Characterization of Chloroplasts Isolated from Triazine-Susceptible and Triazine-Resistant Biotypes of Brassica campestris L

Chloroplasts isolated from triazine-susceptible and triazine-resistant biotypes of Brassica campestris L. were analyzed for lipid composition, ultrastructure, and relative quantum requirements of photosynthesis. In general, phospholipids, but not glycolipids in chloroplasts from the triazine-resistant biotype had a higher linolenic acid concentration and lower levels of oleic and linoleic fatty acids, than chloroplasts from triazine-susceptible plants. Chloroplasts from the triazine-resistant biotype had a 1.6-fold higher concentration of t-Δ3-hexadecenoic acid with a concomitantly lower palmitic acid concentration in phosphatidylglycerol. Phosphatidylglycerol previously has been hypothesized to be a boundary lipid for photosystem II. Chloroplasts from the triazine-resistant biotype had a lower chlorophyll a/b ratio and exhibited increased grana stacking. Light-saturation curves revealed that the relative quantum requirement for whole chain electron transport at limiting light intensities was lower for the susceptible biotype than for the triazine-resistant biotype. Although the level of the chlorophyll a/b light-harvesting complex associated with photosystem II was greater in resistant biotypes, the increased levels of the light-harvesting complex did not increase the photosynthetic efficiency enough to overcome the rate limitation that is inherited concomitantly with the modification of the Striazine binding site.     

Function and evolution of grana

Chloroplasts are descended from cyanobacteria, and they retain many features of the cyanobacterial photosynthetic apparatus. However, land-plant chloroplasts have a strikingly different thylakoid membrane organization to that of cyanobacteria. Usually the two photosystems are laterally segregated; Photosystem II is concentrated in complex stacked-membrane structures known as grana. The function of grana has long been debated. Recent studies on membrane organization in chloroplasts, cyanobacteria and purple bacteria now offer a new perspective. I argue that grana allow the presence of a large light-harvesting antenna for Photosystem II, without excessively restricting electron transport. Other organisms solve this problem in different ways. Land plants evolved from macroalgae that were adapted to high light conditions; they evolved grana as a new solution to the problem of efficient photosynthesis in shade.     

The structure and function of the chloroplast photosynthetic membrane — a model for the domain organization

Recent work on the domain organization of the thylakoid is reviewed and a model for the thylakoid of higher plants is presented. According to this model the thylakoid membrane is divided into three main domains: the stroma lamellae, the grana margins and the grana core (partitions). These have different biochemical compositions and have specialized functions. Linear electron transport occurs in the grana while cyclic electron transport is restricted to the stroma lamellae. This model is based on the following results and considerations. (1) There is no good candidate for a long-range mobile redox carrier between PS II in the grana and PS I in the stroma lamellae. The lateral diffusion of plastoquinone and plastocyanin is severely restricted by macromolecular crowding in the membrane and the lumen respectively. (2) There is an excess of 14±18% chlorophyll associated with PS I over that of PS II. This excess is assumed to be localized in the stroma lamellae where PS I drives cyclic electron transport. (3) For several plant species, the stroma lamellae account for 20±3% of the thylakoid membrane and the grana (including the appressed regions, margins and end membranes) for the remaining 80%. The amount of stroma lamellae (20%) corresponds to the excess (14–18%) of chlorophyll associated with PS I. (4) The model predicts a quantum requirement of about 10 quanta per oxygen molecule evolved, which is in good agreement with experimentally observed values. (5) There are at least two pools of each of the following components: PS I, PS II, cytochrome bf complex, plastocyanin, ATP synthase and plastoquinone. One pool is in the grana and the other in the stroma compartments. So far, it has been demonstrated that the PS I, PS II and cytochrome bf complexes each differ in their respective pools.Abbreviations PS I and PS II Photosystem I and II - P 700 reaction center of PS I - LHC II light-harvesting complex II     

Minor antenna proteins CP24 and CP26 affect the interactions between photosystem II subunits and the electron transport rate in grana membranes of Arabidopsis

