Major Light-harvesting Pigment-protein Complex Research Articles

Effect of Protonation on the Secondary Structure and Orientation of Plant Light-Harvesting Complex II Studied by PM-IRRAS.

The major light-harvesting pigment-protein complex of photosystem II, LHCII, has a crucial role in the distribution of the light energy between the two photosystems, the efficient light capturing and protection of the reaction centers and antennae from overexcitation. In this work direct structural information on the effect of LHCII protonation, which mimics the switch from light-harvesting to photoprotective state of the protein, was revealed by polarization-modulated infrared reflection-absorption spectroscopy (PM-IRRAS). PM-IRRAS on LHCII monolayers verified that the native helical structure of the protein is preserved in both partly deprotonated (pH 7.8, LHCII) and protonated (pH 5.2, p-LHCII) states. At low surface pressure, 10 mN/m, the orientation of the α-helices in these two LHCII states is different-tilted (θ ≈ 40°) in LHCII and nearly vertical (θ ≈ 90°) in p-LHCII monolayers; the partly deprotonated complex is more hydrophilic than the protonated one and exhibits stronger intertrimer interactions. At higher surface pressure, 30 mN/m, which is typical for biological membranes, the protonation affects neither the secondary structure nor the orientation of the transmembrane α-helices (tilted ∼45° relative to the membrane surface in both LHCII states) but weakens the intermolecular interactions within and/or between the trimers.

Utilization of light by fucoxanthin-chlorophyll-binding protein in a marine centric diatom, Chaetoceros gracilis.

The major light-harvesting pigment protein complex (fucoxanthin-chlorophyll-binding protein complex; FCP) was purified from a marine centric diatom, Chaetoceros gracilis, by mild solubilization followed by sucrose density gradient centrifugation, and then characterized. The dynamic light scattering measurement showed unimodality, indicating that the complex was highly purified. The amount of chlorophyll a (Chl a) bound to the purified FCP accounted for more than 60% of total cellular Chl a. The complex was composed of three abundant polypeptides, although there are nearly 30 FCP-related genes. The two major components were identified as Fcp3 (Lhcf3)- and Fcp4 (Lhcf4)-equivalent proteins based on their internal amino acid sequences and a two-dimensional isoelectric focusing electrophoresis analysis developed in this work. Compared with the thylakoids, the FCP complex showed higher contents of fucoxanthin and chlorophyll c but lower contents of the xanthophyll cycle pigments diadinoxanthin and diatoxanthin. Fluorescence excitation spectra analyses indicated that light harvesting, rather than photosystem protection, is the major function of the purified FCP complex, which is associated with more than 60% of total cellular Chl a. These findings suggest that the huge amount of Chl bound to the FCP complex composed of Lhcf3, Lhcf4, and an unidentified minor protein has a light-harvesting function to allow efficient photosynthesis under the dim-light conditions in the ocean.

Ion transport across model lipid membranes containing light-harvesting complex II: an effect of light

The effect of light on proton transport across lipid membranes of small unilamellar liposomes containing incorporated major light-harvesting pigment–protein complex of Photosystem II (LHCII) has been studied with the application of pH-sensitive dyes entrapped inside vesicles. Proton permeability coefficient for LHCII-modified membranes was found to be about twice as high as in the case of the control pure lipid vesicles. Illumination of the samples with light absorbed by the LHCII-bound photosynthetic pigments considerably affects the kinetics of proton transport: it increases the rate and decreases the steady-state level of proton gradient across the membranes. The effect was interpreted in terms of heat-induced conformational changes of LHCII molecular structures that affect proton buffering capacity of this protein. Both the control and the LHCII-modified lipid membranes have been found to be practically impermeable to Ca ++ ions, as demonstrated by fluorescence of liposome-entrapped calcium-sensitive probe calcium crimson. The slight differences in the proton transport across the LHCII-containing membranes under the presence of Ca ++ suggest calcium binding to this antenna protein.

Effect of xanthophyll pigments on fluorescence of chlorophyll a in LHC II embedded to liposomes

A major light-harvesting pigment-protein complex of Photosystem II (LHC II) isolated from rye leaves was incorporated to egg yolk lecithin liposomes containing various amounts (0-0.6 mol % with respect to lipid) of xanthophyll pigments: violaxanthin, lutein or zeaxanthin. It appeared that the addition of xanthophylls increased the level of the chlorophyll a fluorescence emission despite the screening effect of additional pigments in the system. Another phenomenon documented by the 77 K fluorescence emission spectra was the xanthophyll-related desaggregation of LHC II within the lipid phase of liposomes. The analysis of the parameters of chlorophyll a fluorescence decay in comparison to steady state fluorescence emission let us conclude that the fluorescence reabsorption in aggregated LHC II was a major cause of the lower level of fluorescence emission in that state of antenna proteins. Chlorophyll singlet excitations quenching brought about by violaxanthin, lutein and zeaxanthin is also documented in liposomes containing incorporated LHC II and exogenous xanthophyll pigments, and in liposomes containing xanthophyll pigments and chlorophyll b.

