Higher Plant Thylakoids Research Articles

Growth-phase dependent morphological alteration in higher plant thylakoid is accompanied by changes in both photodamage and repair rates.

Thylakoid membranes of young leaves consist of grana and stroma lamellae (stroma-grana [SG] structure). The SG thylakoid is gradually converted into isolated grana (IG), almost lacking the stroma lamellae during growth. This morphological alteration was found to cause a reduction in maximum photosynthetic rate and an enhancement of photoinhibition in photosystem II (PSII). In situ microspectrometric measurements of chlorophyll fluorescence in individual chloroplasts suggested an increase of the PSII/PSI ratio in IG thylakoids of mature leaves. Western blot analysis of isolated IG thylakoids showed relative increases in some PSII components, including the core protein (D1) and light-harvesting components CP24 and Lhcb2. Notably, a nonphotochemical quenching-related factor in the PSII supercomplex, PsbS, decreased by 40%. Changes in the high light response of PSII were detected through parameters of pulse-amplitude modulation fluorometry. Chlorophyll fluorescence lifetime indicated an increase of fluorescence quantum yield in IG. A minimal photodamage-repair rate analysis on a lincomycin treatment of the leaves indicated that repair rate constant of IG is slower than that of SG, while photodamage rate of IG is higher than that of SG. These results suggest that IG thylakoids are relatively sensitive to high light, which is not only due to a higher photodamage rate caused by some rearrangements of PS complexes, but also to the retarded PSII repair that may result from the lack of stroma lamellae. The IG thylakoids found among many plant species thus seem to be an adaptive form to low light environments, although their physiological roles still remain unclear.

Pigment-protein complexes are organized into stable microdomains in cyanobacterial thylakoids

Thylakoids are the place of the light-photosynthetic reactions. To gain maximal efficiency, these reactions are conditional to proper pigment-pigment and protein-protein interactions. In higher plants thylakoids, the interactions lead to a lateral asymmetry in localization of protein complexes (i.e. granal/stromal thylakoids) that have been defined as a domain-like structures characteristic by different biochemical composition and function (Albertsson P-Å. 2001,Trends Plant Science 6: 349–354). We explored this complex organization of thylakoid pigment-proteins at single cell level in the cyanobacterium Synechocystis sp. PCC 6803. Our 3D confocal images captured heterogeneous distribution of all main photosynthetic pigment-protein complexes (PPCs), Photosystem I (fluorescently tagged by YFP), Photosystem II and Phycobilisomes. The acquired images depicted cyanobacterial thylakoid membrane as a stable, mosaic-like structure formed by microdomains (MDs). These microcompartments are of sub-micrometer in sizes (~0.5–1.5 μm), typical by particular PPCs ratios and importantly without full segregation of observed complexes. The most prevailing MD is represented by MD with high Photosystem I content which allows also partial separation of Photosystems like in higher plants thylakoids. We assume that MDs stability (in minutes) provides optimal conditions for efficient excitation/electron transfer. The cyanobacterial MDs thus define thylakoid membrane organization as a system controlled by co-localization of three main PPCs leading to formation of thylakoid membrane mosaic. This organization might represent evolutional and functional precursor for the granal/stromal spatial heterogeneity in photosystems that is typical for higher plant thylakoids.

Pea PSII-LHCII supercomplexes form pairs by making connections across the stromal gap

In higher plant thylakoids, the heterogeneous distribution of photosynthetic protein complexes is a determinant for the formation of grana, stacks of membrane discs that are densely populated with Photosystem II (PSII) and its light harvesting complex (LHCII). PSII associates with LHCII to form the PSII-LHCII supercomplex, a crucial component for solar energy conversion. Here, we report a biochemical, structural and functional characterization of pairs of PSII-LHCII supercomplexes, which were isolated under physiologically-relevant cation concentrations. Using single-particle cryo-electron microscopy, we determined the three-dimensional structure of paired C2S2M PSII-LHCII supercomplexes at 14 Å resolution. The two supercomplexes interact on their stromal sides through a specific overlap between apposing LHCII trimers and via physical connections that span the stromal gap, one of which is likely formed by interactions between the N-terminal loops of two Lhcb4 monomeric LHCII subunits. Fast chlorophyll fluorescence induction analysis showed that paired PSII-LHCII supercomplexes are energetically coupled. Molecular dynamics simulations revealed that additional flexible physical connections may form between the apposing LHCII trimers of paired PSII-LHCII supercomplexes in appressed thylakoid membranes. Our findings provide new insights into how interactions between pairs of PSII-LHCII supercomplexes can link adjacent thylakoids to mediate the stacking of grana membranes.