We investigated the function of chlorophyll a/b binding antenna proteins Chlorophyll Protein 26 (CP26) and CP24 in light harvesting and regulation of photosynthesis by isolating Arabidopsis thaliana knockout lines that completely lacked one or both of these proteins. All three mutant lines had a decreased efficiency of energy transfer from trimeric light-harvesting complex II (LHCII) to the reaction center of photosystem II (PSII) due to the physical disconnection of LHCII from PSII and formation of PSII reaction center depleted domains in grana partitions. Photosynthesis was affected in plants lacking CP24 but not in plants lacking CP26: the former mutant had decreased electron transport rates, a lower DeltapH gradient across the grana membranes, reduced capacity for nonphotochemical quenching, and limited growth. Furthermore, the PSII particles of these plants were organized in unusual two-dimensional arrays in the grana membranes. Surprisingly, overall electron transport, nonphotochemical quenching, and growth of the double mutant were restored to wild type. Fluorescence induction kinetics and electron transport measurements at selected steps of the photosynthetic chain suggested that limitation in electron transport was due to restricted electron transport between Q(A) and Q(B), which retards plastoquinone diffusion. We conclude that CP24 absence alters PSII organization and consequently limits plastoquinone diffusion.     

Spatial relationship of photosystem I, photosystem II, and the light- harvesting complex in chloroplast membranes

We have previously demonstrated (Armond, P. A., C. J. Arntzen, J.-M. Briantais, and C. Vernotte. 1976. Arch. Biochem. Biophys. 175:54-63; and Davis, D. J., P. A. Armond, E. L. Gross, and C. J. Arntzen. 1976. Arch. Biochem. Biophys. 175:64-70) that pea seedlings which were exposed to intermittent illumination contained incompletely developed chloroplasts. These plastids were photosynthetically competent, but did not contain grana. We now demonstrate that the incompletely developed plastids have a smaller photosynthetic unit size; this is primarily due to the absence of a major light-harvesting pigment-protein complex which is present in the mature membranes. Upon exposure of intermittent- light seedlings to continuous white light for periods up to 48 h, a ligh-harvesting chlorophyll-protein complex was inserted into the chloroplast membrane with a concomitant appearance of grana stacks and an increase in photosynthetic unit size. Plastid membranes from plants grown under intermediate light were examined by freeze-fracture electron microscopy. The membrane particles on both the outer (PF) and inner (EF) leaflets of the thylakoid membrane were found to be randomly distributed. The particle density of the PF fracture face was approx. four times that of the EF fracture face. While only small changes in particle density were observed during the greening process under continuous light, major changes in particle size were noted, particularly in the EF particles of stacked regions (EFs) of the chloroplast membrane. Both the changes in particle size and an observed aggregation of the EF particles into the newly stacked regions of the membrane were correlated with the insertion of light-harvesting pigment- protein into the membrane. Evidence is presented for identification of the EF particles as the morphological equivalent of a "complete" photosystem II complex, consisting of a phosochemically active "core" complex surrounded by discrete aggregates of the light-harvesting pigment protein. A model demonstrating the spatial relationships of photosystem I, photosystem II, and the light-harvesting complex in the chloroplast membrane is presented.     

Localization of photophosphorylation and proton transport activities in various regions of the chloroplast lamellae

Membrane fragments released by French pressure cell treatment of whole chloroplasts and isolated by differential centrifugation have been characterized structurally and with respect to phosophorylating and proton transport activities. In agreement with results of other workers, the heavy fraction released by pressure treatment was found by electron microscopy studies to be made up of mostly intact grana stacks while the light fraction was comprised of vesicles derived from the stromal lamellae. Both fractions were found to carry out rapid rates of cyclic photophosphorylation catalyzed by phenazine methosulfate (PMS). However, only the grana membranes demonstrated active proton accumulation in the presence of PMS. No light induced H⁺ uptake could be detected in the stromal lamellae fraction; and as expected, proton gradient dissipating agents such as NH₄Cl, nigericin in the presence of K⁺, and gramicidin were only slightly inhibitory to phosphorylation at concentrations which were very inhibitory in the grana membrane fraction.