Involvement of xanthophyll pigments in regulation of light-driven excitation quenching in light-harvesting complex of Photosystem II

Light-driven chlorophyll a fluorescence quenching and dark reversibility of this process was measured in LHC II — the major light-harvesting pigment-protein complex of Photosystem II. Two preparations of LHC II, complex isolated from dark-adapted (LHC II-D) and light-pretreated (LHC II-L) rye leaves, differed significantly in the level of the xanthophyll pigment violaxanthin, being enriched in LHC II-D. LHC II-L contained only trace amounts of zeaxanthin but additionally antheraxanthin. LHC II-D suspended in buffer showed similar fluorescence quenching kinetics. Chlorophyll fluorescence quenching in LHC II-L was almost fully reversible after 2 h of dark-adaptation whereas only 37% of the initial fluorescence level was recovered under identical conditions. Transfer of LHC II as protein-lipid complex (PLC) in a reverse micellar solution composed of isooctane, phospholipids and limited amount of water, considerably decreased xanthophyll pigment concentrations. Under these conditions, light-driven fluorescence chlorophyll a quenching was the same for LHC II-L and LHC II-D and fluorescence level in both preparations was fully reversible within the dark period. Kinetics of light-induced fluorescence quenching and reversibility of LHC II-L reconstituted in asolectin liposomes was faster than in buffer. However, reversibility of LHC II-L in liposomes was markedly slowed down by addition of exogenous xanthophyll pigments in the following sequence: violaxanthin > zeaxanthin > nonpigmented liposomes. Remarkable light-dependent changes in chlorophyll a fluorescence excitation spectra of LHC II-L in liposomes were observed. These changes are ascribed to energetic uncoupling caused by physical detachment of violaxanthin from the antenna complex — this process being discussed in terms of violaxanthin availability to de-epoxidation in thylakoid membrane. The possible involvement of the xanthophyll cycle pigments violaxanthin and zeaxanthin in the regulation of light-driven excitation quenching in LHC II is also discussed.

A direct measurement of thermal energy dissipation in the photosynthetic apparatus during induction of fluorescence

A calorimetric method was applied to monitor the heat emission during the induction of photosynthesis (fluorescence) in the dark-adapted green alga Chlorella kessleri suddenly illuminated with HeNe laser light. The same method was also applied to record the heat emission from an illuminated preparation of the major light-harvesting pigment-protein complex of photosystem II (LHCII). The results are discussed in terms of the kinetics of recorded fluorescence changes. The direct measurement of thermal energy dissipation by the calorimetric technique is compared with indirect methods based on photothermal techniques such as photoacoustic spectroscopy.

LHCII, the major light-harvesting pigment-protein complex is a zeaxanthin epoxidase

Several experiments are presented indicating that LHCII, the major light-harvesting pigment-protein complex of Photosystem II isolated from rye leaves, exhibits properties of an enzyme catalyzing the epoxidation of zeaxanthin to violaxanthin. On the basis of this as well as other recent findings, a new model of a spatial organization of the xanthophyll cycle within a thylakoic membrane is proposed.

The light-harvesting complex of Mamiella gilva: a character linking scaly and naked members of the Micromonadophyceae (Chlorophyta)

The major light-harvesting pigment-protein complex of Mamiella gilva (Parke et Rayns) Moestrup (Micromonadophyceae, Chlorophyta) was isolated by lithium dodecyl sulphate polyacrylamide gel electrophoresis at 4°C. This complex is similar to that of Mantoniella squamata (Manton et Parke) Desikachary and Micromonas pusilla (Butcher) Manton et Parke with regard to molecular weight, absorption spectrum, and immunochemical cross-reactivity. On the other hand, the light-harvesting complex of Chlamydomonas reinhardtii P. Dangeard is quite different. The distribution of this unusual light-harvesting complex, which has to date been reported only in Mamiella, Mantoniella, and Micromonas, suggests a line of evolution in the Micromonadophyceae separate from all other green algae. In addition, the inclusion of the naked Micromonas in this group suggests that Micromonas should be included with Mamiella and Mantoniella in the Mamiellales, an order which is presently defined by scale type.

Biogenesis and Light Regulation of the Major Light Harvesting Chlorophyll-Protein of Diatoms

The apoprotein of the major light harvesting pigment-protein complex from the diatom Phaeodactylum tricornutum (UTEX 646) is composed of two similar polypeptides of 17.5 and 18.0 kilodaltons (kD). The in vivo synthesis of these polypeptides is inhibited by the 80s protein synthesis inhibitor cycloheximide, but not by the 70s ribosome inhibitor chloramphenicol. When total poly(A)(+) RNA was used in in vitro protein synthesis, a number of polypeptides were synthesized with a dominant product at 22 kD. When the polypeptides were immunoprecipitated with monospecific antibodies to the 17.5 and 18.0 polypeptides, a single protein zone of 22 kD was detected. Immunoprecipitation with preimmune serum failed to precipitate detectable levels of protein at any relative molecular weight (M(r)). These findings indicate that the two apoprotein polypeptides of the diatom light harvesting pigment-protein are translated from polyadenylated message on cytoplasmic ribosomes as either a single or two (or more) similar M(r) precursor proteins. These findings also suggest that this protein is encoded in the nucleus.Photosynthetic light adaptation features of P. tricornutum UTEX 646 indicate that it responds to low light by increasing cell size and numbers of photosystem I and II reaction centers per cell, but does not change photosynthetic rate per cell or photosynthetic unit sizes significantly. When low light cells are exposed to higher photon flux densities, the in vivo incorporation of label into the apoprotein of the light harvesting complex decreases. In contrast, high light grown cells show rapid (<3 hour) increases in apoprotein synthesis when exposed to low light levels. This is the first demonstration of a specific role of photon flux density in regulating the synthesis of a major light harvesting pigment-protein during photosynthetic light adaptation.

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.