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• W2090225411 abstract "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 organismArabidopsis 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 Arabidopsisgenome and estimated that the thylakoid lumen of the chloroplast contains ∼80 proteins. 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 organismArabidopsis 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 Arabidopsisgenome and estimated that the thylakoid lumen of the chloroplast contains ∼80 proteins. Proteome map of the chloroplast lumen of Arabidopsis thaliana.Journal of Biological ChemistryVol. 278Issue 15PreviewThe preparation of lumen content fromArabidopsis chloroplasts (page 8355) should be completed as follows. Arabidopsis chloroplasts were prepared from 100 g of leaves (wet weight) as described (1). The leaves were divided in portions of 20 g that were blended five times for 1 s using a Heidolph DIAX 900 homogenizer in 170 ml of 20 mm Tricine-NaOH (pH 8.4), 300 mm sorbitol, 10 mm EDTA, 10 mm KCl, 0.25% (w/v) bovine serum albumin, 4.5 mm sodium ascorbate, and 5 mm cysteine. The preparation was continued as described (1), but the chlo-roplasts were washed and resuspended in 20 mm Hepes-NaOH (pH 7.8), 300 mm sorbitol, 5 mm MgCl2, 2.5 mm EDTA, and 10 mm KCl. Full-Text PDF Open Access 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid matrix-assisted laser desorption/ionization time-of-flight peptidyl-prolyl cis-trans isomerase 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 cytochromeb 6 f complex, and the ATP-synthase, produce NADPH and ATP that fuel the further synthesis of carbohydrates (1Albertsson P.-Å. Photosynth. Res. 1995; 46: 141-149Crossref PubMed Scopus (117) Google Scholar, 2Wollman F.-A. Minai L. Nechushtai R. Biochim. Biophys. Acta. 1999; 1411: 21-85Crossref PubMed Scopus (218) Google Scholar). 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 (3Hind G. Nakatani H.Y. Izawa S. Proc. Natl. Acad. Sci. 1974; 71: 1484-1488Crossref PubMed Scopus (184) Google Scholar, 4Pottosin I.I. Schönknecht G. J. Membr. Biol. 1995; 148: 143-156Crossref PubMed Scopus (42) Google Scholar, 5Pottosin I.I. Schönknecht G. J. Membr. Biol. 1996; 152: 223-233Crossref PubMed Scopus (77) Google Scholar, 6Ettinger W.F. Clear A.M. Fanning K.J. Peck M.L. Plant Physiol. 1999; 119: 1379-1385Crossref PubMed Scopus (122) Google Scholar). 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 (7Hager H. Holocher K. Planta. 1994; 192: 581-589Crossref Scopus (207) Google Scholar), polyphenol oxidase (8Sommer A. Ne'eman E. Steffens J.C. Mayer A.M. Harel E. Plant Physiol. 1994; 105: 1301-1311Crossref PubMed Scopus (110) Google Scholar, 9Sokolenko A. Fulgosi H. Gal A. Altschmied L. Ohad I. Herrmann R.G. FEBS Lett. 1995; 371: 176-180Crossref PubMed Scopus (39) Google Scholar), the extrinsic photosystem I protein PsaN (10He W.-Z. Malkin R. FEBS Lett. 1992; 308: 298-300Crossref PubMed Scopus (20) Google Scholar), and the carboxyl-terminal processing protease for the D1 protein (11Oelmüller R. Herrmann R.G. Pakrasi H.B. J. Biol. Chem. 1996; 271: 21848-21852Abstract Full Text Full Text PDF PubMed Scopus (65) Google Scholar). 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 that of the stroma (12Kieselbach T. Hagman Å. Andersson B. Schröder W.P. J. Biol. Chem. 1998; 273: 6710-6716Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar). Several new luminal proteins could be characterized in more detail. The 17.4-kDa protein (TL17) had a remarkable new pentapeptide motif and led to the discovery of a whole family of unknown pentapeptide proteins inSynechocystis sp. PCC 6803 (13Kieselbach T. Mant A. Robinson C. Schröder W.P. FEBS Lett. 1998; 428: 241-244Crossref PubMed Scopus (21) Google Scholar). In addition, a novel 16-kDa protein (TL16) was found to be routed into the thylakoid lumen by the Tat translocation pathway (14Mant A. Kieselbach T. Schröder W.P. Robinson C. Planta. 