Further evidence that stromal lamellae do not have active proton transport in the intact chloroplast was obtained by comparing various chloroplasts having different amounts of stromal and grana membranes. Comparative studies on young and old chloroplasts from lettuce, mesophyll and bundle sheath cell plastids from sorghum, and greening plastids from etiolated corn seedlings revealed a direct correlation between the extent of grana formation and the amount of proton transport activity. Samples which had larger amounts of stromal lamellae had high rates of ATP formation but a reduced capacity for H⁺ accumulation.     

Freeze-fracture studies on barley plastid membranes. VII. Structural changes associated with phosphorylation of the light-harvesting complex

Chloroplast thylakoid membranes were isolated from barley at room temperature under redox conditions which ensured that the light-harvesting complex was either non-phosphorylated or phosphorylated. The ultrastructural appearance of these membranes was characterised by rotary shadowed, freeze-fracture electron microscopy. Upon phosphorylation, there was a slight (5%) decrease in the extent of thylakoid stacking, as evidenced by an increase in EFu face particle density. It was concluded from detailed measurements of particle density and size distribution that phosphorylation of the light-harvesting complex results in the movement of some of the Photosystem II EFs particles and some of the PFs particles containing the light-harvesting complex from grana to stroma membranes. There was also a slight increase in PFs particle size and the appearance of a population of large particles on this face, which may be due to conformational changes in the light-harvesting complex or to the movement of some Photosystem I particles from stroma to grana membranes.     

Photosynthetic Acclimation to Temperature in the Desert Shrub, Larrea divaricata: II. Light-harvesting Efficiency and Electron Transport

The response of photosynthetic electron transport and light-harvesting efficiency to high temperatures was studied in the desert shrub Larrea divaricata Cav. Plants were grown at day/night temperatures of 20/15, 32/25, or 45/33 C in rough approximation of natural seasonal temperature variations. The process of acclimation to high temperatures involves an enhancement of the stability of the interactions between the light-harvesting pigments and the photosystem reaction centers. As temperature is increased, the heat-induced dissociation of these complexes results in a decrease in the quantum yield of electron transport at limiting light intensity, followed by a loss of electron transport activity at rate-saturating light intensity. The decreased quantum yield can be attributed to a block of excitation energy transfer from chlorophyll b to chlorophyll a, and changes in the distribution of the excitation energy between photosystems II and I. The block of excitation energy transfer is characterized by a loss of the effectiveness of 480 nm light (absorbed primarily by chlorophyll b) to drive protochemical processes, as well as fluorescence emission by chlorophyll b.     

Measurements of cytochrome f and P-700 in intact leaves of Sinapis alba grown under high-light and low-light conditions

The oxidation and reduction of cytochrome f and P-700 is measured spectrophotometrically in leaves of low-light and high-light plants. After illumination with red light, an induction phenomenon for cytochrome f oxidation is observed which indicates a regulation of photosystem I activity through energy distribution between the pigment systems by the energy state of the membrane. After far-red excitation the reduction of cytochrome f in the dark is much slower in low-light leaves. This shows that cyclic electron transport is not improved in low-light plants under these conditions. P-700 is oxidized on excitation with far-red light. However, with high intensities of far-red light, P-700 is partially reduced again which is due to a low extent of photosystem II excitation with the far-red used in the experiments. The low-light leaves show greater sensitivity of photosystem II to this excitation. The initial rate of the cytochrome f oxidation-rate is the same in low-light and high-light leaves. This shows that several P-700 are connected with only one electron transport chain. The consequences of these results concerning the tripartite concept and the photosynthetic unit are discussed. In the high-light plants the experimental data can be well explained by the tripartite organization of the photosynthetic unit. In low-light plants, however, a multipartite organization has to be postulated. In the partition regions of the grana, several antennae systems I, antennae systems II, and light-harvesting complexes can communicate with one electron transport chain.Abbreviations CP I P-700-chlorophyll a-protein - Cyt f cytochrome f - DCMU 3-(3,4 dichlorophenyl)-1,1-dimethylurea - DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - LA leaf-area - PhAR photosynthetically active radiation - PS photosystem     

