Chlorophyll-protein complex composition and photochemical activity in developing chloroplasts from greening barley seedlings

The time course for the observation of intact chlorophyll-protein (CP) complexes during barley chloroplast development was measured by mild sodium dodecyl sulfate polyacrylamide gel electrophoresis. The procedure required extraction of thylakoid membranes with sodium bromide to remove extrinsic proteins. During the early stages of greening, the proteins extracted with sodium bromide included polypeptides from the cell nucleus that associate with developing thylakoid membranes during isolation and interfere with the separation of CP complexes by electrophoresis. Photosystem I CP complexes were observed before the photosystem II and light-harvesting CP complexes during the initial stages of barley chloroplast development. Photosystem I activity was observed before the photosystem I CP complex was detected whereas photosystem II activity coincided with the appearance of the CP complex associated with photosystem II. Throughout chloroplast development, the percentage of the total chlorophyll associated with photosystem I remained constant whereas the amount of chlorophyll associated with photosystem II and the light-harvesting complex increased. The CP composition of thylakoid membranes from the early stages of greening was difficult to quantitate because a large amount of chlorophyll was released from the CP complexes during detergent extraction. As chloroplast development proceeded, a decrease was observed in the amount of chlorophyll released from the CP complexes by detergent action. The decrease suggested that the CP complexes were stabilized during the later stages of development.Abbreviations Chl chlorophyll - CP chlorophyll-protein - CPI P700 chlorophyll-a protein complex of photosystem I - CPa electrophoretic band that contains the photosystem II reaction center complexes and a variable amount of the photosystem I light-harvesting complex - CP A/B the major light-harvesting complex associated with photosystem II - DCIP 2,6-dichlorophenolindophenol - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DPC diphenyl carbazide - MV methyl viologen - PAR photosynthetically active radiation - PSI photosystem I - PSII photosystem II - SDS sodium dodecyl sulfate - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis - TEMED N,N,N,N-tetramethylethylenediamine - TMPD N,N,N,N-tetramethyl-p-phenylenediamine Cooperative investigations of the United States Department of Agriculture, Agricultural Research Service, and the North Carolina Agricultural Research Service, Raleigh, NC 27695-7601. Paper No. 9949 of the Journal Series of the North Carolina Agricultural Research Service, Raleight, NC 27695-7601.     

Chlorophyll-protein complex composition during chloroplast development: A species comparison

Barley, maize, pea, soybean, and wheat exhibited differences in chlorophyll a/b ratio and chlorophyll-protein (CP) complex composition during the initial stages of chloroplast development. During the first hours of greening, the chlorophyll a/b ratios of barley, pea, and wheat were high (a/b8) and these species contained only the CP complex of photosystem I as measured by mild sodium dodecyl sulfate polyacrylamide gel electrophoresis. A decrease in chlorophyll a/b ratio and the observation of the CP complexes associated with photosystem II and the light-harvesting apparatus occurred at later times in barley, pea, and wheat. In contrast, maize and soybean exhibited low chlorophyll a/b ratios (a/b<8) and contained the CP complexes of both photosytem I and the light-harvesting apparatus at early times during chloroplast development. The species differences were not apparent after 8 h of greening. In all species, the CP complexes were stabilized during the later stages of chloroplast development as indicated by a decrease in the percentage of chlorophyll released from the CP complexes during detergent extraction. The results demonstrate that CP complex synthesis and accumulation during chloroplast development may not be regulated in the same way in all higher plant species.Abbreviations Chl chlorophyll - CP chlorophyll-protein - CPI P700 chlorophyll-a protein complex of photosystem I - CPa electrophoretic band that contains the photosystem II reaction center complexes and a variable amount of the photosystem I light-harvesting complex - LHC the major light-harvesting complex associated with photosystem II - PSI photosystem I - PSII photosystem II - SDS sodium dodecyl sulfate - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis Cooperative investigations of the United States Department of Agriculture, Agricultural Research Service, and the North Carolina Agricultural Research Service, Raleigh, NC 27695-7601. Paper No. 10335 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh, NC 27695-7601.     

