Main Light-harvesting Antenna Research Articles

S2P2-the chloroplast-located intramembrane protease and its impact on the stoichiometry and functioning of the photosynthetic apparatus of A. thaliana.

S2P2 is a nuclear-encoded protease, potentially located in chloroplasts, which belongs to the zinc-containing, intramembrane, site-2 protease (S2P) family. In A. thaliana cells, most of the S2P proteases are located within the chloroplasts, where they play an important role in the development of chloroplasts, maintaining proper stoichiometric relations between polypeptides building photosynthetic complexes and influencing the sensitivity of plants to photoinhibitory conditions. Among the known chloroplast S2P proteases, S2P2 protease is one of the least known. Its exact location within the chloroplast is not known, nor is anything known about its possible physiological functions. Therefore, we decided to investigate an intra-chloroplast localization and the possible physiological role of S2P2. To study the intra-chloroplast localization of S2P2, we used specific anti-S2P2 antibodies and highly purified chloroplast fractions containing envelope, stroma, and thylakoid proteins. To study the physiological role of the protease, we used two lines of insertion mutants lacking the S2P2 protease protein. Here, we present results demonstrating the thylakoid localization of S2P2. Moreover, we present experimental evidence indicating that the lack of S2P2 in A. thaliana chloroplasts leads to a significant decrease in the level of photosystem I and photosystem II core proteins: PsaB, PsbA, PsbD, and PsbC, as well as polypeptides building both the main light-harvesting antenna (LHC II), Lhcb1 and Lhcb2, as well as Lhcb4 and Lhcb5 polypeptides, constituting elements of the minor, peripheral antenna system. These changes are associated with a decrease in the number of PS II-LHC II supercomplexes. The consequence of these disorders is a greater sensitivity of s2p2 mutants to photoinhibition. The obtained results clearly indicate that the S2P2 protease is another thylakoid protein that plays an important role in the proper functioning of A. thaliana chloroplasts, especially in high-light-intensity conditions.

Substituted Methylenation at the 132 -Position of a Chlorophyll-a Derivative via Mixed Aldol Condensation, Optical Properties of the Synthetic Bacteriochlorophyll-d Analogs, and Self-aggregation of Their Zinc Complexes.

Chlorophyll-a derivatives possessing a substituted methylene group at the 132 -position were prepared by the mixed aldol condensation of methyl 3-hydroxymethyl-pyropheophorbide-a with aldehydes bearing a methyl, p-nitro/cyanophenyl, or pentafluorophenyl group. Their electronic absorption spectra were dependent on the substituents at the methylene terminal. The Soret bands were broadened by increasing the group electronegativity of the substituents, which was ascribable to the charge transfer from the core chlorin to the peripheral substituent in a molecule. Although their Qy absorption and fluorescence emission bands resembled each other, the emission intensities decreased with an increase in the electronegativity because of intramolecular electron transfer quenching. Some of their zinc complexes self-aggregated in a less polar organic solvent to give red-shifted and broadened absorption bands with intense circular dichroism couplets, which were similar to those of bacteriochlorophyll-c/d aggregates in natural chlorosomes as the main light-harvesting antennas of green photosynthetic bacteria and their models. The J-aggregation was suppressed with an enhancement in the size of the 132 -substituents.

In situ cryo-ET structure of phycobilisome-photosystem II supercomplex from red alga.

Phycobilisome (PBS) is the main light-harvesting antenna in cyanobacteria and red algae. How PBS transfers the light energy to photosystem II (PSII) remains to be elucidated. Here we report the in situ structure of the PBS-PSII supercomplex from Porphyridium purpureum UTEX 2757 using cryo-electron tomography and subtomogram averaging. Our work reveals the organized network of hemiellipsoidal PBS with PSII on the thylakoid membrane in the native cellular environment. In the PBS-PSII supercomplex, each PBS interacts with six PSII monomers, of which four directly bind to the PBS, and two bind indirectly. Additional three 'connector' proteins also contribute to the connections between PBS and PSIIs. Two PsbO subunits from adjacent PSII dimers bind with each other, which may promote stabilization of the PBS-PSII supercomplex. By analyzing the interaction interface between PBS and PSII, we reveal that αLCM and ApcD connect with CP43 of PSII monomer and that αLCM also interacts with CP47' of the neighboring PSII monomer, suggesting the multiple light energy delivery pathways. The in situ structures illustrate the coupling pattern of PBS and PSII and the arrangement of the PBS-PSII supercomplex on the thylakoid, providing the near-native 3D structural information of the various energy transfer from PBS to PSII.