Lateral heterogeneity in the distribution of chlorophyll-protein complexes of the thylakoid membranes of spinach chloroplasts

The lateral distribution of the main chlorophyll-protein complexes between appressed and non-appressed thylakoid membranes has been studied. The reaction centre complexes of Photosystems I and II and the light-harvesting complex have been resolved by an SDS-polyacrylamide gel electrophoretic method which permits most of the chlorophyll to remain protein-bound.

The analyses were applied to subchloroplast fractions shown to be derived from different thylakoid regions. Stroma thylakoids were separated from grana stacks by centrifugation following chloroplast disruption by press treatment or digitonin. Vesicles derived from the grana partitions were isolated by aqueous polymer two-phase partition. A substantial depletion in the amount of Photosystem I chlorophyll-protein complex and an enrichment in the Photosystem II reaction centre complex and the light-harvesting complex occurred in the appressed grana partition region. The high enrichment in this fraction compared to grana stack fractions derived from press or digitonin treatments, suggests that the grana Photosystem I is restricted mainly to the non-appressed grana end membranes and margins, and that the grana partitions possess mainly Photosystem II reaction centre complex and the light-harvesting complex.

In contrast, stroma thylakoids are highly enriched in the Photosystem I reaction centre complex. They possess also some 10–20% of the total Photosystem II reaction centre complex and the light-harvesting complex.

The ratio of light-harvesting complex to Photosystem II reaction centre complex is rather constant in all subchloroplast fractions suggesting a close association between these complexes. This was not so for the ratio of light-harvesting complex and the Photosystem I reaction centre complex.

The lateral heterogeneity in the distribution of the photosystems between appressed and non-appressed membranes must have a profound impact on current understanding of both the distribution of excitation energy and photosynthetic electron transport between the photosystems.     

Low-light-induced formation of semicrystalline photosystem II arrays in higher plant chloroplasts

Remodeling of photosynthetic machinery induced by growing spinach plants under low light intensities reveals an up-regulation of light-harvesting complexes and down-regulation of photosystem II and cytochrome b6f complexes in intact thylakoids and isolated grana membranes. The antenna size of PSII increased by 40-60% as estimated by fluorescence induction and LHCII/PSII stoichiometry. These low-light-induced changes in the protein composition were accompanied by the formation of ordered particle arrays in the exoplasmic fracture face in grana thylakoids detected by freeze-fracture electron microscopy. Most likely these highly ordered arrays consist of PSII complexes. A statistical analysis of the particles in these structures shows that the distance of neighboring complexes in the same row is 18.0 nm, the separation between two rows is 23.7 nm, and the angle between the particle axis and the row is 26 degrees . On the basis of structural information on the photosystem II supercomplex, a model on the supramolecular arrangement was generated predicting that two neighboring complexes share a trimeric light-harvesting complex. It was suggested that the supramolecular reorganization in ordered arrays in low-light grana thylakoids is a strategy to overcome potential diffusion problems in this crowded membrane. Furthermore, the occurrence of a hexagonal phase of the lipid monogalactosyldiacylglycerol in grana membranes of low-light-adapted plants could trigger the rearrangement by changing the lateral membrane pressure.     

Adaptability of tomato and pepper leaves to changes in photoperiod: Effects on the composition and function of the thylakoid membrane