Redox regulation of the antimycin A sensitive pathway of cyclic electron flow around photosystem I in higher plant thylakoids

The chloroplast must regulate supply of reducing equivalents and ATP to meet rapid changes in downstream metabolic demands. Cyclic electron flow around photosystem I (CEF) is proposed to balance the ATP/NADPH budget by using reducing equivalents to drive plastoquinone reduction, leading to the generation of proton motive force and subsequent ATP synthesis. While high rates of CEF have been observed in vivo, isolated thylakoids show only very slow rates, suggesting that the activity of a key complex is lost or down-regulated upon isolation. We show that isolation of thylakoids while in the continuous presence of reduced thiol reductant dithiothreitol (DTT), but not oxidized DTT, maintains high CEF activity through an antimycin A sensitive ferredoxin:quinone reductase (FQR). Maintaining low concentrations (~2mM) of reduced DTT while modulating the concentration of oxidized DTT leads to reversible activation/inactivation of CEF with an apparent midpoint potential of −306mV (±10mV) and n=2, consistent with redox modulation of a thiol/disulfide couple and thioredoxin-mediated regulation of the plastoquinone reductase involved in the antimycin A-sensitive pathway, possibly at the level of the PGRL1 protein. Based on proposed differences in regulatory modes, we propose that the FQR and NADPH:plastoquinone oxidoreductase (NDH) pathways for CEF are activated under different conditions and fulfill different roles in chloroplast energy balance.

The significance of CP29 reversible phosphorylation in thylakoids of higher plants under environmental stresses

Reversible phosphorylation of proteins is a key event in many fundamental cellular processes. Under stressful conditions, many thylakoid membrane proteins in photosynthetic apparatus of higher plants undergo rapid phosphorylation and dephosphorylation in response to environmental changes. CP29 is the most frequently phosphorylated protein among three minor antennae complexes in higher plants. CP29 phosphorylation in dicotyledons has been known for several decades and is well characterized. However, CP29 phosphorylation in monocotyledons is less studied and appears to have a different phosphorylation pattern. In this review, we discuss recent advancements in CP29 phosphorylation and dephosphorylation studies and its physiological significance under environmental stresses in higher plants, especially in the monocotyledonous crops. Physiologically, the phosphorylation of CP29 is likely to be a prerequisite for state transitions and the disassembly of photosystem II supercomplexes, but not involved in non-photochemical quenching (NPQ). CP29 is phosphorylated in monocots exposed to environmental cues, with its subsequent lateral migration from grana stacks to stroma lamellae. However, neither CP29 phosphorylation nor its lateral migration occurs in dicotyledonous plants after drought, cold, or salt stress. Since the molecular mechanisms of differential CP29 phosphorylation under stresses are not fully understood, this review provides insights for future studies regarding the physiological function of CP29 reversible phosphorylation.

Three-Dimensional Architecture of Grana and Stroma Thylakoids of Higher Plants as Determined by Electron Tomography

We have investigated the three-dimensional (3D) architecture of the thylakoid membranes of Arabidopsis (Arabidopsis thaliana), tobacco (Nicotiana tabacum), and spinach (Spinacia oleracea) with a resolution of approximately 7 nm by electron tomography of high-pressure-frozen/freeze-substituted intact chloroplasts. Higher-plant thylakoids are differentiated into two interconnected and functionally distinct domains, the photosystem II/light-harvesting complex II-enriched stacked grana thylakoids and the photosystem I/ATP synthase-enriched, nonstacked stroma thylakoids. The grana thylakoids are organized in the form of cylindrical stacks and are connected to the stroma thylakoids via tubular junctions. Our data confirm that the stroma thylakoids are wound around the grana stacks in the form of multiple, right-handed helices at an angle of 20° to 25° as postulated by a helical thylakoid model. The junctional connections between the grana and stroma thylakoids all have a slit-like architecture, but their size varies tremendously from approximately 15 × 30 nm to approximately 15 × 435 nm, which is approximately 5 times larger than seen in chemically fixed thylakoids. The variable slit length results in less periodicity in grana/stroma thylakoid organization than proposed in the original helical model. The stroma thylakoids also exhibit considerable architectural variability, which is dependent, in part, on the number and the orientation of adjacent grana stacks to which they are connected. Whereas some stroma thylakoids form solid, sheet-like bridges between adjacent grana, others exhibit a branching geometry with small, more tubular sheet domains also connecting adjacent, parallel stroma thylakoids. We postulate that the tremendous variability in size of the junctional slits may reflect a novel, active role of junctional slits in the regulation of photosynthetic function. In particular, by controlling the size of junctional slits, plants could regulate the flow of ions and membrane molecules between grana and stroma thylakoid membrane domains.