Facile 132-methylation of chlorophyll-a derivative and (132R/S)-stereoselective self-aggregation of zinc bacteriochlorophyll-d analogs

Zinc methyl 3-hydroxymethyl-(132R/S)-methyl-pyropheophorbides-a were prepared as model compounds of bacteriochlorophyll-d by the chemical modification of naturally occurring chlorophyll-a. Both the HPLC-separated 132-epimers as well as the 132-unsubstituted counterpart self-aggregated in an aqueous Triton X-100 micellar solution to form large oligomers with red-shifted electronic absorption and intense circular dichroism bands, similarly as in natural chlorosomes, the main light-harvesting antennas of photosynthetic green bacteria. The substitution with a methyl group at the 132-position did not disturb the formation of chlorosomal self-aggregates in a J-type fashion, which were slightly dependent on the stereochemistry.

Synthesis of zinc 13-oxime-functionalized chlorophyll-a derivatives and their (131E/Z)-dependent self-aggregation

Zinc methyl 3-hydroxymethyl-pyropheophorbides-a possessing hydroxyimino, acetoxyimino, and benzoyloxyimino groups at the 131-position were prepared by chemical modification of naturally occurring chlorophyll-a. The (131E)- and (131Z)-stereoisomers of the obtained unsymmetric oximes were readily separated by column chromatography, and their stereochemistry was determined by 1H NMR spectroscopy. The synthetic O-(un)substituted oximes were monomeric in tetrahydrofuran and their electronic absorption spectra were dependent on the stereochemistry as well as the O-substituents in the oxime moiety. These chlorophyll-a derivatives self-aggregated in an aqueous Triton X-100 micellar solution to give red-shifted absorption bands. The J-aggregation ability of the (131E)-isomers was higher than that of the corresponding the (131Z)-isomers. Such stereoselective self-aggregation was comparable to that of bacteriochlorophylls-c, d, e, and f in the main light-harvesting antennas of green photosynthetic bacteria.

Effect of additional hydroxy group on self-aggregation of synthetic zinc bacteriochlorophyll-c analogs

Zinc methyl 3-hydroxymethyl-pyropheophorbides-a possessing a hydroxymethyl or 2-hydroxyethyl group at the 20-position were prepared by chemical modification of naturally occurring chlorophyll-a. The synthetic bacterio-chlorophyll-c analog bearing the 201-hydroxy group self-aggregated in an aqueous Triton X-100 micellar solution to give large oligomers with red-shifted and broadened electronic absorption bands, which was similar to the chlorophyllous J-aggregates in the main light-harvesting antennas of green photosynthetic bacteria. By contrast, the other 202-hydroxylated analog did not aggregate under the same conditions to remain its monomeric species. The red shift in the longest wavelength (Qy) band by the self-aggregation of the former analog was greater than that of the corresponding 201-methyl substitute with a similar size (CH3 ≈ OH). These results could be ascribable to the less sterically demanding and more electron-withdrawing 20-hydroxymethyl group, compared with more interacting 2-hydroxyethyl and more electron-donating ethyl groups at the 20-position.

Self-Aggregation abilities of synthetic bacteriochlorophyll-d analogs bearing a propargyl- or benzyl-type alcohol

The propargyl- or benzyl-type alcoholic moiety attached chlorophyll-a derivatives have been synthesized via Pd-mediated coupling reactions. These compounds could act as models of bacteriochlorophyll molecules in the main light-harvesting antenna (chlorosome) of photosynthetic green bacteria. Their self-aggregation abilities were investigated in less polar organic solvents.

BciC-Catalyzed C132 -Demethoxycarbonylation of Metal Pheophorbide a Alkyl Esters.

Bacteriochlorophyll c molecules self-aggregate to form large oligomers in the core part of chlorosomes, which are the main light-harvesting antenna systems of green photosynthetic bacteria. In the biosynthetic pathway of bacteriochlorophyll c, a BciC enzyme catalyzes the removal of the C132 -methoxycarbonyl group of chlorophyllide a, which possesses a free propionate residue at the C17-position and a magnesium ion as the central metal. The in vitro C132 -demethoxycarbonylations of chlorophyll a derivatives with various alkyl propionate residues and central metals were examined by using the BciC enzyme derived from one green sulfur bacteria species, Chlorobaculum tepidum. The BciC enzymatic reactions of zinc pheophorbide a alkyl esters were gradually suppressed with an increase of the alkyl chain length in the C17-propionate residue (from methyl to pentyl esters) and finally the hexyl ester became inactive for the BciC reaction. Although not only the zinc but also nickel and copper complexes were demethoxycarbonylated by the BciC enzyme, the reactions were largely dependent on the coordination ability of the central metals: Zn>Ni>Cu. The above substrate specificity indicates that the BciC enzyme would not bind directly to the carboxy group of chlorophyllide a, but would bind to its central magnesium to form the stereospecific complex of BciC with chlorophyllide a, giving pyrochlorophyllide a, which lacks the (132 R)-methoxycarbonyl group.