The adaptability of the thylakoid membrane to extended photoperiod (from natural to 24 h) was studied using a photoperiod-sensitive species ( Lycopersicon esculentum Mill. cv. Trend) and a non-photoperiod-sensitive species ( Capsicum annuum L. cv. Delphin). Our results have shown that thylakoid membranes of both species adapt to an extended photoperiod by increasing their photosystem II to photosystem I ratio (PSII/PSI) in order to provide a more balanced energy distribution between both photosystems to improve quantum yield. In tomato plants, these results correspond with a lower chlorophyll (Chl) a/b ratio, a decrease in Chl associated with PSI light-harvesting chlorophyll a/b protein complexes and with an increase in Chl associated with PSII light-harvesting chlorophyll a/b protein complexes. In spite of these changes, the electron transport capacity through PSII and PSI per unit of Chl and the light saturation point of PSII remained unchanged. The inability of tomato plants to use supplemental light for an extended photoperiod is not the result of photoinhibitory conditions. In pepper plants a significant increase in electron transport capacity and in the light saturation point of PSII was found. There was a significant increase in CO₂ assimilation when the light period was increased from 12 to 24 h. In contrast to tomato, pepper plants adapt to a 24-h photoperiod by increasing their carboxylation capacity which is accompanied by an increase in electron transport capacity and the light saturation point.     

Chloroplast structure in normal and pigment-deficient soybeans grown in continuous red or far-red light

Clark L1, a normal green soybean [ Glycine max (L.) Merrill] and Clark y₉y₉, a backross-developed isoline exhibiting pigment deficiency, were grown under continuous red (11 W m⁻² and far-red (9 W m⁻²) light. Chloroplast thylakoids from the unifoliolate leaf (9–10 days old) were isolated and analyzed for pigments, pigment-protein, membrane polypeptides, electron transport and ultrastructural differences. Chloroplasts of soybean plants grown under far-red light have decreased chlorophyll a to chlorophyll b ratio, increased light-harvesting complexes, and grana structure with few stroma-type thylakoids. Photosystem II/photosystem I ratios (PSII/PSI) are higher in far-red due to decreased synthesis of PSI reaction center and/or less antenna associated with PSI.     

类囊体膜的垛叠、松散与它的功能关系

菠菜完整叶绿体置于4mM MgCl_2或20 mM KCl低浓度介质中低渗10秒钟后,得到由Mg~(++)或K~+离子诱导的类囊体垛叠膜和松散膜。它们在功能上表现出明显的差异。垛叠膜有较高的毫秒级延迟光发射(ms-DLE),松散膜显著降低DLE的快相,垛叠膜比松散膜的9-AA荧光猝灭快,并保持稳定;而松散膜有H~+渗漏。在非循环或Fd催化的循环光合磷酸化中,垛叠膜比松散膜活力高。但是,若在同样的低渗介质中低渗1分钟以上,Mg~(++)离子诱导的垛叠膜,在显微结构上不同于低渗过10秒钟的垛叠膜,它垛叠较松,而且在磷酸化活力上也与松散膜差别不大。揭示了H~+传递速度受二个光系统、电子载体间的距离及偶联程度的限制。新鲜制备的垛叠或松散膜,在NADP~+还原系统中,具有相同的电子传递放O_2速度,说明电子传递速度在一定范围内不受膜间的距离和偶联程度的影响。但是松散膜不稳定,随着膜的老化而解联,牛血清白蛋白(BSA)能稳定松散膜的电子传递。     

Regulation of photosystem stoichiometry,chlorophyll a and chlorophyll b content and relation to chloroplast ultrastructure

The structural-functional organization of higher plant chloroplasts has been investigated in relation to the particular light conditions during plant growth. (1) Light intensity variations during growth caused changes in the $\text{Chl}\text{a}\text{Chl}\text{b}$ ratio, in the light-saturated uncoupled rates of electron transport to a Hill oxidant and in the distribution of the chloroplast volume between the membrane and stroma phases. (2) Light quality differences during growth had an effect on the PS II/PS I reaction center ratio and on the chloroplast membrane phase differentiation into grana and stroma thylakoids. Plants grown under far-red-enriched (680–710 nm) illumination contained higher (20–25%) amounts of PS II and simultaneously lower (20–25%) amounts of PS I reaction centers. They also showed a higher grana density along with thicker grana stacks in their chloroplasts. (3) The size of the light-harvesting antenna pool of PS II centers was estimated from the fluorescence time course of 3-(3′,4′-dichlorophenyl)-1,1-dimethylurea-poisoned chloroplasts and was found to be fairly constant (±10%) in spite of the variable PS II/PS I reaction center ratio. The results are compatible with the hypothesis that the structural entities of grana facilitated the centralization and relative concentration increase of a certain group of PS II reaction centers.     