Redox state of the photosynthetic electron transport chain in wild-type and mutant leaves of Arabidopsis thaliana: Impact on photosystem II fluorescence

In addition to the photosynthetic linear electron transport, several alternative electron transport routes exist in thylakoids of higher plants. The plastoquinone (PQ) pool acts as a common electron carrier in these pathways. In the cyclic electron flow around photosystem I (PSI), reduced ferredoxin is used by the ferredoxin-quinone reductase (FQR) to reduce the PQ pool. Chlororespiratory pathway consists in the reduction of the PQ pool by the NAD(P)H dehydrogenase (NDH). These alternative pathways and their role in photosynthesis are still not fully understood. In the present study, the accumulation kinetics of quinone acceptors was measured by fluorescence induction in leaves of Arabidopsis thaliana wild-type and mutants altered in alternative electron pathways after various light- and dark-adaptation conditions. Results show that NDH activity can be probed by fluorescence induction during light-to-dark transition of plants. Also, the activity of FQR pathway did not affect directly the FI kinetics. However, the accumulation kinetics of reduced PQ under actinic light was dependant on the redox state of PSI acceptors prior to illumination.

Determination of phosphoproteins in higher plant thylakoids.

Redox-dependent thylakoid protein phosphorylation regulates both short- and long-term acclimation of the photosynthetic apparatus to changes in environmental conditions. The major thylakoid phosphoproteins belong to photosystem (PS)II (D1, D2, CP43, PsbH) and its light-harvesting antenna (Lhcb1, Lhcb2, CP29) but a number of minor phosphoproteins have also been identified. The detection methods traditionally include the radiolabeling techniques, electrophoretic separation of the phosphorylated and unphosphorylated forms of the protein, and the use of phosphoamino acid antibodies. The recent progress in mass spectrometry (MS) techniques and methods of proteomics allow for successful identification and analyses of protein phosphorylation. In mass spectrometry approaches, exogenous tracer is not needed and natural phosphorylation of proteins can be characterized with high sensitivity yielding the mapping of exact phosphorylation sites in the proteins as well. Various methods for detection of thylakoid phosphoproteins, including the preparation of phosphopeptides for mass spectrometric analyses and techniques for phosphopeptide identification by electrospray ionization MS are described. The experimental protocols for simultaneous identification of multiple phosphopeptides in complex peptide mixtures and for assess of stoichiometry for in vivo phosphorylation of multiple proteins are outlined.

A Note on Three-Dimensional Models of Higher-Plant Thylakoid Networks

Ever since the electron microscope became commercially available in 1939, plant biologists have exploited this powerful tool to elucidate the intricate structure of higher-plant thylakoid networks to correlate structure with function and chromatic adaptability as well as to gain insight into the

Thylakoid Membrane Remodeling during State Transitions inArabidopsis

Adaptability of oxygenic photosynthetic organisms to fluctuations in light spectral composition and intensity is conferred by state transitions, short-term regulatory processes that enable the photosynthetic apparatus to rapidly adjust to variations in light quality. In green algae and higher plants, these processes are accompanied by reversible structural rearrangements in the thylakoid membranes. We studied these structural changes in the thylakoid membranes of Arabidopsis thaliana chloroplasts using atomic force microscopy, scanning and transmission electron microscopy, and confocal imaging. Based on our results and on the recently determined three-dimensional structure of higher-plant thylakoids trapped in one of the two major light-adapted states, we propose a model for the transitions in membrane architecture. The model suggests that reorganization of the membranes involves fission and fusion events that occur at the interface between the appressed (granal) and nonappressed (stroma lamellar) domains of the thylakoid membranes. Vertical and lateral displacements of the grana layers presumably follow these localized events, eventually leading to macroscopic rearrangements of the entire membrane network.