Photoactivated Supramolecular Assembly Using “Caged Chlorophylls” for the Generation of Nanotubular Self-Aggregates Having Controllable Lengths

Living supramolecular polymerization is a breakthrough system to control the size of supramolecular aggregates. Multidimensional (2D or 3D) self-aggregates of chiral and unsymmetric molecules are observed in nature. For instance, a chlorosome, which is the main light-harvesting antenna in green photosynthetic bacteria, possesses tubular structures of chlorophyll pigments with J-type slipped cofacial and circular arrangements. Here, we report size-controllable construction of chlorosome-like aggregates by photoactivated supramolecular assembly of “caged chlorophyll”. The caged chlorophylls, which were nonassembling (inert) species, were illuminated with UV-light to give the active species one by one, and then the resulting active monomers spontaneously assembled to construct tubular self-aggregates. The length of such chlorosome-like aggregate tubes was dependent on the UV-irradiation duration and intensity of UV-light. The photoactivated supramolecular assembly system would be useful for application of (o...

Phototriggered Dynamic and Biomimetic Growth of Chlorosomal Self-Aggregates.

Supramolecular polymerizations mimicking native systems, which are step-by-step constructions to form self-aggregates, were recently developed. However, a general system to successively and spontaneously form self-aggregates from monomeric species remains challenging. Here, we report a photoinduced supramolecular polymerization system as a biomimetic formation of chlorophyll aggregates which are the main light-harvesting antennas in photosynthetic green bacteria, called "chlorosomes". In this system, inert chlorophyll derivatives were UV-irradiated to gradually produce active species through deprotection. Such active monomers spontaneously assembled to form fiberlike chlorosomal self-aggregates in a similar manner as a dynamic growth of natural chlorosomal self-aggregates. The study would be useful for elucidation of the formation process of the chlorosomal aggregates and construction of other supramolecular structures in nature.

Synthesis of zinc 20-ethenylated bacteriochlorophyll-d analogs and their self-aggregation in an aqueous micelle solution

Zinc complexes of 3-hydroxymethyl-131-oxo-chlorins possessing a vinyl or its trans-substituted group at the 20-position were prepared through regioselective bromination and successive palladium-catalyzed cross-coupling reactions from naturally occurring chlorophyll-a. The synthetic zinc chlorophyll-a derivatives self-aggregated in an aqueous Triton X-100 micelle solution to give red-shifted and broadened electronic absorption bands, which were similar to those of bacteriochlorophyll-c/d self-aggregates in the main light-harvesting antenna systems of green photosynthetic bacteria. Because of the sterically demanded perpendicular orientation of the (un)substituted ethenyl group to the chlorin π-system, the present model compounds had less J-aggregation ability in a slipped π–π interaction fashion than the corresponding saturated ethyl-type analogs, while halogenations at the terminal of the 20-vinyl group enhanced the ability due to their electron-withdrawing effect.

Structure of phycobilisome from the red alga Griffithsia pacifica.

Life on Earth depends on photosynthesis for its conversion of solar energy to chemical energy. Photosynthetic organisms have developed a variety of light-harvesting systems to capture sunlight. The largest light-harvesting complex is the phycobilisome (PBS), the main light-harvesting antenna in cyanobacteria and red algae. It is composed of phycobiliproteins and linker proteins but the assembly mechanisms and energy transfer pathways of the PBS are not well understood. Here we report the structure of a 16.8-megadalton PBS from a red alga at 3.5 Å resolution obtained by single-particle cryo-electron microscopy. We modelled 862 protein subunits, including 4 linkers in the core, 16 rod-core linkers and 52 rod linkers, and located a total of 2,048 chromophores. This structure reveals the mechanisms underlying specific interactions between linkers and phycobiliproteins, and the formation of linker skeletons. These results provide a firm structural basis for our understanding of complex assembly and the mechanisms of energy transfer within the PBS.