The accumulation of the light-harvesting 2 complex during remodeling of the Rhodobacter sphaeroides intracytoplasmic membrane results in a slowing of the electron transfer turnover rate of photochemical reaction centers

A functional proteomic analysis of the intracytoplasmic membrane (ICM) development process was performed in Rhodobacter sphaeroides during adaptation from high-intensity illumination to indirect diffuse light. This initiated an accelerated synthesis of the peripheral light-harvesting 2 (LH2) complex relative to that of LH1-reaction center (RC) core particles. After 11 days, ICM vesicles (chromatophores) and membrane invagination sites were isolated by rate-zone sedimentation and subjected to clear native gel electrophoresis. Proteomic analysis of gel bands containing the RC-LH1 and -LH2 complexes from digitonin-solubilized chromatophores revealed high levels of comigrating electron transfer enzymes, transport proteins, and membrane assembly factors relative to their equivalent gel bands from cells undergoing adaptation to direct low-level illumination. The GroEL chaperonin accounted for >65% of the spectral counts in the RC-LH1 band from membrane invagination sites, which together with the appearance of a universal stress protein suggested that the viability of these cells was challenged by light limitation. Functional aspects of the photosynthetic unit assembly process were monitored by near-IR fast repetition rate analysis of variable fluorescence arising from LH-bacteriochlorophyll a components. The quantum yield of the primary charge separation during the early stages of adaptation showed a gradual increase (variable/maximal fluorescence = 0.78-0.83 between 0 and 4 h), while the initial value of ~70 for the functional absorption cross section (σ) gradually increased to 130 over 4 days. These dramatic σ increases showed a direct relation to gradual slowing of the RC electron transport turnover rate (τ(QA)) from ~1.6 to 6.4 ms and an ~3-fold slowing of the rate of reoxidation of the ubiquinone pool. These slowed rates are not due to changes in UQ pool size, suggesting that the relation between increasing σ and τ(QA) reflects the imposition of constraints upon free diffusion of ubiquinone redox species between the RC and cytochrome bc(1) complex as the membrane bilayer becomes densely packed with LH2 rings.     

Arrangement of photosystem II supercomplexes in crystalline macrodomains within the thylakoid membrane of green plant chloroplasts

The chloroplast thylakoid membrane of green plants is organized in stacked grana membranes and unstacked stroma membranes. We investigated the structural organization of Photosystem II (PSII) in paired grana membrane fragments by transmission electron microscopy. The membrane fragments were obtained by a short treatment of thylakoid membranes with the mild detergent n-dodecyl-alpha, d-maltoside and are thought to reflect the grana membranes in a native state. The membranes frequently show crystalline macrodomains in which PSII is organized in rows spaced by either 26.3 nm (large-spaced crystals) or 23 nm (small-spaced crystals). The small-spaced crystals are less common but better ordered. Image analysis of the crystals by an aperiodic approach revealed the precise positions of the core parts of PSII in the lattices, as well as features of the peripheral light-harvesting antenna. Together, they indicate that the so-called C(2)S(2) and C(2)S(2)M supercomplexes form the basic motifs of the small-spaced and large-spaced crystals, respectively. An analysis of a pair of membranes with a well-ordered large-spaced crystal reveals that many PSII complexes in one layer face only light-harvesting complexes (LHCII) in the other layer. The implications of this type of organization for the efficient transfer of excitation energy from LHCII to PSII and for the stacking of grana membranes are discussed.     

