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1.
The major light-harvesting complex in higher plants is LHC IIb. The LHC IIb of Arabidopsis thaliana contains 2 pigment-binding apoproteins of 28 and 25 kDa. To determine the relationship between them and the LHC IIb gene family members, each protein was purified to homogeneity, subjected to direct protein sequencing, and the sequences compared with those deduced from LHC IIb genes in this organism. The 28 kDa protein is the product of Type I LHC IIb genes. The 25 kDa LHC IIb component is distinctly different from the 28 kDa LHC IIb protein, and is more closely related to the type III LHC IIb gene product of barley. Type III gene products lack the first 9-11 residues found in proteins of the Type I and II genes, a region that contains a phosphorylatable threonine residue. The lack of the N-terminal residues explains why this LHC IIb apoprotein has never been seen to be phosphorylated, and partly or wholly why it is smaller. The implication of the missing N-terminus on uptake of LHC II precursor proteins into the plastid and of the relative organization of the LHC IIb subunits in the PS II antenna is discussed.  相似文献   

2.
3.
Monospecific antibodies directed against typical domains of type 1, 2, and 3 light-harvesting complex (LHC) II apoproteins have been used (a) to identify these apoproteins on denaturing sodium dodecyl sulfate gels of barley (Hordeum vulgare) thylakoids, (b) to determine their distribution between grana and stroma membranes, and (c) to follow their accumulation during light-induced greening of etioplasts. In addition, we have studied the light-induced assembly of chlorophyll-protein complexes with a native green gel system (K.D. Allen, L.A. Staehelin [1991] Anal Biochem 194: 214-222). Western blot analysis of the three major LHCII apoprotein bands has identified the highest molecular mass band at 27.5 kD as containing the type 2 LHCII apoproteins, the middle band at 26.9 kD as containing the type 1 LHCII apoproteins, and the lowest band at 26.0 kD as containing the type 3 LHCII apoproteins. During light-induced greening of 6-d-old etiolated barley seedlings, the type 1, 2, and 3 LHCII apoproteins accumulate simultaneously and at similar rates but appear somewhat sooner (< 4 h) in thylakoids from apical than from basal (4-8 h) leaf segments. LHCI polypeptides accrue with similar kinetics, whereas the 33-kD oxygen-evolving complex polypeptides can be detected already in the 0-h light samples. During the most rapid phase of thylakoid development (8-24 h), two slightly larger (28.3 and 28.7 kD) type 2 LHCII apoproteins (precursor intermediates?) also accumulate in the thylakoids. No corresponding higher molecular mass forms of type 1 and 3 LHCII apoproteins could be detected. It is interesting that differences are still apparent in the composition of chlorophyll-protein complexes of light-control plants and those of etiolated plants greened for 8 d.  相似文献   

4.
Allen KD  Staehelin LA 《Plant physiology》1992,100(3):1517-1526
The photosystem (PS) II antenna system comprises several biochemically and spectroscopically distinct complexes, including light-harvesting complex II (LHCII), chlorophyll-protein complex (CP) 29, CP26, and CP24. LHCII, the most abundant of these, is both structurally and functionally diverse. The photosynthetic apparatus is laterally segregated within the thylakoid membrane into PSI-rich and PSII-rich domains, and the distribution of antenna complexes between these domains has implications for antenna function. We report a detailed analysis of the differences in the polypeptide composition of LHCII, CP29, and CP26 complexes associated with grana and stroma thylakoid fractions from spinach (Spinacia oleracea L.), making use of a very high-resolution denaturing gel system, coupled with immunoblots using monospecific antibodies to identify specific antenna components. We first show that the polypeptide composition of the PSII antenna system is more complex than previously thought. We resolved at least five type I LHCII apoproteins and two to three type II LHCII apoproteins. We also resolved at least two apoproteins each for CP29 and CP26. In state 1-adapted grana and stroma thylakoid membranes, the spectrum of LHCII apoproteins is surprisingly similar. However, in addition to overall quantitative differences, we saw subtle but reproducible qualitative differences in the spectrum of LHCII apoproteins in grana and stroma membrane domains, including two forms of the major type II apoprotein. The implications of these findings for models of PSII antenna function in spinach are discussed.  相似文献   