1999; 207: 624-627Crossref PubMed Scopus (5) Google Scholar). The luminal immunophilin TL40 was suggested to participate in signal transduction over the thylakoid membrane (15Fulgosi H. Vener A.V. Altschmied L. Herrmann R.G. Andersson B. EMBO J. 1998; 17: 1577-1587Crossref PubMed Scopus (110) Google Scholar), and Hcf136 was identified as a luminal assembly factor for photosystem II (16Meurer J. Plücken H. Kowallik K.V. Westhoff P. EMBO J. 1998; 17: 5286-5297Crossref PubMed Scopus (181) Google Scholar). Recently, the 29-kDa peroxidase homologue TL29 and a novel plastocyanin were added to the list of new luminal proteins (17Kieselbach T. Bystedt M. Hynds P. Robinson C. Schröder W.P. FEBS Lett. 2000; 480: 271-276Crossref PubMed Scopus (81) Google Scholar) along with several proteins from the thylakoids of pea that were not further characterized (18Peltier J.-B. Friso G. Kalume D.E. Roepstorff P. Nilsson F. Adamska I. van Wijk K.J. Plant Cell. 2000; 12: 319-341Crossref PubMed Scopus (297) Google Scholar). The completion of the Arabidopsis thaliana genome sequencing project by the end of 2000 (19The Arabidopsis Genome Initiative Nature. 2000; 408: 796-815Crossref PubMed Scopus (7211) Google Scholar) started a new era in plant research. To apply the knowledge of the Arabidopsis genome to an investigation into luminal proteins, we designed a method to isolate the luminal proteins from Arabidopsis chloroplasts and studied them by proteomics. In this study, we performed the first systematic characterization of the chloroplast lumen ofArabidopsis and compared its proteins with those from the chloroplast lumen of spinach. We found 36 luminal proteins inArabidopsis, of which 22 could be identified in spinach also. By comparing the experimentally identified lumen proteins ofArabidopsis with a theoretical prediction of a luminal proteome in this organism, we estimated that the chloroplast lumen ofArabidopsis comprises ∼80 proteins. Plants were cultivated hydroponically according to Norén et al.(20Norén H. Svensson P. Andersson B. Biosci. Rep. 1999; 19: 499-509Crossref PubMed Scopus (20) Google Scholar). A. thaliana ecotype Columbia was grown for 13 weeks with 8 h light per day and a light intensity of 100 μmol of photons m−2 s−1, and spinach (Spinacia oleracea) was raised for 5 weeks with 10 h of light per day. The lumen fraction from spinach chloroplasts was isolated according to Kieselbach et al. (12Kieselbach T. Hagman Å. Andersson B. Schröder W.P. J. Biol. Chem. 1998; 273: 6710-6716Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar). The luminal content fromArabidopsis chloroplasts was isolated in the same way with the following minor changes. To avoid protein degradation, 1 mm of EDTA was added to the last washing buffer, and 50 μg/ml of Pefablock was added to the thylakoids just prior to the Yedapress treatment. The lumen fraction was concentrated using Centriprep YM-3 concentrators (Millipore Corp.), and protein quantification was carried out according to Bradford (21Bradford M.M. Anal. Biochem. 1976; 72: 248-254Crossref PubMed Scopus (217544) Google Scholar) using bovine serum albumin as a standard. The luminal proteins were separated by isoelectric focusing in the first and by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) in the second dimension. The samples for analytical two-dimensional gels contained 100 μg of protein, and those for Western blots for microsequencing contained 200 μg of protein. The luminal proteins were solubilized in 5 m urea, 2 m thiourea, 4% (w/v) CHAPS,1 50 mmdithiothreitol, and 0.8% (v/v) carrier ampholytes (IPG buffer, pH 3–10 nonlinear or 4–7 linear Amersham Biosciences, Inc.) and applied during rehydration to a nonlinear IPG strip, pH 3–10, or a to linear IPG strip, pH 4–7. The strips were allowed to rehydrate overnight at 20 °C