Isolation of chlorophyll-protein complexes and quantification of electron transport components in Synura petersenii and Tribonema aequale

The chlorophyll-protein complexes of the yellow alga Synura petersenii (Chrysophyceae) and the yellow-green alga Tribonema aequale (Xanthophyceae) were studied. The sodiumdodecylsulfate/sodiumdesoxycholate solubilized photosynthetic membranes of these species yielded three distinct pigment-protein complexes and a non-proteinuous zone of free pigments, when subjected to SDS polyacrylamid gel electrophoresis. The slowest migrating protein was identical to complex I (CP I), the P-700 chlorophyll a-protein, which possessed 60 chlorophyll a molecules per reaction center in Tribonema and 108 in Synura. The zone of intermediate mobility contained chlorophyll a and carotenoids. The absorption spectrum of this complex was very similar to the chlorophyll a-protein of photosystem II (CP a), which is known from green plants. The fastest migrating pigment protein zone was identified as a light-harvesting chlorophyll-protein complex. In Synura this protein was characterized by the content of chlorophyll c and of fucoxanthin. Therefore this complex will be named as LH Chl a/c-fucocanthin protein. In addition to the separation of the chlorophyll-protein complexes the cellular contents of P-700, cytochrome f (bound cytochrome) and cytochrome c-553 (soluble cytochrome) were measured. The stoichiometry of cytochrome f: cytochrome c-553:P-700 was found to be 1:4:2.4 in Tribonema and 1:6:3.4 in Synurá.Abbreviations CP a chlorophyll a-protein of photosystem II - CP I P-700 chlorophyll a-protein - FP free pigment - LH Chl a/c light-harvesting chlorophyll a/c-protein - PAGE polyacrylamidgelelectrophoresis - SDS Sodiumdodecylsulfate - SDOC sodium-desoxycholate     

NITROGEN LIMITATION IN ISOCHRYSIS GALBANA (HAPTOPHYCEAE). II. RELATIVE ABUNDANCE OF CHLOROPLAST PROTEINS1

Using spectroscopic, biophysical and immunological techniques, we assayed the relative abundance often chloroplast proteins and protein complexes in the marine haptophyte, Isochrysis galbana Green, grown at nine steady-state dilution rates in nitrogen-limited chemostats. The proteins included Photosystem I reaction center (RCI) chlorophyll protein, CP1; Photosystem II reaction center (RC II) protein, D1; two chlorophyll a-binding apoproteins, CP 43 and CP 47; 33 KDa oxygen evolving protein, OEC 33; α subunit of coupling factor, CF1α; large (LSU) and small subunits (SSU) of ribulose 1,5-bisphosphate carboxylase, RuBisCO; the chlorophyll a/c/fucoxanthin protein complex, LHCP; and cytochrome b₆/f. Seven of the ten protein complexes are encoded in the chloroplast, two are encoded in the nucleus and one shares chloroplast and nuclear genomes. Over the range of dilution rates (0.96-0.18 d^?1) cell N decreased 42% and cellular chlorophyll a decreased 50%; however, the stoichiometric proportion of RC II: cytochrome b₆/f: RC I remained constant, averaging 1:3.3:0.8. In contrast, RuBisCO / PS II decreased by 58%. The light harvesting chlorophyll a/c/fucoxanthin protein complex increased relative to RC II; however, as cells became more nitrogen limited the fraction of total cell nitrogen contained in RuBisCO decreased from 21.3 to 6.7%, whereas that of the light harvesting complex remained relatively constant, averaging 6.8%. Our results generally support the hypothesis that in nitrogen limited cells, proteins encoded in the nuclear genome are synthesized preferentially over those encoded in the chloroplast.     

Energy transfer in the light-harvesting complex II of Dunaliella tertiolecta is unusually sensitive to Triton X-100