5.
Immunoblotting was used to probe the reactivity of rabbit polyclonal antibodies against PS1I and PSI light-harvesting chlorophyll a/b-proteins of spinach ( Spinacea oleracea L.) with the light-harvesting complexes of a siphonaceous marine alga, Codium , that have more chlorophyll b, siphonaxanthin and siphonein instead of the lutein. The spinach LHCII antibodies cross-reacted only with the apoproteins of Cod-ium LHCII. Antisera against the spinach LHCI apoproteins showed strong affinity for the apoproteins of Codium LHCI, and also reacted with the polypeptides of spinach LHCII and Codium LHCII. Our results indicate some similarities in the amino acid sequences between the Codium siphonaxanthin-Chl a/fe-proteins of LHCII and LHCI and the corresponding spinach lutein-chlorophyll a/b-proteins.  相似文献   

6.
The main light-harvesting complex of Photosystem II (LHC II) in higher plants consists of two sub-populations. The 'inner' pool consists only of a 27 kDa polypeptide, whereas in the 'outer' pool both the 27 kDa and a 25 kDa polypeptide are found. We purified the 25 and the 27 kDa LHC II polypeptides from Scots pine and 25 kDa LHC II polypeptide from spinach. Protein sequencing after cleavage with endoproteinase Lys-C showed that the 25 kDa polypeptide is encoded by the Type 2 cab-II genes and the 27 kDa polypeptide by the Type I cab-II genes. A fatty acid was not covalently attached to the peptides assembled into the pigment-protein complex. Our results show that the different polypeptides seen on a gel are different gene products, and not the result of different processing.  相似文献   

7.
Three different pigment-binding proteins of the light-harvesting complex (LHC I) of maize photosystem I (PS I) have been isolated. Absorption and fluorescence excitation spectral analyses showed that each pigment-protein can transfer absorbed energy from its carotenoid and/or chlorophyll b components to chlorophyll alpha. Their apoproteins with apparent sizes of 24 (LHC Ia), 21 (LHC Ib), and 17 (LHC Ic) kDa have been purified to homogeneity. Differences in their pigment and amino acid compositions and in their reactions with antibodies demonstrate that the two smaller pigment-proteins are not proteolytically derived from the largest one. LHC Ib's apoprotein is particularly enriched in cysteine residues. None of the three apoproteins cross-reacted with antibodies raised against the major light-harvesting chlorophyll a/b-protein of photosystem II (LHC IIb) or against the PS I core complex (CC I) subunits. Studies of the biogenesis of PS I during greening of etiolated plants showed that all of the CC I subunits accumulated to a detectable level prior to the appearance of the 17-kDa subunit of LHC I, the accumulation of which preceded those of the 24- and 21-kDa subunits of LHC I. In addition, subunit VI of CC I is shown to be differentially expressed in mesophyll and bundle sheath cells; a slightly larger form of it accumulates in mesophyll than in bundle sheath thylakoids during plastid development.  相似文献   