and then transferred to IPGphor cup-loading strip holders and covered with paraffin oil. The proteins were focused for 10 min at 300 V followed by a 3-h gradient from 300 to 3500 V and a 30-min gradient from 3500 to 8000 V. The isoelectric focusing was then completed at a constant voltage of 8000 V until 60,000 V-h was reached. Subsequently, the strips were equilibrated first for 15 min in 50 mm Tris-HCl (pH 6.8), 6 m urea, 30% (v/v) glycerol, 2% (w/v) SDS, and 1% (w/v) dithiothreitol and then for 10 min in the same buffer without dithiothreitol but with 2.5% (w/v) iodoacetamide and a trace of bromphenol blue. In the second dimension, SDS-PAGE according to Laemmli (22Laemmli U.K. Nature. 1970; 227: 680-685Crossref PubMed Scopus (207537) Google Scholar) was carried out in a gradient slab gel (T = 9–16%) using a Protean II XL system from Bio-Rad. Before the polymerization of the gel, 5 mm of sodium thiosulfate was added to the monomer solution to decrease the background staining with silver. Proteins were detected by silver staining according to Bjellqvist et al. (23Bjellqvist B. Pasquali C. Ravier F. Sanchez J.-C. Hochstrasser D. Electrophoresis. 1993; 14: 1357-1365Crossref PubMed Scopus (365) Google Scholar). The two-dimensional gels were scanned using an image scanner and evaluated with the Image Master two-dimensional software (both from Amersham Biosciences). The apparent masses of the proteins that were detected on the two-dimensional gels were determined manually or with the Image Master two-dimensional software using identified proteins of known masses as a reference. MALDI-TOF analysis of in-gel digested proteins was carried out with a Reflex III mass spectrometer from Bruker. The in-gel digests were performed using sequencing grade modified trypsin (Promega) and analyzed as described (24Shevchenko A. Wilm M. Vorm O. Mann M. Anal. Chem. 1996; 68: 850-858Crossref PubMed Scopus (7831) Google Scholar, 25Pandey A. Andersen J.S. Mann M. Science's stke. 2000; 37: l1Google Scholar). Data base searches were done with the MS BioTools software from Bruker using the Mascot search engine (available on the World Wide Web at www.matrixscience.com). If a protein could not unambiguously be identified by a fingerprint spectrum, its identity was confirmed by a postsource decay analysis of single peptides (26Gevaert K. Demol H. Martens L. Hoorelbeke B. Puype M. Goethals M. De Van Damme J. Boeck S. Vandekerckhove J. Electrophoresis. 2001; 22: 1645-1651Crossref PubMed Scopus (61) Google Scholar). Amino-terminal microsequencing was carried out with a Procise sequencer from Applied Biosystems. Proteins were sequenced from polyvinylidene difluoride membrane following resolution by two-dimensional electrophoresis essentially as described (27Matsudaira P. J. Biol. Chem. 1987; 262: 10035-10038Abstract Full Text PDF PubMed Google Scholar). The individual analysis of protein sequences by similarity searches (28Pearson W.R. Methods Enzymol. 1990; 183: 63-98Crossref PubMed Scopus (1652) Google Scholar, 29Altschul S.F. Madden T.L. Schaffer A.A. Zhang J. Zhang Z. Miller W. Lipman D.J. Nucleic Acids Res. 1997; 25: 3389-3402Crossref PubMed Scopus (60233) Google Scholar), pattern and profile searches (30Hofmann K. Bucher P. Falquet L. Bairoch A. Nucleic Acids Res. 1999; 27: 215-219Crossref PubMed Scopus (1006) Google Scholar, 31Bateman A. Birney E. Durbin R. Eddy S.R. Howe K.L. Sonnhammer E.L. Nucleic Acids Res. 2000; 28: 263-266Crossref PubMed Scopus (1225) Google Scholar), alignments (32Thompson J.D. Higgins D.G. Gibson T.J. Nucleic Acids Res. 1994; 22: 4673-4680Crossref PubMed Scopus (56002) Google Scholar), and hydrophobicity plots (33Kyte J. Doolittle R.F. J. Mol. Biol. 1982; 157: 105-132Crossref PubMed Scopus (17296) Google Scholar) was carried out using the ExPaSy tools (available on the World Wide Web at www.expasy.ch). The prediction of