Triton X-100, a detergent commonly used to solubilize higher plant thylakoid membranes, was found to be deleterious to Dunaliella LHC II. It disrupted the transfer of excitation energy from chlorophyll b to chlorophyll a. Based on analysis of pigments and immunoassays of LHC II apoproteins from sucrose density gradient fractions, Triton X-100 caused aggregation of the complex, but apparently did not remove chlorophyll b from the apoprotein. Following solubilization with Triton X-100 only CPI could be resolved by electrophoresis. In contrast, solubilization of Dunaliella thylakoids with octyl--D-glucopyranoside preserved energy transfer from chlorophyll b to chlorophyll a. This detergent also effectively prevented aggregation on sucrose gradients and preserved CPI oligomers, as well as LHCP1 and LHCP3 on non-denaturing gels. Solubilization with Deriphat gave similar results. We propose that room temperature fluorescence excitation and emission spectroscopy be used in conjunction with other biophysical and biochemical probes to establish the effects of detergents on the integrity of light harvesting chlorophyll protein complexes. Methods used here may be applicable to other chlorophytes which prove refractory to protocols developed for higher plants.Abbreviations LHC II light harvesting chlorophyll protein complex associated with photosystem II - LHCP1 and LHCP3 monomeric and oligomeric forms of LHC II, respectively, observed on non-denaturing gels - LiDS lithium dodecylsulphate - PMSF phenylmethylsulfonyl fluoride     

Properties of protein-chlorophyll complexes from pea (Pisum sativum L.) leaves. The organization of chlorophyll.

Chlorophyll-protein-detergent complexes were prepared from pea chloroplasts by using sodium dodecylbenzenesulphonate and polyacrylamide-gel electrophoresis. Circular-dichroism spectra showed that complex CPI has a dimeric arrangement of chlorophyll a, with additional weaker interactions. Ellipticities were determined for both complexes and for purified chlorophylls in solution, and it is argued that the circular dichroism of complex CPII is derived from chlorophyll-protein interaction rather than from interaction between chlorophylls a and b. The detergent could be removed from the complexes by using urea and gel filtration, leaving the chlorophyll-protein in solution, although in each case a diminished ellipticity indicated some loss of organization. Three-peaked circular-dichroism spectra of chloroplast fragments before and after addition of detergent were compared with a curve obtained by summing graphically the spectra of complexes CPI, CPII and the free-pigment fraction. There was good correspondence at 650 nm, and the longer-wavelength peaks agreed in form and magnitude, but with discrepancies in position. It was concluded that complexes CPI and CPII pre-exist in the original material, but that there is an environmental effect which is destroyed when the complexes are extracted.     

Chlorophyll-protein and polypeptide composition of Mn-deficient sugar beet thylakoids

The chlorophyll-protein and polypeptide composition of manganese deficient and control sugar beet thylakoids was examined using three different detergent-electrophoresis systems. On a per chlorophyll basis, manganese deficiency reduced the amounts of CPa complex (separated by sodium dodecylsulfate (SDS)-polyacrylamide gel electrophoresis), and CP 47 and CP 43 complexes (separated by octylglucoside/SDS-polyacrylamide gel electrophoresis) without decreasing the amounts of light harvesting complexes. Lithium dodecylsulfate/Triton X-100 polyacrylamide gel electrophoresis showed that manganese deficiency decreased several thylakoid polypeptides, including a chlorophyll b containing 30 kilodalton chlorophyll-protein complex, but did not decrease the amounts of 28 and 29 kilodalton light-harvesting chlorophyll b-containing polypeptides.     

Are specific chlorophyll-protein complexes required for photosynthesis?

Using analytical polyacrylamide gel electrophoresis of detergent extracts of Scenedesmus chloroplast fragments, we have shown that the electrophoresis pattern of Bishop's mutant 8 (which lacks a functional Photoreaction I) differs from the pattern shown by the wild-type and mutant II. Gels from mutant 8 extracts do not show a band corresponding to the chlorophyll-protein-detergent complex I nor is a corresponding protein band present. Thus chlorophyll-protein-detergent complex I reflects solubilization of a real and essential chlorophyll-protein complex of System I of photosynthesis, and lack of a functional System I in this mutant is due to absence of this chlorophyll-protein complex. This implies that at least this chlorophyll-protein complex is required for photosynthesis.     

Intrinsic membrane proteins of the thylakoids of Chlamydomonas reinhardii

Upon protoolytle treatment of thylakoid membranes, extrinsic proteins are selectively degraded. The remaining resistant proteins have been analyzed by SDS-PAGE. In the thylakoids of Chlamydomonas reinhardii, six polypeptides or protein fragments of 20 kD or higher are resistant to proteolysis. These intrinsic proteins have been identified as: the apoproteins of the chlorophyll-protein complexes CP I and LHCP; a polypeptide whose presence is related to the chlorophyll b content of the cells; and a portion of a chloroplast-made 34 kD polypeptide. Furthermore, after proteolytic treatment of the membranes, the LHCP complex can be resolved into two subcomplexes, apparently differing in their polypeptide composition.