8.
The structure and heterogeneity of LHC II were studied by in vitro reconstitution of apoproteins with pigments (Plumley and Schmidt 1987, Proc Natl Acad Sci 84: 146–150). Reconstituted CP 2 complexes purified by LDS-PAGE were subsequently characterized and shown to have spectroscopic properties and pigment-protein compositions and stoichiometries similar to those of authentic complexes. Heterologous reconstitutions utilizing pigments and light-harvesting proteins from spinach, pea and Chlamydomonas reinhardtii reveal no evidence of specialized binding sites for the unique C. reinhardtii xanthophyll loroxanthin: lutein and loroxanthin are interchangeable for in vitro reconstitution. Proteins modified by the presence of a transit peptide, phosphorylation, or proteolytic removal of the NH2-terminus could be reconstituted. Evidence suggests that post-translational modification are not responsible for the presence of six electrophoretic variants of C. reinhardtii CP 2. Reconstitution is blocked by iodoacetamide pre-treatment of the apoproteins suggesting a role for cysteine in pigment ligation and/or proper folding of the pigment-protein complex. Finally, no effect of divalent cations on pigment reassembly could be detected.Abbreviations cab chlorophyll a/b-binding protein genes - Chl chlorophyll - CP2 light-harvesting chlorophyll A+b-protein complex fractionated by mildly denaturing LDS-PAGE from Photosystem II in thylakoids - CP 43 and CP 47 chlorophyll a-antenna complexes fractionated from Photosystem II in thylakoids by mildly denaturing LDS-PAGE at 4°C - IgG gamma immunoglobulin - LDS lithium dodecyl sulfate - LDS-PAGE lithium dodecyl sulfate polyacrylamide gel electrophoresis at 4°C - LHC I and LHC II thylakoid light-harvesting chlorophyll a+b-protein holocomplexes associated with Photosystems I and II, respectively - PS II Photosystem II - TX100 Triton X-100 - TX100-derived LHC light-harvesting complexes enriched in LHC II following fractionation of thylakoids by TX100  相似文献   

9.
We investigated the primary structure of a cDNA encoding a light-harvesting protein from the marine chrysophyteIsochrysis galbana. Antibodies raised against the major fucoxanthin, chlorophylla/c-binding light-harvesting protein (FCP) ofI. galbana were used to select a cDNA clone encoding one of the FCP apoproteins. The nucleic acid and deduced amino acid sequences reveal conserved regions within the first and third transmembrane spans with Chla/b-binding proteins and with FCPs of another chromophyte. However, the amino acid identity betweenI. galbana FCP and othercab genes of FCPs is only ca. 30%. Phylogenetic analyses demonstrated that the FCP genes of both diatoms and chrysophytes sequenced to date are more closely related tocab genes encoding LHC I, CP 29, and CP 24 of higher plants than tocab genes encoding LHC II of chlorophytes. We propose that LHC I, CP 24 and CP 29 and FCP might have originated from a common ancestral chl binding protein and that the major LHC II of Chla/b-containing organisms arose after the divergence between the chromophytes and the chlorophytes.  相似文献   

10.
The light-induced assembly of light-harvesting complex (LHC) II has been followed during the biogenesis of the plastid. Seedlings grown in intermittent light (IML) accumulate only small amounts of chlorophyll b. The minor LHC II apoproteins are present; however, the apoprotein levels of the major LHC II complex, LHC IIb, are severely depressed after exposure to IML. The levels of all LHC II apoproteins increase rapidly upon exposure to continuous illumination. The 25-kD, type 3 LHC IIb subunit appears to be more abundant during the early hours of greening in relation to its level in mature thylakoids. The LHC IIb apoproteins are initially associated with pigments to form monomeric pigment-protein complexes. The abundance of monomeric LHC IIb complexes gradually decreases during exposure to continuous light and a concomitant increase occurs in the amount of the trimeric and higher-order oligomeric forms. Pulse-chase experiments verify that labeled LHC IIb monomeric complexes are intermediates in the formation of trimeric and higher-order oligomeric LHC IIb-pigmented complexes. Therefore, the assembly of LHC II occurs via the initial pigmentation of the apoproteins to form monomeric complexes and proceeds in a sequential manner.  相似文献   