chloroplast-targeted proteins encoded within theA. thaliana genome was performed by subjecting the latest version of the proteome, currently consisting of 25,657 proteins, to an analysis using the program TargetP via the World Wide Web interface (34Emanuelsson O. Nielsen H. Brunak S. von Heijne G. J. Mol. Biol. 2000; 300: 1005-1016Crossref PubMed Scopus (3638) Google Scholar). The NH2-terminal portion of each protein sequence, not exceeding 140 residues in length, was analyzed by the plant version of TargetP, and all chloroplast-predicted proteins (rank 1–5) were used. The prediction of signal peptides and the peptide cleavage products was performed using the portable version of SignalP-2.0 (35Nielsen H. Engelbrecht J. Brunak S. von Heijne G. Protein Eng. 1997; 10: 1-6Crossref PubMed Scopus (4942) Google Scholar,36Nielsen H. Brunak S. von Heijne G. Protein Eng. 1999; 12: 3-9Crossref PubMed Scopus (536) Google Scholar). The programs TargetP and SignalP are available on the World Wide Web at www.cbs.dtu.dk/services, and the Arabidopsis proteome is accessible on the Internet atftp://ftp.tigr.org/pub/data/a_thaliana/ath1/SEQUENCES/ATH1.pep. WhileArabidopsis is an excellent model organism for molecular biological studies, biochemical work has been difficult due to the small leaf size and the small amount of material. The introduction of a hydroponical culture made it possible to overcome this drawback and to grow plants with considerably larger leaves that provided sufficient amounts of plant material in a high quality well suited for biochemical preparations. An essential element in this technique was the restriction of light to 8 h/day, which enabled us to grow the plants for 13 weeks without flowering. Using Arabidopsis plants cultivated in this way, we were able to purify the luminal fraction in the same high quality as was obtained in the original method for the isolation of thylakoid lumen from spinach chloroplasts (12Kieselbach T. Hagman Å. Andersson B. Schröder W.P. J. Biol. Chem. 1998; 273: 6710-6716Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar). A typical lumen preparation from Arabidopsis started with 100–200 g of wet weight leaf material and yielded in ∼2 mg protein/100 g of leaves. An important prerequisite for a concise mapping of the luminalArabidopsis proteins was a reproducible two-dimensional electrophoresis system that was capable of resolving the major part of the luminal proteins. To meet these requirements, we used a combination of nonlinear pH gradients (pH 3–10) and a polyacrylamide gradient gel that allowed us to detect proteins with isoelectric points between 4 and 9 and masses between 200 and 9 kDa. Samples from 14 different lumen preparations were analyzed in more than 20 experiments. The two-dimensional maps of the luminal Arabidopsis proteins showed an excellent reproducibility, and a representative experiment is shown in Fig. 1 A. From the complete set of experiments, 13 two-dimensional gels were selected for a detailed image analysis. The total number of protein spots that were detected on the two-dimensional gels was between 400 and 700, and 277 of those were present on all 13 gels. The protein pattern of the two-dimensional gel in Fig. 1 Ashows clearly that most proteins were detected in the acidic region of the pH gradient, while the group of basic proteins was relatively small. It should be noticed, too, that there is a distinct gap between the proteins in the acidic and the basic region of the pH gradient, where only few proteins could be detected. Since 80% of the luminal proteins were found in the acidic range of the pH gradient, we analyzed these proteins in more detail using a two-dimensional electrophoresis system with a linear pH gradient from 4 to 7. A typical two-dimensional map of the luminal proteins with isoelectric points in this pH range is shown in Fig. 1 B. An evaluation of the