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Zusammenfassung Durch proteolytische Behandlung von Thylakoidmembranen worden die extrinsischen Proteinanteile selektiv abgebaut. Die resistenten, in der Membran verbleibenden Polypeptide können mit SDS-PAGE analysiert werden. In Thylakoiden von Chlamydomonas reinhardii finden sich sechs resistente Polypeptide mit Molekulargewichten über 20'000. Durch quantitative Bestimmung während der Proteolyse und durch limitierte Fragmentierung nach Cleveland wurden die resistenten Polypeptide identifiziert als: (a) Apoprotein von CP 1, (b) Apoproteine des LHCP-Komplexes, (c) Teil des chloroplastidial synthetisierten 34 kD-Proteins und (d) Polypeptid, welches möglicherweise mit dem Chlorophyll b-Gehalt in Verbindung steht. Im weiteren führte die proteolytische Behandlung der Thylakoide zu einer Auftrennung des LHCP-Komplexes in zwei Subkomplexe mit unterschiedlicher Proteinzusammensetzung.

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Abbreviations CP 1 pigment-protein complex with slower mobility in SDS electrophoresis, corresponding to the P700 chlorophyll a protein complex - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl urea - EDTA ethylene-diaminotetraacetic acid - kD kilo Dalton - LHCP pigment-protein complex with intermediate mobility in SDS electrophoresis, corresponding to the light-harvesting chlorophyll a/b protein complex - SDS sodiumdodecyl sulfate - SDS-PAGE electrophoresis in SDS-containing polyacrylamide gels - Tris tris (hydroxymethyl) aminomethane     

Multiple forms of chlorophyll-protein complexes from a thermophilic cyanobacterium Synechococcus sp

Eight chlorophyll-proteins were resolved from the thylakoid membranes, or digitonin particles, of a thermophilic cyanobacterium Synechococcus sp. by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Six chlorophyll-proteins with slower electrophoretic mobilities were shown to be P700-chlorophyll a-protein complexes (CP1), whereas faster-moving proteins (CP2) were related to photosystem 2. Extraction of CP1 complexes from the membranes with different detergent/chlorophyll ratios and reelectrophoresis of extracted CP1 complexes indicated that the chlorophyll-proteins are closely interrelated with each other; any CP1 complex could be transformed to other CP1 complexes with faster electrophoretic mobilities. This, together with the Ferguson plot and the polypeptide composition, showed that six CP1 complexes are different in terms of polypeptide composition, oligomerization, SDS-binding, or conformation of the proteins but represent, in the order of increasing electrophoretic mobility, increasing degree of modification of the native P700-chlorophyll a-protein.     

The action of lipases on chloroplast membranes. III. The effect of lipid hydrolysis on chlorophyll-protein complexes in thylakoid membranes

Bean thylakoid membranes treated with various lipolytic enzymes (bean galactolipase, phospholipases A₂, C, D) showed marked changes in their acyl lipid composition. As a consequence of acyl lipids hydrolysis, destruction of some chlorophyll a-protein complexes (CP1a, CP1, CPa) or monomerization of the oligomeric of light harvesting chlorophyll a/b protein complex (LHCP) was observed. It is concluded that galactolipids and phosphatidylcholine are responsible for the stability of CP1a, CP1 and CPa, respectively. Phosphatidylglycerol and to some extent monogalactosyldiacylglycerol are essential for the stabilization of oligomeric structures of light harvesting chlorophyll a/b protein complex.Abbreviations chl chlorophyll - CP1a, CP1 chl a-protein complexes, of PSI - CPa chl a-protein complex of PSII - DGDG diagalactosyldiacylglycerol - FC free chl - GL galactolipase - LHCP^1–3 light harvesting chl a/b protein complex - MGDG monogalactosyldiacylglycerol - PAGE polyacrylamide gel electrophoresis - PC phosphatidylcholine - PG phosphatidylglycerol - PLA₂ phospholipase A₂ - PL phospholipase C - PLD phospholipase D - PSI photosystem I - PSII photosystem II - SDS sodium dodecyl sulphate - SQDG sulfoquinovosyl-diacylglycerol - TCA trichloroacetic acid - Tricine N-tris-(hydroxymethyl)-methylglycine - Tris Tris-(hydroxymethyl)-aminomethan     