11.
Summary cDNAs encoding three different LHC I polypeptides (Type I, Type II and Type III) from the gymnosperm Scots pine (Pinus sylvestris L.) were isolated and sequenced. Comparisons of the deduced amino acid sequences with the corresponding tomato sequences showed that all three proteins were highly conserved although less so than the LHC II proteins. The similarities between mature Scots pine and tomato Types I, II and III LHC I proteins were 80%, 87% and 85%, respectively. Two of the five His residues that are found in AXXXH sequences, which have been identified as putative chlorophyll ligands in the Type I and Type II proteins, were not conserved. The same two regions of high homology between the different LHC proteins, which have been identified in tomato, were also found in the Scots pine proteins. Within the conserved regions, the Type I and Type II proteins had the highest similarity; however, the Type II and Type III proteins also showed a similarity in the central region. The results suggest that all flowering plants (gymnosperms and angiosperms) probably have the same set of LHC polypeptides. A new nomenclature for the genes encoding LHC polypeptides (formerly cab genes) is proposed. The names lha and lhb are suggested for genes encoding LHC I and LHC II proteins, respectively, analogous to the nomenclature for the genes encoding other photosynthetic proteins.  相似文献   

12.
Photosystem I (PSI) holocomplexes were fractionated to study the organization of the light-harvesting complex I (LHC I) pigment-proteins in barley (Hordeum vulgare) plastids. LHC Ia and LHC Ib can be isolated as oligomeric, presumably trimeric, pigment-protein complexes. The LHC Ia oligomeric complex contains both the 24- and the 21.5-kD apoproteins encoded by the Lhca3 and Lhca2 genes and is slightly larger than the oligomeric LHC Ib complex containing the Lhca1 and Lhca4 gene products of 21 and 20 kD. The synthesis and assembly of LHC I during light-driven development of intermittent light-grown plants occurs rapidly upon exposure to continuous illumination. Complete PSI complexes are detected by nondenaturing Deriphat (disodium N-dodecyl-[beta]-iminodipropionate-160)-PAGE after 2 h of illumination, and their appearance correlates with that of the 730- to 740-nm emission characteristic of assembled LHC I. However, the majority of the newly synthesized LHC I apoproteins are present as monomeric complexes in the thylakoids during the early hours of greening. We propose that during development of the protochloroplast the LHC I apoproteins are first assembled into monomeric pigmented complexes that then aggregate into trimers before becoming attached to the pre-existing core complex to form a complete PSI holocomplex.  相似文献   

13.
Monospecific polyclonal antibodies have been raised against synthetic peptides derived from the primary sequences from different plant light-harvesting Chl a/b-binding (LHC) proteins. Together with other monospecific antibodies, these were used to quantify the levels of the 10 different LHC proteins in wild-type and chlorina f2 barley (Hordeum vulgare L.), grown under normal and intermittent light (ImL). Chlorina f2, grown under normal light, lacked Lhcb1 (type I LHC II) and Lhcb6 (CP24) and had reduced amounts of Lhcb2, Lhcb3 (types II and III LHC II), and Lhcb4 (CP 29). Chlorina f2 grown under ImL lacked all LHC proteins, whereas wild-type ImL plants contained Lhcb5 (CP 26) and a small amount of Lhcb2. The chlorina f2 ImL thylakoids were organized in large parallel arrays, but wild-type ImL thylakoids had appressed regions, indicating a possible role for Lhcb5 in grana stacking. Chlorina f2 grown under ImL contained considerable amounts of violaxanthin (2-3/reaction center), representing a pool of phototransformable xanthophyll cycle pigments not associated with LHC proteins. Chlorina f2 and the plants grown under ImL also contained early light-induced proteins (ELIPs) as monitored by western blotting. The levels of both ELIPs and xanthophyll cycle pigments increased during a 1 h of high light treatment, without accumulation of LHC proteins. These data are consistent with the hypothesis that ELIPs are pigment-binding proteins, and we suggest that ELIPs bind photoconvertible xanthophylls and replace "normal" LHC proteins under conditions of light stress.  相似文献   