images of 10 independent experiments showed that ∼200 protein spots were present on each two-dimensional gel of the pH range from 4 to 7. Having established a reproducible two-dimensional map of the chloroplast lumen from Arabidopsis, we systematically analyzed the luminal proteins by both MALDI-TOF mass spectrometry and amino-terminal microsequencing. Using this combination of methods, we were able to determine the amino termini of the mature proteins and to correct errors in the gene models of several proteins. This approach could be successfully applied to 90 protein spots, but others of the 277 spots that were detected in all two-dimensional gels were too weak for an analysis by both mass spectrometry and microsequencing. In total, 49 proteins were detected, and each protein was identified in at least two independent experiments except for the three putative fibrillins with apparent masses at 25.5, 25.3, and 24.7 kDa. Although these proteins were analyzed only once, the identification was specific in each single case, and the corresponding protein spots were detected in all two-dimensional gels analyzed. Hence, there is no doubt that these proteins were correctly identified. The two-dimensional gel in Fig. 1 A shows that 40 of the 49 identified proteins were detected in the acidic range of the pH gradient, while nine proteins were found in the basic region. The analysis of the proteins by mass spectrometry and microsequencing showed that the major part of the proteins from the luminal fraction was intact. There were only few degradation products found. As Fig.1 B shows, there are two fragments of PsbP1 that were detected in all two-dimensional gels analyzed. In addition, we also detected a degradation product for PsaN, but this fragment occurred less frequently and did not appear in a distinct pattern as those from PsbP1. One of the principal objectives of this study was to identify a representative group of proteins from the chloroplast lumen of Arabidopsis and to find indications for their possible functions. Hence, it was important to confirm that the proteins that were identified in the luminal fraction from Arabidopsischloroplasts were truly lumen-located. The purification method alone was no proof for a luminal location. As Fig. 1 B shows, we detected among the luminal proteins a putative glyoxylase and the Cyp4 cyclophilin that are putative stroma proteins but were present in similar amounts as known luminal proteins such as TL29 and Hcf136. To resolve whether the proteins that were identified in the luminal fraction were resident in the chloroplast lumen or not, we performed a concise analysis of their transit peptides. All proteins that are targeted to the chloroplast lumen are synthesized in the cytosol as precursors and cross the chloroplast envelope and the thylakoid membrane in a two-stage import via a stromal intermediate. Accordingly, the signal peptides of luminal precursors comprise two parts, of which one is designed for the transit through the envelope and the second one for the import into the thylakoid lumen. Once the precursor of a luminal protein has reached the chloroplast stroma, the envelope transit region is cleaved off, and the intermediate is routed into the thylakoid lumen by either the Sec pathway or the ΔpH-dependent twin arginine translocation (Tat) pathway. While the Sec machinery is used by proteins that tolerate partial unfolding during the import, proteins that need to be imported in a folded state cross the thylakoid membrane via the Tat complex (37Robinson C. Thompson S.J. Woolhead C. Traffic. 