Isolation of chlorophyll-protein complexes and quantification of electron transport components in Synura petersenil and Tribonema aequale

The chlorophyll-protein complexes of the yellow alga Synura petersenii (Chrysophyceae) and the yellow-green alga Tribonema aequale (Xanthophyceae) were studied. The sodiumdodecylsulfate/sodiumdesoxycholate solubilized photosynthetic membranes of these species yielded three distinct pigment-protein complexes and a non-proteinous zone of free pigments, when subjected to SDS polyacrylamid gel electrophoresis. The slowest migrating protein was identical to complex I (CP I), the P-700 chlorophyll a-protein, which possessed 60 chlorophyll a molecules per reaction center in Tribonema and 108 in Synura. The zone of intermediate mobility contained chlorophyll a and carotenoids. The absorption spectrum of this complex was very similar to the chlorophyll a-protein of photosystem II (CP a), which is known from green plants. The fastest migrating pigment protein zone was identified as a light-harvesting chlorophyll-protein complex. In Synura this protein was characterized by the content of chlorophyll c and of fucoxanthin. Therefore this complex will be named as LH Chl a/c-fucocanthin protein. In addition to the separation of the chlorophyll-protein complexes the cellular contents of P-700, cytochrome f (bound cytochrome) and cytochrome c-553 (soluble cytochrome) were measured. The stoichiometry of cytochrome f: cytochrome c-553:P-700 was found to be 1:4:2.4 in Tribonema and 1:6:3.4 in Synurá.     

Preferential accumulation of apoproteins of the light-harvesting chlorophyll a/b-protein complex in greening barley leaves treated with 5-aminolevulinic acid

In order to study the coordinate accumulation of chlorophyll (Chl) and apoproteins of Chl-protein complexes (CPs) during chloroplast development, we examined changes in the accumulation of the apoproteins in barley (Hordeum vulgare L.) leaves when the rate of Chl synthesis was altered by feeding 5-aminolevulinic acid (ALA), a precursor of Chl biosynthesis. Pretreatment with ALA increased the accumulation of Chl a and Chl b 1.5- and 2.3-fold, respectively, after 12 cycles of intermittent light (2 min light followed by 28 min darkness). Apoproteins of the light-harvesting Chl a/b-protein complex of photosystem II (LHCII) were increased 2.4-fold with ALA treatment. However, apoproteins of the P700-Chl a-protein complex (CP1) and the 43-kDa apoprotein of a Chl a-protein complex of photosystem II (CPa) were not increased by ALA application. With respect to CPs themselves, LHCII was increased when Chl synthesis was raised by ALA feeding, whereas CP1 exhibited no remarkable increase. These results indicate that LHCII serves a role in maintaining the stoichiometry of Chl to apoproteins by acting as a temporary pool for Chl molecules.Abbreviations ALA 5-aminolevulinic acid - Chl chlorophyll - CP chlorophyll-protein complex - CPa chlorophyll a-protein complex of PSII - CP1 P700-chlorophyll a-protein complex - LDS lithium dodecyl sulfate - LHCII light-harvesting chlorophyll a/b-protein complex of PSII This work was supported by the Grants-in-Aid for Scientific Research (04304004) from the Ministry of Education, Science and Culture, Japan.     

Light- and MgCl2-dependent characteristics of four chlorophyll-protein complexes isolated from the marine dinoflagellate,Glenodinium sp.

Chlorophyll-protein complexes previously isolated from low-light (80 μE·m⁻²·s⁻¹) log cultures of the marine dinoflagellate, Glenodinium sp., were further characterized. SDS solubilization in combination with polyacrylamide gel electrophoresis in the presence of Deriphat 160-C resolved four discrete chlorophyll-protein bands. In order to elucidate the functional role of Glenodinium sp., room-temperature absorption and fluorescence spectra, protein composition, and pigment molar ratios were obtained for each complex. Results indicated that complex I was analogous to the green plant Photosystem I complex and was also associated with light-harvesting chlorophyll c₂. Complex II was highly enriched in chlorophyll c₂, devoid of peridinin, and demonstrated energy transfer from chlorophyll c to chlorophyll a within the complex, indicating the presence of a light-harvesting component. Based on peridinin: chlorophyll a ratios and fluorescence excitation spectra analyses for complexes III and IV, it was concluded that these complexes contained functional peridinin-chlorophyll a-protein complexes. Changing the ionic environment during isolation of the complexes, or altering the growth irradiance of Glenodinium sp. cultures, resulted in a significant alteration of distribution of chlorophyll a among the chlorophyll-protein complexes.     