20-Substitution effect on self-aggregation of synthetic zinc bacteriochlorophyll-d analogs

Abstract Zinc methyl 20-substituted 3 1 -demethyl-bacteriopheophorbides- d were prepared through palladium-catalyzed cross-coupling of chlorophyll- a derivatives. The synthetic zinc 3 1 -hydroxy-13 1 -oxo-chlorins possessing a(n) (un)substituted ethynyl or butadiynyl group at the 20-position were good models for bacteriochlorophylls- c / d found solely in chlorosomes, the main light-harvesting antennas of green photosynthetic bacteria. Similarly as in natural chlorosomes, the synthetic models self-aggregated in an aqueous Triton X-100 solution to form large oligomers with red-shifted and broadened electronic absorption bands. While a phenyl group directly connected with the chlorin π-system at the 20-position disturbed the chlorosomal self-aggregation, the insertion of an ethynylene group between the two functional groups reduced the steric hindrance around the 20-position and was useful for the facile formation of the self-aggregates. A similar substitution effect was observed in the 20-ethynyl to butadiynyl moieties bearing a sterically bulky trimethylsilyl group at the terminal position.

Self-aggregation of synthetic zinc methyl 20-substituted 3-hydroxymethyl-pyropheophorbides as models of bacteriochlorophyll-c.

Zinc 3-hydroxymethyl-131-oxo-chlorins bearing a variety of primary alkyl groups at the 20-position were prepared as models of bacteriochlorophyll-c by chemical modification of naturally occurring chlorophyll-a. The synthetic chlorophyll-a derivatives self-aggregated in an aqueous Triton X-100 solution to afford large oligomers whose Soret and Qy bands were red-shifted and broadened, compared with the bands of their monomers in tetrahydrofuran. The oligomeric bands are similar to those of bacteriochlorophyll-c self-aggregates in chlorosomes, the main light-harvesting antennae of photosynthetic green bacteria. The 20-alkylation led to bathochromic shifts of the visible Soret maxima in J-type self-aggregates of the synthetic models, while elongation of the 20-alkyl group decreased the chlorosomal Qy maxima due to an increase in steric hindrance. Considering the light-harvesting and energy-transferring processes in a chlorosome, the 20-methylation in bacteriochlorophyll-c would be more suitable for efficient culturing of green bacteria than the 20-ethylation and propylation as well as the 20-unsubstitution in bacteriochlorophyll-d.

Self-aggregation of synthetic chlorophyll-c derivative and effect of C17-acrylate residue on bridging green gap in chlorosomal model

Abstract A zinc bacteriochlorophyll- d analog possessing a fully π-conjugated porphyrin skeleton and an acrylate residue at the 17-position was prepared by chemical modification of an accessory photosynthetic pigment, chlorophyll- c 1 from a diatom Chaetoceros species. The synthetic zinc chlorophyll- c 1 derivative self-aggregated in a less polar organic solvent and in a solid film to give a red-shifted and broadened visible absorption spectrum, which is similar to that of the main light-harvesting antenna of a green photosynthetic bacterium, where magnesium chlorins with the 17-propionate residue including bacteriochlorophyll- d aggregate in a J -type fashion to form large oligomers. The resulting self-aggregates showed specific and intense absorption bands at around 500 and 600 nm due to the π-extension along the molecular x-axis, which could bridge the “green gap” observed in the above natural antenna system. Visible and infrared absorption spectral analyses revealed that the supramolecular structure of self-aggregates of the zinc chlorophyll- c 1 derivative are comparable to those of the natural antenna, in spite of double dehydrogenation at the 17,18- and 17 1 ,17 2 -positions in the molecule.

Visible and near-infrared spectra of chlorosomal zinc chlorin self-aggregates dependent on their peripheral substituents at the 8-position

Zinc complexes of 3-hydroxymethyl-131-oxo-chlorins possessing various π-conjugatable substituents at the 8-position were prepared by chemical modification of naturally occurring chlorophyll-a. The synthetic zinc chlorophyll-a derivatives self-aggregated in an aqueous Triton X-100 solution to give red-shifted and broadened electronic absorption bands compared with their monomeric species in tetrahydrofuran. Their self-assemblies in an aqueous micelle showed similar electronic absorption spectra to those of the main light-harvesting antennas in green photosynthetic bacteria. The self-aggregated species afforded largely bathochromically shifted Qy maxima to absorb near-infrared light.

In vitro stereospecific hydration activities of the 3-vinyl group of chlorophyll derivatives by BchF and BchV enzymes involved in bacteriochlorophyll c biosynthesis of green sulfur bacteria.