Time-resolution of the Antheraxanthin- and ΔpH-dependent Chlorophyll a Fluorescence Components Associated with Photosystem II Energy Dissipation in Mantoniella squamata¶

The electronic excited-state behavior of photosystem II (PSII) in Mantoniella squamata, as influenced by the xanthophyll cycle and the transthylakoid pH gradient (ΔpH), was examined in vivo.Mantoniella is distinguished from other photosynthetic organisms by two main features namely (1) a unique light-harvesting complex that serves both photosystems I (PSI) and II (PSII); and (2) a violaxanthin (V) cycle that undergoes only one de-epoxidation step in excess light to accumulate the monoepoxide antheraxanthin (A) as opposed to the epoxide-free zeaxanthin (Z). The cells were treated first with high light to induce the ΔpH and A accumulation, followed by herbicide-induced closure of PSII traps and a chilling treatment, to sustain and stabilize the ΔpH and nigericin-sensitive fluorescence level in the dark. De-epoxidation was controlled with subsaturating concentrations of dithiothreitol (DTT) and was 5–10 times more sensitive to DTT than higher plant thylakoids. The PSII energy dissipation involved two steps: (1) the pH activation of the xanthophyll binding site that was associated with a narrowing and slight attenuation of the main 2 ns (ns = 10−9 s) fluorescence lifetime distribution; and (2) the concentration-dependent binding of A to the activated binding site yielding a second distribution centered around 0.9 ns. Consistent with the model of Gilmore et al. (1998) (Biochemistry 37, 13 582–13 593), the fractional intensity of the 0.9 ns component depended almost entirely on the A concentration and correlated linearly with the decrease of the steady-state chlorophyll a fluorescence intensity.

Intramembrane formation of hydrogen peroxide during oxygen reduction in thylakoids of higher plants.

monitored by measuring light scattering and chlorophyll fluorescence quantum yield in the presence of diuron in suspensions of the control and unstacked thylakoids [9]. The content of chlorophyll in thylakoids was determined by 95% ethanol extract [10]. The reaction medium ( 21°e ) was illuminated through a KC-10 red light filter ( λ > 600 nm) with light with an intensity of 500 µ mol photons s ‐1 m ‐2 . The content of e 2 in vessel was measured with a Clark-type oxygen electrode. Cytochrome c reduction was determined by changes in the optical density ∆ A 550–540 using the differential extinction coefficient 19 mM ‐1 cm ‐1 [6]. It was found that the rate of cytochrome c reduction reaches a pla

Mechanistic aspects of the xanthophyll dynamics in higher plant thylakoids

Plant thylakoids have a highly conserved xanthophyll composition, consisting of β‐carotene, lutein, neoxanthin and a pool of violaxanthin that can be converted to antheraxanthin and zeaxanthin in excess light conditions. Recent work has shown that xanthophylls undergo dynamic changes, not only in their composition but also in their distribution among Lhc proteins. Xanthophylls are released from specific binding site in the major trimeric LHCII complex of photosystem II and are subsequently bound to different sites into monomeric Lhcb proteins and dimeric Lhca proteins. In this work we review available evidence from in vivo and in vitro studies on the structural determinants that control xanthophyll exchange in Lhc proteins. We conclude that the xanthophyll exchange rate is determined by the structure of individual Lhc gene products and it is specifically controlled by the lumenal pH independently from the activation state of the violaxanthin de‐epoxidase enzyme. The xanthophyll exchange induces important modifications in the organization of the antenna system of Photosystem II and, possibly of Photosystem I. Major changes consist into a modulation of the light harvesting efficiency and an increase of the protection from lipid peroxidation. The xanthophyll cycle thus appears to be a signal transduction system for co‐ordinated regulation of the photoprotection mechanisms under persistent stress from excess light.

Segregation of the photosystems in higher plant thylakoids and short- and long-term regulation by a mesoscopic approach

In this paper we consider the relationship between the lateral segregation of photosystems I and II in the grana and characteristics of the short- and long-term regulation in thylakoids following the mesoscopic approach. Our study is thermodynamic; it is based on the Flory–Huggins theory for binary mixtures and the McMillan–Mayer theory of heterogeneous solutions. We demonstrate that state transitions promote rearrangement of photosystems by either favoring their mixing after LHCII phosphorylation or enhancing their segregation after LHCII dephosphorylation. This regulation influences the entire system properties locally. We also demonstrate that the variations of the photosystem ratio promote rearrangement of the photosystems preserving their segregation. This regulation influences the entire system properties globally. The studies presented are another indication of the importance of the segregation of the photosystems in the grana thylakoids of higher plants. Segregation of PSIs and PSIIs is a signature of the spinodal decomposition, which is a fine regulatory mechanism, related to both the short- and long-term adaptations of the photosynthetic apparatus in higher plant thylakoids.