Purification and characterization of photosystem I and photosystem II core complexes from wild-type and phycocyanin-deficient strains of the cyanobacterium Synechocystis PCC 6803

Highly photoactive Photosystem I (PS I) and Photosystem II (PS II) core complexes have been isolated from the cyanobacterium Synechocystis Pasteur Culture Collection (PCC) 6803 and a phycocyanin-deficient mutant, enriched in PS II. Cell breakage using glass beads was followed by sucrose density gradient centrifugation and two high-performance liquid chromatography steps involving anion-exchange and hydroxyapatite. The PS I core complex has an apparent molecular mass of 300 +/- 20 kDa (including a detergent shell of about 50 kDa) and contains subunits of approximately 60, approximately 60, 18.5, 18.5, 16, 15, 10.5, 9.5, and 6.5 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblots; its antenna size is 75 +/- 5 chlorophyll/P-700. The PS II core complex has an apparent molecular mass of 310 +/- 20 kDa (including the detergent shell); subunits of 43, 37, 33, 29, and 10-11 kDa were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting. The antenna size of the average PS II complex is 45 +/- 5 chlorophyll/primary quinone electron acceptor (QA). This preparation procedure also yields, as a byproduct, a highly purified cytochrome b6f complex. This complex contains four subunits of 38, 24, 19, and 15 kDa and b- and c-type cytochromes in a ratio of 2:1. Its apparent molecular mass of 180 +/- 20 kDa (including the detergent shell) is consistent with a monomeric complex.     

The action of lipase on chloroplast membranes: II. Polypeptide patterns of bean galactolipase- and phospholipase A2-treated thylakoid membranes

Thylakoid membranes obtained from bean chloroplasts treated with bean galactolipase or phospholipase A₂ (from Crotalus terr. terr.) showed marked changes in their polypeptide patterns when separated on SDS-PAGE. The obtained results have been discussed with regard to the relationship between chloroplast lipids and polypeptides originating from chlorophyll-protein complexes of bean thylakoids. A coexistence between galactolipids and the peripheral antennae in PS I complex and LHCP³ as well as a conspicuous role of phospholipids in PSI and PSII centre chlorophyll-protein complexes has to be underlined.Abbreviations CP1 chlorophyll a-protein complex of PSI - CPa chlorophyll a-protein complex of PSII - D₁₀ digitonin subchloroplast particles enriched in PSII - D₁₄₄ digitonin subchloroplast particles enriched in PSI - DCMU 3-(3,4-dichlorophenyl)-1, 1-dimethylurea - LHCP^1-3 light harvesting chlorophyll a/b protein complexes - PAGE polyacrylamide gel electrophoresis - PSI photosystem I - PSII photosystem II - SDS sodium dodecyl sulphate - TCA trichloroacetic acid - Tricine N-Tris-(hydroxymethyl)-methylglycine - Tris Tris-(hydroxymethyl)-aminomethan     

Site-directed mutagenesis of the psbC gene of photosystem II: isolation and functional characterization of CP43-less photosystem II core complexes

Two mutants of Synechocystis PCC 6803 lacking the psbC gene product CP43 were constructed by site-directed mutagenesis. Analysis of cells and thylakoid membranes of these mutants indicates that PS II reaction centers accumulate to a concentration of about 10% of that of WT cells. PS II core complexes isolated from mutants lacking the CP43 subunit show light-driven electron transfer from the secondary electron donor Z to the primary quinone electron acceptor QA with a quantum yield similar to that of wild type, indicating that CP43 is not required for binding or function of QA. The use of mutants for the removal of CP43 thus avoids the loss of QA function associated with biochemical extraction of CP43 from intact core complexes. Both absorbance and fluorescence emission maxima of the mutant complexes show a blue shift in comparison to the WT PS II core complex, indicating that the absorbance spectrum of CP43 is red-shifted relative to that of the remainder of the core complex. The antenna size of these CP43-less complexes is about 70% of that of WT, indicating that approximately 15 chlorophyll molecules are bound by CP43. The molecular mass of the PS II complex, including the detergent shell, shifts from 310 +/- 15 kDa in WT to 285 +/- 15 kDa in the CP43-less mutants.     