The photosynthetic green sulfur bacterium Chlorobaculum (Cba.) tepidum produces bacteriochlorophyll (BChl) c pigments bearing a chiral 1-hydroxyethyl group at the 3-position, which self-aggregate to construct main light-harvesting antenna complexes, chlorosomes. The secondary alcoholic hydroxy group is requisite for chlorosomal aggregation and biosynthesized by hydrating the 3-vinyl group of their precursors. Using recombinant proteins of Cba. tepidum BchF and BchV, we examined in vitro enzymatic hydration of some 3-vinyl-chlorophyll derivatives. Both the enzymes catalyzed stereoselective hydration of zinc 3-vinyl-8-ethyl-12-methyl-bacteriopheophorbide c or d to the zinc 31 R-bacteriopheophorbide c or d homolog, respectively, with a slight amount of the 31 S-epimric species. A similar R-stereoselectivity was observed in the BchF-hydration of zinc 3-vinyl-8-ethyl- and propyl-12-ethyl-bacteriopheophorbides c, while their BchV-hydration gave a relatively larger amount of the 31 S-epimers. The in vitro stereoselective hydration confirmed the in vivo production of the S-epimeric species by BchV. The enzymatic hydration for the above 8-propylated substrate proceeded more slowly than that for the 8-ethylated, and the 8-isobutylated substrate was no longer hydrated. Based on these results, biosynthetic pathways of BChl c homologs and epimers are proposed.

Design principles of natural light-harvesting as revealed by single molecule spectroscopy

Biology offers a boundless source of adaptation, innovation, and inspiration. A wide range of photosynthetic organisms exist that are capable of harvesting solar light in an exceptionally efficient way, using abundant and low-cost materials. These natural light-harvesting complexes consist of proteins that strongly bind a high density of chromophores to capture solar photons and rapidly transfer the excitation energy to the photochemical reaction centre. The amount of harvested light is also delicately tuned to the level of solar radiation to maintain a constant energy throughput at the reaction centre and avoid the accumulation of the products of charge separation. In this Review, recent developments in the understanding of light-harvesting by plants will be discussed, based on results obtained from single molecule spectroscopy studies. Three design principles of the main light-harvesting antenna of plants will be highlighted: (a) fine, photoactive control over the intrinsic protein disorder to efficiently use intrinsically available thermal energy dissipation mechanisms; (b) the design of the protein microenvironment of a low-energy chromophore dimer to control the amount of shade absorption; (c) the design of the exciton manifold to ensure efficient funneling of the harvested light to the terminal emitter cluster.

Synthesis of 20-iodochlorophyll derivatives and their properties including reactivity, electronic absorption, and self-aggregation

Methyl 20-iodopyropheophorbides-a possessing several substituents at the 3-position were prepared by treatment of the corresponding 20-unsubstituted chlorophyll-a derivatives with iodine and [bis(trifluoroacetoxy)iodo]benzene in chloroform and water at room temperature. The sterically demanding 20-iodination was clean, efficient, and regioselective without alteration of the 3-substituents, and also highly reproducible. The resulting 20-iodides were so reactive that they were readily transformed into 20-acetylchlorin (Pd-catalyzed cross-coupling) and 20-unsubstituted chlorin (acidic deiodination). Electronic absorption spectra in dichloromethane showed that the 20-iodination shifted the maxima to longer wavelengths in their monomeric states. Zinc complexes of 31-hydroxy-20-iodo-131-oxochlorins self-aggregated in non-polar organic solvents to give red-shifted and broadened absorption bands, similarly as in the corresponding 20-unsubstituted molecules, which are good models of the main light-harvesting antenna systems in photosynthetic green bacteria.

Synthesis of zinc 20-substituted bacteriochlorophyll-d analogs and their self-aggregation

Zinc complexes of methyl 31-demethyl-bacteriopheophorbides-d possessing a 2-(un)substituted ethynyl or ethenyl group at the 20-position were prepared through palladium-catalyzed cross-coupling of their 20-bromide with the corresponding alkyne or alkene. The synthetic zinc 20-substituted 3-hydroxymethyl-131-oxo-chlorins self-aggregated in an aqueous micellar solution to give similar large oligomers as self-aggregates of bacteriochlorophylls in the core part of the main light-harvesting antenna of photosynthetic green bacteria. The zinc 20-ethynyl-chlorin fully gave the self-aggregates, but its 202-substitution disturbed the self-aggregation. The 20-(2-phenyl)ethenyl moieties induced the formation of relatively smaller oligomers as well as residual monomers due to their steric bulkiness.