Diatom Fucoxanthin Chlorophyll a/c-binding Protein (FCP) and Land Plant Light-harvesting Proteins Use a Similar Pathway for Thylakoid Membrane Insertion

The light-harvesting proteins in plastids of different lineages including algae and land plants represent a superfamily of chlorophyll-binding proteins that seem to be phylogenetically related, although some of the light-harvesting complex (LHC) proteins bind different carotenoids. LHCs can be divided into chlorophyll a/b-binding proteins found in green algae, euglenoids, and higher plants and into chlorophyll a/c-binding proteins of various algal taxa. LHC proteins from diatoms are named fucoxanthin-chlorophyll a/c-binding proteins (FCP). In contrast to chlorophyll a/b-binding proteins, there is no information so far about the way FCPs integrate into thylakoid membranes. The diatom FCP preproteins have a bipartite presequence that is necessary to enable transport into the four membrane-bound diatom plastids, but similar to chlorophyll a/b-binding proteins there is apparently no presequence present for targeting to the thylakoid membrane. By establishing an in vitro import assay for diatom thylakoids, we demonstrated that thylakoid integration of diatom FCP depends on the presence of stromal factors and GTP. This indicates that a pathway involving signal recognition particles (SRP) is involved in membrane integration just as shown for LHCs in higher plants. We also demonstrate integration of diatom FCP into thylakoids of higher plants and vice versa SRP-dependent targeting of LHCs from pea and Arabidopsis into diatom thylakoids. The similar SRP-dependent modes of thylakoid integration of land plant LHCs and FCPs support recent analyses indicating a common origin of chlorophyll a/b- and a/c-binding proteins.

An assessment of the pathways of dark reduction and oxidation of the plastoquinone pool in thylakoid membranes of higher plants and green algae

Several possible `chlororespiratory? pathways have been proposed for both reduction and oxidation of the plastoquinone pool in the dark in cells of both higher plants and green algae. However, bioinformatic analysis of the recently published genome of Arabidopsis thaliana sets limits to the types of respiratory-related components that could be present in higher plant thylakoids: for example, homologues of cytochrome c oxidase, succinate dehydrogenase and alternative NDH-2 enzymes in thylakoids can be ruled out and there are also no thylakoid-targetted homologues of the NADH-reactive fragment of mitochondrial complex I (the 75kDa, 51 kDa and 30 kDa subunits) that might provide an NADH dehydrogenase fragment for the other complex I homologues that are encoded in the chloroplast genome. In parallel to these considerations, direct assay of pathways of dark reduction and oxidation of plastoquinone has been made in intact leaves and in cells of Chlamydomonas reinhardtii by monitoring P700+ reduction, the slow phase of the carotenoid bandshift and turnover kinetics of the cytochrome bf complex. Combination of these measurements with effects of a wide range of specific inhibitors of various quinone-reactive sites has allowed further definition of the types of pathways and their extents.

Time-resolution of the antheraxanthin- and delta pH-dependent chlorophyll a fluorescence components associated with photosystem II energy dissipation in Mantoniella squamata.

The electronic excited-state behavior of photosystem II (PSII) in Mantoniella squamata, as influenced by the xanthophyll cycle and the transthylakoid pH gradient (delta pH), was examined in vivo. Mantoniella is distinguished from other photosynthetic organisms by two main features namely (1) a unique light-harvesting complex that serves both photosystems I (PSI) and II (PSII); and (2) a violaxanthin (V) cycle that undergoes only one de-epoxidation step in excess light to accumulate the monoepoxide antheraxanthin (A) as opposed to the epoxide-free zeaxanthin (Z). The cells were treated first with high light to induce the delta pH and A accumulation, followed by herbicide-induced closure of PSII traps and a chilling treatment, to sustain and stabilize the delta pH and nigericin-sensitive fluorescence level in the dark. De-epoxidation was controlled with subsaturating concentrations of dithiothreitol (DTT) and was 5-10 times more sensitive to DTT than higher plant thylakoids. The PSII energy dissipation involved two steps: (1) the pH activation of the xanthophyll binding site that was associated with a narrowing and slight attenuation of the main 2 ns (ns = 10(-9) s) fluorescence lifetime distribution; and (2) the concentration-dependent binding of A to the activated binding site yielding a second distribution centered around 0.9 ns. Consistent with the model of Gilmore et al. (1998) (Biochemistry 37, 13,582-13,593), the fractional intensity of the 0.9 ns component depended almost entirely on the A concentration and correlated linearly with the decrease of the steady-state chlorophyll alpha fluorescence intensity.