Spectral anisotropy of chloroplasts, subchloroplast particles and pigment-protein complexes

Anisotropic properties of pea chloroplasts, subchloroplast fragments (photosystem 1 particles and pigment-protein complexes) and the blue-green algae oriented in polyacrylamide gel were investigated. It was shown that linear dichroism spectra of chloroplasts are the superposition of the corresponding spectra for the main light harvesting complex (HMLC) and P 700 chlorophyll a--protein complex (CP 1). Anisotropic properties of the photosystem 1 particles and blue-green algae are mainly caused by CP 1 anisotropy. Qy-transition moments tend to perpendicular orientation to the membrane plane for the Chl. b 649, Chl. a 660 and parallel orientation--for Chl. b 654, Chl. a 682. The degree of Qy-transition moments parallel orientation is higher for the longwave forms (Chl. a 690, Chl. a 702, Chl. a 712), than for the shortwave ones and coincides with this degree for the reaction centre pigment P 700 transition moment. It is suggested that the specific orientation of the pigment-protein complexes in the chloroplast membrane is important for the regulation of the spillover between two photosystems.     

Reinvestigation of the chlorophyll distribution among the chlorophyll-proteins and chlorophyll-protein complexes of Hordeum vulgare L.

Solubilization of barley (Hordeum vulgare L.) thylakoid membranes with sodium dodecylsulphate plus sodium deoxycholate with or without Triton X-100 and subsequent fractionation in the polyacrylamide gel electrophoresis system described in this paper resulted: (1) in the resolution of the chlorophyll-proteins and chlorophyll-protein complexes commonly known as CP1a, CP1, LHCP¹, LHCP², CPa and LHCP³; (2) in the highly increased stability of CP1 and CP1a, as judged by their chlorophyll content, (3) at the expense of the free pigment concentration (4) which could be reduced to a negligible amount. Some 40% of the total chlorophyll contained in the mature higher plant thylakoid membrane is associated with CP1 and CP1 a and as already suggested before [19] no significant amount of free chlorophyll occurs in vivo.Abbreviations chl chlorophyll - CP1 P₇₀₀-chla-protein - CPa P680-chla-protein - DOC sodium deoxychlolate - FC free chlorophyll - LHCP light-harvesting chlorophyll a/b-protein - PAGE(S) polyacrylamide gel electrophoresis (system) - SDS sodium dodecylsulphate - TX-100 Triton X-100     

The action of lipase on chloroplast membranes: II. Polypeptide patterns of bean galactolipase- and phospholipase A2-treated thylakoid membranes

Thylakoid membranes obtained from bean chloroplasts treated with bean galactolipase or phospholipase A₂ (from Crotalus terr. terr.) showed marked changes in their polypeptide patterns when separated on SDS-PAGE. The obtained results have been discussed with regard to the relationship between chloroplast lipids and polypeptides originating from chlorophyll-protein complexes of bean thylakoids. A coexistence between galactolipids and the peripheral antennae in PS I complex and LHCP³ as well as a conspicuous role of phospholipids in PSI and PSII centre chlorophyll-protein complexes has to be underlined.Abbreviations CP1 chlorophyll a-protein complex of PSI - CPa chlorophyll a-protein complex of PSII - D₁₀ digitonin subchloroplast particles enriched in PSII - D₁₄₄ digitonin subchloroplast particles enriched in PSI - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - LHCP^1–3 light harvesting chlorophyll a/b protein complexes - PAGE polyacrylamide gel electrophoresis - PSI photosystem I - PSII photosystem II - SDS sodium dodecyl sulphate - TCA trichloroacetic acid - Tricine N-Tris-(hydroxymethyl)-methylglycine - Tris Tris-(hydroxymethyl)-aminomethan