2001; 2: 1-7Crossref Scopus (54) Google Scholar). The bipartite transit peptides for the import into the thylakoid lumen are a distinguishing feature of the luminal proteins. They are marked by a hydrophilic serine- and threonine-rich region for the transit through the envelope and a thylakoid targeting region with a typical hydrophobic core close to the processing site. In addition, the transit peptides of proteins that are routed by the Tat complex reveal a distinctive twin arginine motif in the beginning of the hydrophobic core region and a highly hydrophobic residue two or three positions after the twin arginine motif. By contrast, the transit peptides of proteins that are translocated by the Sec machinery do not have a twin arginine motif but a single lysine residue next to the amino-terminal end of the hydrophobic core region (37Robinson C. Thompson S.J. Woolhead C. Traffic. 2001; 2: 1-7Crossref Scopus (54) Google Scholar, 38Robinson C. Bolhuis A. Nat. Rev. Mol. Cell. Biol. 2001; 2: 350-356Crossref PubMed Scopus (145) Google Scholar). The transit peptides of all proteins that were identified in the luminal fraction of Arabidopsis chloroplasts were comprehensively analyzed whether or not they revealed the features of bipartite signal peptides. Of the 49 proteins that were identified in the luminal fraction, 35 had, indeed, a bipartite transit peptide. Fig.2 shows the thylakoid targeting regions of these transit peptides aligned with the program ClustalW. The alignment shows that all transit peptides possess a hydrophobic core; 19 signal peptides have a twin-arginine motif that marks them for translocation by the Tat pathway, and 16 have a lysine residue close to the hydrophobic core, which is a characteristic of signal peptides routed by the Sec machinery. Only the D1-processing protease appears to be an exception from this rule and has an arginine instead of lysine residue next to the amino-terminal end of the hydrophobic core. The only protein for which it was hard to decide whether the transit peptide really contained a targeting signal for the thylakoid lumen was the 20-kDa protein (Q9LXX5). The prepeptide of this protein had a rather indistinct hydrophobic core region and an arginine triplet instead of a conventional twin-arginine motif. To resolve this apparent conflict, we searched the expressed sequence tag databases for homologues and examined their transit peptides. The soybean clone Gm-c1032-2020 and the tomato clone cLET42E20 encoded for full-length homologues of the 20-kDa protein, and both had typical bipartite transit peptides with a plain twin arginine motif (not shown). This ensured that the prepeptide of the 20-kDa protein contained only an unusual variant of the twin arginine motif and, indeed, was a true bipartite transit peptide. In the case of the 19-kDa protein (P82658), no Arabidopsisgene was available that could be used for an examination of the signal peptide. It could only be mapped to the tentative consensus sequence TC115875 in the Arabidopsis Gene Index, which cannot be used to assess its subcellular location. Hence, we searched the expressed sequence tag databases and found several cDNAs from other plants that encoded the complete precursor of a homologue of the 19-kDa protein. Two representative examples were the tomato cDNA cTOF22B19 and the soybean cDNA Gm-c1013-3374. An analysis of the signal peptides of these homologues demonstrated clearly that they had all features of a bipartite transit peptide (not shown), which indicates that the 19-kDa protein from Arabidopsis is a lumen-located protein as well. The other 14 proteins that were identified in the luminal fraction from Arabidopsis came either from the chloroplast stroma such as the small subunit of ribulose-bisphosphate carboxylase or the thylakoid membrane such as the Ftsh protease or the α- and β-subunits of the ATP synthase. As for the four putative fibrillins and the 26-kDa protein, the subcellular location could not be predicted reliably, and, hence, it was left undecided. As Table I shows, the entire range of the luminal proteins of Arabidopsis that were identified in this work covers both well established classical