14.
Seedlings of wheat (Triticum aestivum L. cv. Walde, Weibull) grown in continuous weak red light (16 mW m−2) with or without SAN-9789, contained significantly lower amounts of chlorophylls and carotenoids compared to untreated plants grown in a greenhouse. The Chl alb ratios were 3.6 in the greenhouse-grown plants, 5.1 in untreated and ca 16 in SAN-treated plants grown in weak red light, respectively. The main difference in polypeptide composition of thylakoids isolated from red light-grown plants, compared to those grown in the greenhouse, was a lower amount of proteins of the light-harvesting complex (LHC) II. The amount of apo-LHC and LHC were correlated to the xanthophyll to β-carotene ratios in these plants. The absence of grana and the absence of proteins of the light-harvesting complex 11 in SAN-treated plants, support the general dogma that these proteins are involved in the formation of grana. Since the amount of apo-LHC and LHC could be correlated to the presence of carotenoids as well as the chlorophylls, it is concluded that the carotenoids are necessary for the correct assembly and stabilization of the apoproteins of LHC II in the thylakoid membranes.  相似文献   

15.
The 20 kDa polypeptide, the apoprotein of the chlorophyll a/b antenna complex CP24 associated with photosystem II, is a remote relative of light-harvesting complex (LHC) apoproteins and thus a member of the extended cab gene family. LHC apoproteins are poly-topic integral components of the thylakoid membrane with probably three transmembrane segments which originate in nuclear genes and are made in the cytosol as precursors. They possess exclusively stroma-targeting transit peptides for import into the organelle and integrate into the thylakoid membrane via uncleaved hydrophobic domains of the mature protein. The CP24 apoprotein displays intriguing structural differences to LHC apoproteins with a potential impact on the routing and targeting processes during biogenesis. In particular, it lacks a pronounced second hydrophobic segment in the mature polypeptide chain found in LHCPs, and carries a transit peptide that is reminiscent of thylakoid-targeting transit peptides. We have used in organello assays with isolated intact chloroplasts and the authentic precursor of the 20 kDa apoprotein from spinach, or appropriate chimaeric polypeptides consisting of a transit peptide and the mature part of various nuclear-encoded thylakoid proteins of known location and targeting epitopes, in order to resolve the characteristics of its targeting properties, as well as to determine the contribution of the individual parts of the precursor molecule to its import and subsequent intra-organellar routing. Our experiments demonstrate that the transit peptide of the CP24 apoprotein is required only for the import of the protein into the organelle. All subsequent steps, such as the integration of the protein into the thylakoid membrane, binding of chlorophyll, assembly into the CP24 complex and migration to the grana lamellae, still take place if the authentic transit peptide is replaced by a targeting signal of a nuclear-encoded stromal protein.  相似文献   

16.
The marine chlorophyte Dunaliella tertiolecta Butcher responds to a one-step transition from a high growth irradiance level (700 micromoles quanta per square meter per second) to a low growth irradiance level (70 micromoles quanta per square meter per second) by increasing the total amount of light-harvesting chlorophyll (Chl) a/b binding protein associated with photosystem II (LHC II), and by modifying the relative abundance of individual LHC II apoproteins. When high light-adapted cells were incubated with gabaculine, which inhibits Chl synthesis, and transferred to low light, the LHC II apoproteins were still synthesized and the 35S-labeled LHC II apoproteins remained stable after a 24 hour chase. These results suggest that Chl synthesis is not required for stability of the LHC II apoproteins in this alga. However, when the control cells are transferred from high light to low light, the amount of the four LHC II apoproteins per cell increases, whereas it does not in the presence of gabaculine. These results suggest that Chl synthesis is required for a photoadaptive increase in the cellular level of LHC II.  相似文献   