Control of the photosynthetic electron transport by PQ diffusion microdomains in thylakoids of higher plants

We investigate the role of plastoquinone (PQ) diffusion in the control of the photosynthetic electron transport. A control analysis reveals an unexpected flux control of the whole chain electron transport by photosystem (PS) II. The contribution of PSII to the flux control of whole chain electron transport was high in stacked thylakoids (control coefficient, CJ(PSII)=0.85), but decreased after destacking (CJ(PSII)=0.25). From an ‘electron storage’ experiment, we conclude that in stacked thylakoids only about 50 to 60% of photoreducable PQ is involved in the light-saturated linear electron transport. No redox equilibration throughout the membrane between fixed redox groups at PSII and cytochrome (cyt) bf complexes, and the diffusable carrier PQ is achieved. The data support the PQ diffusion microdomain concept by Lavergne et al. [J. Lavergne, J.-P. Bouchaud, P. Joliot, Biochim. Biophys. Acta 1101 (1992) 13–22], but we come to different conclusions about size, structure and size distribution of domains. From an analysis of cyt b 6 reduction, as a function of PSII inhibition, we conclude that in stacked thylakoids about 70% of PSII is located in small domains, where only 1 to 2 PSII share a local pool of a few PQ molecules. Thirty percent of PSII is located in larger domains. No small domains were found in destacked thylakoids. We present a structural model assuming a hierarchy of specific, strong and weak interactions between PSII core, light harvesting complexes (LHC) II and cyt bf. Peripheral LHCII’s may serve to connect PSII–LHCII supercomplexes to a flexible protein network, by which small closed lipid diffusion compartments are formed. Within each domain, PQ moves rapidly and shuttles electrons between PSII and cyt bf complexes in the close vicinity. At the same time, long range diffusion is slow. We conclude, that in high light, cyt bf complexes located in distant stromal lamellae (20 to 30%) are not involved in the linear electron transport.

Organization of the pigment molecules in the thylakoids and the chlorophyll a/c light-harvesting complex of a xanthophyte alga, Pleurochloris meiringensis. A linear dichroism study

We have studied the orientation of the absorption transition dipoles of the pigment molecules in thylakoid membranes, in the isolated core complex of photosystem I (CCI) and in the chlorophyll a/c light-harvesting complex of photosystem II (Chl a/c LHC) of the xanthophyte alga Pleurochloris meiringensis. In thylakoids, linear dichroism (LD) spectra reveal two distinct orientations of the Q y transition dipoles of the Chl a molecules: the long-wavelength Chl a molecules tend to lie in the plane of the thylakoid membrane, while those at shorter wavelengths tilt out of the membrane plane. Our data show that in CCl, the orientation of Q y dipoles of Chl a, and the overall orientation pattern of all transition dipoles in the visible region, are very similar to those in higher-plant photosystem I particles. The Chl a molecules with their Q y dipoles tilting out of the membrane plane are associated with the LHC, and constitute a substantial part of the Chl a population. The majority of the carotenoid molecules of LHC also tend to tilt out of the membrane plane. These data further corroborate our earlier conclusion that the molecular organization of thylakoid membranes and most particularly of the Chl a/c LHC differs significantly from the organization of higher-plant thylakoids and the main Chl a/b complexes, respectively.

Immunological studies on chlorophyll-a/b proteins and their distribution in thylakoid membrane domains

The immunological relationships between chlorophyll-a/b proteins from higher-plant thylakoid membranes have been studied by assaying purified chlorophyll proteins (CPs) with polyclonal and monoclonal antibodies. Although low levels of cross-reactions were observed between all light-harvesting proteins, the peripheral antennae (LHCII) were largely distinct from the inner antennae (CP 26 and CP 29). Chlorophyll-protein 24 and LHCI-680 have been proposed to have a role in connecting the inner and outer antennae, respectively, in photosystems I and II, and were closely related. The immunological relationships closely corresponded to the spectral properties. Antibodies were also used for locating chlorophyll-a/b proteins in grana, stroma and bundle-sheath membranes showing a strong lateral heterogeneity, which was maintained following State I State II transition. The only exception to this pattern was a specific LHCII population enriched in State-II stroma membranes. Chlorophyll proteins from bundlesheath chloroplasts, that have only cyclic electron flow, had epitopes distinct from those of their mesophyll homologues.