Energization and ultrastructural pattern of thylakoids formed under periodic illumination followed by continuous light

Bean leaves grown under periodic illumination (56 cycles of 2 min light and 98 min darkness) were subsequently exposed to continuous illumination, and in connection with granum formation and accumulation of the light-harvesting pigment-protein complex thermoluminescence and light-induced shrinkage of thylakoid membranes were studied. Juvenile chloroplasts with large double sheets of thylakoids obtained under periodic light exhibited low temperature spectra of polarized fluorescence yielding fluorescence polarization (FP) values < 1 at 695 nm, characteristic for pheophytin emission. In the course of maturation under continuous light when normal grana appeared and the chlorophyll a/b light-harvesting photosystem II complex was incorporated into the membrane, at 695 nm the relative intensity of fluorescence dropped and FP changed to a value of > 1, suggesting an overlap between the emission of pheophytin and that of the chlorophyll a/b light-harvesting photosystem II complex. Thermoluminescence glow curves recorded with juvenile thylakoids displayed a relatively high proportion of emission at low temperatures (around -10°C) while with mature chloroplasts, more thermoluminescence originated from energetically deeper traps (discharged around 28°C). This means that during thylakoid development the capacity of the membrane to stabilize the separated charges increases, which might be favourable for the ultimate conservation of energy. The more extensive energization of mature thylakoids was also indicated by a light-induced decrease in the thickness of the membranes upon illumination; a change which could not be detected in juvenile thylakoids.Abbreviations EDTA ethylenediamine tetraacetic acid - Hepes 4-(2-hydroxy ethyl)-1-piperazine ethane sulfonic acid Dedicated to Prof. L.N.M. Duysens on the occasion of his retirement.     

Comparative studies of two membrane fractions isolated from chemotrophically and phototrophically grown cells of Rhodopseudomonas capsulata.

Light and heavy membrane fractions have been isolated by equilibrium sucrose density centrifugation from Rhodopseudomonas capsulata 938 GCM grown aerobically in the dark (chemotrophically) and anaerobically in the light (phototrophically). The densities of the light and heavy fractions from phototrophic cells were 1.1004 to 1.1006 and 1.1478, respectively, and the densities of the light and heavy fractions from chemotrophic cells were 1.0957 to 1.0958 and 1.1315, respectively. Both fractions were active in photochemical and respiratory functions and in electron transport-coupled phosphorylation. The light membrane fraction isolated from chemotrophic cells contained the reaction center and the light-harvesting pigment-protein complex B 870, but not the variable light-harvesting complex B 800-850. A small amount of the complex B 800-850 was present in the light fraction isolated from phototrophically grown cells, but it was not energetically coupled to the photosynthetic apparatus. From inhibitor studies, difference spectroscopy, and measurement of enzyme activities it was tentatively concluded that the light membrane fraction contains only the reduced nicotinamide adenine dinucleotide-oxidizing electron transport chain having a KCN-insensitive, low-potential cytochrome c oxidase, whereas the heavy fraction contains additionally the succinate dehydrogenase and a high-potential cytochrome b terminal oxidase sensitive to KCN. The light membrane fraction was more labile than the heavy fraction in terms of phosphorylating activity.     

Immuno-gold localization of cytochrome f,light-harvesting complex,ATP synthase and ribulose 1,5-bisphosphate carboxylase/oxygenase

The proteins ribulose 1,5-bisphosphate carboxylase/oxygenase, ATP synthase, light-harvesting chlorophyll a/b protein, and cytochrome f, have been localized in mesophyll chloroplasts of barley (Hordeum vulgare L.) by electron microscopy of immunogold-labelled sections. The light-harvesting chlorophyll a/b protein and cytochrome f are shown to be present in the grana, both within the stacks and at the margins, and in the stromal membranes. Although the absolute amount of labelling for these proteins is greater in the grana than in the stromal membranes, when expressed as label/membrane length the partitioning appears approximately equal between appressed and non-appressed membranes for both the light-harvesting chlorophyll a/b protein and cytochrome f. ATP synthase is restricted to the non-appressed thylakoid membranes, and ribulose 1,5-bisphosphate carboxylase/oxygenase is uniformly distributed through the stromal contents.Abbreviations CF₁ ATP synthase - LHCPII light-harvesting chlorophyll a/b protein - Rubisco ribulose 1,5-bisphosphate carboxylase/oxygenase