proteins such as violaxanthin de-epoxidase and the extrinsic subunits of photosystem II, as well as novel proteins such as the large group of cyclophilins and FKBP-type peptidyl-prolyl cis-trans isomerases. In addition, there are many new proteins for which yet no function is known, such as, for instance, the PsbP domain proteins.Table IProteins from the chloroplast lumen of A. thalianaProtein nameGene locus (Tigr), accession (SP)IdentificationMass exper./theor.1-aExperimental/theoretical mass (kDa) is shown.Experimental NH2-terminal sequencekDaXanthophyll cycleViolaxanthin de-epoxidaseAT1g08550,Q39249MALDI, Micro44.2 /39.8VDALKTCACLLKGCRIELAKCIANPACAXNPhotosystem II assemblyH136_ARATHAT5g23120, O82660MALDI, MicroND /35.8DEQLSEWERVFLPIDPhotosystem II subunitsPSO1_ARATHAT5g66570, P23321MALDI, MicroND /26.6EGAPKRLTPSO2_ARATHAT3g50820, Q9S841MALDI, Micro24.7 /26.6EGAPKRLTPSP1_ARATHAT1g06680,Q42029MALDI, MicroND /20.2AYGEAANVFGKPKTNPSP2_ARATHAT2G30790, O49344Not detectedPSQ1_ARATHAT4g21280, Q9XFT3MALDI, MicroND /16.3DAISIKVGPPPAPSPSQ2_ARATHAT4g05180, Q41932MALDI, Micro15.3 /16.3EAIPIKVGGPPLPSProteins with a PsbP domain35.8 kDa proteinAT5g11450, P82715MALDI, Micro24.3 /25.6EEDVKMSGEELKMGTMVDDITL30_ARATHAT1g77090,O49292MALDI, Micro19.4 /22.2VVKQGLLAGRVPGLSEPDET215_ARATHAT4g15510, O23403MALDI, Micro21.5 /21.3STPVFREYIDTFDGYSFKYPTL26_ARATHAT3g55330,P82538MALDI, Micro16.3 /17.8AESKKGFLAVSDNKDAYAFLYPFGWQEV- VIEGQDKV20-kDa proteinAT3g56650, Q9LXX5MALDI, Micro20.0 /21.5REVEVGSYLPLSPSDPXFVL15.9-kDa proteinAT1g76450, Q9S720MALDI, Micro15.9 /15.8ETNASEAFRVYTDETNKFEISIPQPlastocyaninsPLAS_ARATHAT1g76100, P11490MicroND /10.5MEVLLGSDPLAT_ARATHAT1g20340, P42699Micro16.2 /10.5IEVLLGGGDGSLAFIPNDFSIAKGEKIVFPutative peroxidaseTL29_ARATHAT4g09010, P82281MALDI, Micro23.2 /29.3ADLNQRRQRSEFQSKIKILLSTTIKAKPELProteasesTail-specific proteasesCarboxyl-terminal proteinase D1-like proteinAT5g46390, Q9FL23MALDI, Micro49.0 /45.8ATNDPYLSD1-processing proteaseAT4g17740,O23614MALDI, Micro39.6 /41.9LTEENLLFXEASerine proteases, trypsin family36-kDa proteinAT5g39830, Q9LU10MALDI, PSD, Micro31.3 /37.5LGDPSVATVEDVSPTVFPAGPLFDegP protease (DEG1_ARATH)AT3g27925,1-bCorrected in this work.O22609MALDI, Micro31.7 /35.2FVVSTPKKLQTDELAHhoA proteaseAT4g18370,1-bCorrected in this work. Q9SEL7MALDI, Micro27.4 /23.5LEQFKEXEEXLPutative immunophilinsCyclophilin-type PPIases40-kDa proteinAT3g01480, Q9SSA5MALDI, PSD, Micro36.1 /38.2VLISGPPIKDPEALLRYALPID38-kDa proteinAT3g15520,1-bCorrected in this work.P82869MALDI, PSD, Micro38.0 /37.3VLYSPDTKVPRTGELALRRAIPAN18.5-kDa proteinAT5G13120,1-bCorrected in this work.Q9ASS6Micro18.5 /20.0NAEVVTEPQSKIFKBP-type PPIases18-kDa proteinAT5g13410, Q9LYR5MALDI, Micro18.0 /18.7SQFADMPALKGKDYGKTKMKYPDY17.8-kDa proteinAT1g20810, Q9LM71MALDI, Micro17.8 /17.9RERRSRKVIP17.5-kDa proteinAT2g43560,1-bCorrected in this work.O228701-bCorrected in this work.MALDI, Micro17.5 /15.7AGLPPEDKPRLCEAXCXKXL16.9-kDa proteinAT3g60370, Q9M222MALDI, Micro16.9 /16.4KTKSKSPYDERRLLEQN14.7-kDa proteinAT5g45680, Q9SCY2MALDI, Micro14.7 /13.6ETTSCEFSVSPSGLAFCDKVPentapeptide proteinsTL17_ARATHAT5g53490, P81760MALDI, Micro15.4 /17.4ANQRLPPLSTEPDR11.6-kDa proteinAT2g44920,O22160MALDI, Micro14.7 /11.5FKGGGPYGQGVTRGQDLSGKProteins with unknown function19-kDa proteinNot available, P82658Micro19.0 /NDEGNQTYKIYYGTAASAANYGG18.3-kDa protein" @default.
• W2090225411 created "2016-06-24" @default.
• W2090225411 creator A5019458580 @default.
• W2090225411 creator A5033472075 @default.
• W2090225411 creator A5036688322 @default.
• W2090225411 creator A5040823337 @default.
• W2090225411 creator A5068623714 @default.
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• W2090225411 date "2002-03-01" @default.
• W2090225411 modified "2023-10-16" @default.
• W2090225411 title "Proteome Map of the Chloroplast Lumen of Arabidopsis thaliana" @default.
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