17.
The soybean light-harvesting complex II (LHC II) was composed of one major and three minor chlorophyll a/b (Cab) binding proteins. This study demonstrated that the soybean genome contained at least 11 genes that code for these Cab proteins. Three members of the soybean Cab gene family were characterized. Cab 3 coded for a 25.7 kD mature apoprotein with a 32 amino acid transit peptide. Comparisons with previously published Cab protein sequences indicated that Cab 3 coded for the major Cab protein of LHC II. Cab 2 coded for a novel Cab protein with an apparent molecular weight of 24.6 kD. Cab 2 retained a high degree of similarity with Cab 3, but distinguished itself from previously reported minor photosystem II type II Cab genes and products. Finally, Cab 1 was determined to be a pseudogene that had two deletions relative to Cab 2 and Cab 3.  相似文献   

18.
The high selectivity offered by reversed-phase high-performance liquid chromatography on-line coupled to electrospray ionization mass spectrometry has been utilized to characterize the major and minor light-harvesting proteins of photosystem II (Lhcb). Isomeric forms of the proteins, revealed either on the basis of different hydrophobicity enabling their chromatographic separation or on the basis of different molecular masses identified within one single chromatographic peak, were readily identified in a number of monocot and dicot species. The presence of several Lhcb1 isoforms (preferably in dicots) can explain the tendency of dicot Lhcb1 to form trimeric aggregates. The Lhcb1 molecular masses ranged from 24,680 to 25,014 among different species, whereas within the same species, the isoforms differed by 14-280 mass units. All Lhcb1 proteins appear to be highly conserved among different species such that they belong to a single gene group that has several different gene family members. In all species examined, the number of isoforms corresponded more or less to the genes cloned previously. Two isoforms of Lhcb3 were found in petunia and tomato. For Lhcb6, the most divergent of all light-harvesting proteins, the greatest number of isoforms was found in petunia, tobacco, tomato, and rice. Lhcb2, Lhcb4, and Lhcb5 were present in only one form. The isoforms are assumed to play an important role in the adaptation of plants to environmental changes.  相似文献   

19.
 A fucoxanthin-chlorophyll protein (FCP) cDNA from the raphidophyte Heterosigma carterae encodes a 210-amino acid polypeptide that has similarity to other FCPs and to the chlorophyll a/b-binding proteins (CABs) of terrestrial plants and green algae. The putative transit sequence has characteristics that resemble a signal sequence. The Heterosigma fcp genes are part of a large multigene family which includes members encoding at least two significantly different polypeptides (Fcp1, Fcp2). Comparison of the FCP sequences to the recently determined three-dimensional structure of the pea LHC II complex indicates that many of the key amino acids thought to participate in the binding of chlorophyll and the formation of complex-stabilizing ionic interactions are well conserved. Phylogenetic analyses of sequences of light-harvesting proteins shows that the FCPs of several chromophyte phyla form a natural group separate from the intrinisic peridinin-chlorophyll proteins (iPCPs) of the dinoflagellates. Although the FCP and CAB genes shared a common ancestor, these lineages diverged from each other prior to the separation of the CAB LHC I and LHC II sequences in the green algae and terrestrial plants. Received: 8 July 1996 / Accepted: 21 August 1996  相似文献   

20.
The light-harvesting complex of photosystem II (LHC II) contains one major (LHC IIb) and at least three minor chlorophyll-protein components. The apoproteins of LHC IIb (LHCP) are encoded by nuclear genes and synthesized in the cytoplasm as a higher molecular weight precursor(s) (pLHCP). Several genes coding for pLHCP have been cloned from various higher plant species. The expression of these genes is dependent upon a variety of factors such as light, the developmental stage of the plastids and the plant. After its synthesis in the cytoplasm, pLHCP is imported into plastids, inserted into thylakoids, processed to its mature form, and assembled into LHC IIb. The pathway of assembly of LHC IIb in the thylakoid membranes is currently being investigated in several laboratories. We present a model that gives some details of the steps in the assembly process. Many of the steps involved in the synthesis and assembly are dependent on light and the stage of plastid development.Abbreviations PS Photosystem - LHC II Light-harvesting complex of PS II - LHCP Apoproteins of LHC IIb - pLHCP Precursor of LHCP - PAGE Polyacrylamide gel electrophoresis  相似文献   

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