephosphorylation, LHCII reassociates with PSII (state I, refs. 7-9). The state transition process involves lateral migration of LHCII (10) and cytochrome bf complex from the appressed to the nonappressed thylakoid domains, thereby enhancing PSI cyclic electron flow (11).The signal transduction loop interconnecting light-driven electron flow with the thylakoid protein kinase(s) activation involves the interaction of reduced plastoquinone with the quinol oxidation site of the cytochrome bf ...

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“…The ensemble of known luminal proteins was small and consisted of the three extrinsic photosystem II proteins (PsbO, PsbP, and PsbQ) and plastocyanin. This group was later joined by some new proteins such as violaxanthin de-epoxidase (7), polyphenol oxidase (8,9), the extrinsic photosystem I protein PsaN (10), and the carboxylterminal processing protease for the D1 protein (11).…”

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confidence: 99%

Proteome Map of the Chloroplast Lumen of Arabidopsis thaliana

Schubert¹,

Petersson²,

Haas³

et al. 2002

Journal of Biological Chemistry

408

391

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The thylakoid membrane of the chloroplast is the center of oxygenic photosynthesis. To better understand the function of the luminal compartment within the thylakoid network, we have carried out a systematic characterization of the luminal thylakoid proteins from the model organism Arabidopsis thaliana. Our data show that the thylakoid lumen has its own specific proteome, of which 36 proteins were identified. Besides a large group of peptidyl-prolyl cis-trans isomerases and proteases, a family of novel PsbP domain proteins was found. An analysis of the luminal signal peptides showed that 19 of 36 luminal precursors were marked by a twin-arginine motif for import via the Tat pathway. To compare the model organism Arabidopsis with another typical higher plant, we investigated the proteome from the thylakoid lumen of spinach and found that the luminal proteins from both plants corresponded well. As a complement to our experimental investigation, we made a theoretical prediction of the luminal proteins from the whole Arabidopsis genome and estimated that the thylakoid lumen of the chloroplast contains ϳ80 proteins.The ability to perform oxygenic photosynthesis belongs to the distinguishing characteristics of higher plants, algae, and cyanobacteria. In higher plants, the center of the photosynthetic process is the thylakoid membrane of the chloroplast. Here, in a synergistic series of reactions, four protein complexes, the photosystems I and II, the cytochrome b 6 f complex, and the ATP-synthase, produce NADPH and ATP that fuel the further synthesis of carbohydrates (1, 2).A key feature in the energy conversion of photosynthesis is the link between the electron transfer from photosystem II to I via the cytochrome b 6 f complex and the generation of a proton gradient over the thylakoid membrane. To balance the flow of electrical charges during the formation of the proton gradient, there is a busy traffic of chloride and calcium ions from the stroma into the lumen and of magnesium ions from the lumen into the stroma (3-6). This ion traffic plays a fundamental role for the proper function of photosynthesis. For a long time it was believed that accumulating protons and balancing the ion currents over the thylakoid membrane was the main function of the luminal compartment. The ensemble of known luminal proteins was small and consisted of the three extrinsic photosystem II proteins (PsbO, PsbP, and PsbQ) and plastocyanin. This group was later joined by some new proteins such as violaxanthin de-epoxidase (7), polyphenol oxidase (8, 9), the extrinsic photosystem I protein PsaN (10), and the carboxylterminal processing protease for the D1 protein (11).To achieve a more profound understanding of content and functions of the thylakoid lumen, we designed a method that enabled us to isolate a highly pure fraction of luminal proteins from spinach thylakoids. For the first time, we showed that the lumen of the thylakoid membrane contained at least 20 proteins and that the protein concentration of this compartment was similar to tha...

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The 64 kDa polypeptide of spinach may not be the LHCII kinase, but a lumen‐located polyphenol oxidase

Cited by 49 publications

References 43 publications

The Chloroplast Protein Import Receptors Toc34 and Toc159 Are Phosphorylated by Distinct Protein Kinases

The Chloroplast Protein Import Receptors Toc34 and Toc159 Are Phosphorylated by Distinct Protein Kinases

Regulation of thylakoid protein phosphorylation at the substrate level: Reversible light-induced conformational changes expose the phosphorylation site of the light-harvesting complex II

Proteome Map of the Chloroplast Lumen of Arabidopsis thaliana

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