Chlorosomes are light-harvesting antennae of photosynthetic bacteria containing large numbers of self-aggregated bacteriochlorophyll (BChl) molecules. They have developed unique photophysical properties that enable them to absorb light and transfer the excitation energy with very high efficiency. However, the molecular-level organization, that produces the photophysical properties of BChl molecules in the aggregates, is still not fully understood. One of the reasons is heterogeneity in the chlorosome structure which gives rise to a hierarchy of structural and energy disorder. In this report, we for the first time directly measure absorption linear dichroism (LD) on individual, isolated chlorosomes. Together with fluorescence-detected three-dimensional LD, these experiments reveal a large amount of disorder on the single-chlorosome level in the form of distributions of LD observables in chlorosomes from wild-type bacterium Chlorobaculum tepidum . Fluorescence spectral parameters, such as peak wavelength and bandwidth, are measures of the aggregate excitonic properties. These parameters obtained on individual chlorosomes are uncorrelated with the observed LD distributions and indicate that the observed disorder is due to inner structural disorder along the chlorosome long axis. The excitonic disorder that is also present is not manifested in the LD distributions. Limiting values of the LD parameter distributions, which are relatively free of the effect of structural disorder, define a range of angles at which the excitonic dipole moment is oriented with respect to the surface of the two-dimensional aggregate of BChl molecules. Experiments on chlorosomes of a triple mutant of Chlorobaculum tepidum show that the mutant chlorosomes have significantly less inner structural disorder and higher symmetry, compatible with a model of well-ordered concentric cylinders. Different values of the transition dipole moment orientations are consistent with a different molecular level organization of BChl's in the mutant and wild-type chlorosomes.

The self-aggregated state of bacteriochlorophyll (BChl) c molecules in chlorosomes belonging to a bchQ bchR mutant of the green sulfur bacteria Chlorobaculum tepidum, which mostly produces a single 17(2)-farnesyl-(R)-[8-ethyl,12-methyl]BChl c homologue, was characterized by solid-state nuclear magnetic resonance (NMR) spectroscopy and high-resolution electron microscopy. A nearly complete (1)H and (13)C chemical shift assignment was obtained from well-resolved homonuclear (13)C-(13)C and heteronuclear (1)H-(13)C NMR data sets collected from (13)C-enriched chlorosome preparations. Pronounced doubling (1:1) of specific (13)C and (1)H resonances revealed the presence of two distinct and nonequivalent BChl c components, attributed to all syn- and all anti-coordinated parallel stacks, depending on the rotation of the macrocycle with respect to the 3(1)-methyl group. Steric hindrance from the 20-methyl functionality induces structural differences between the syn and anti forms. A weak but significant and reproducible reflection at 1/0.69 nm(-1) in the direction perpendicular to the curvature of cylindrical segments observed with electron microscopy also suggests parallel stacking of BChl c molecules, though the observed lamellar spacing of 2.4 nm suggests weaker packing than for wild-type chlorosomes. We propose that relaxation of the pseudosymmetry observed for the wild type and a related BChl d mutant leads to extended domains of alternating syn and anti stacks in the bchQ bchR chlorosomes. Domains can be joined to form cylinders by helical syn-anti transition trajectories. The phase separation in domains on the cylindrical surface represents a basic mechanism for establishing suprastructural heterogeneity in an otherwise uniform supramolecular scaffolding framework that is well-ordered at the molecular level.

Chlorophyllous pigments are essential for photosynthesis. Bacteriochlorophyll (BChl) b has the characteristic C8-ethylidene group and therefore is the sole naturally occurring pigment having an absorption maximum at near-infrared light wavelength. Here we report that chlorophyllide a oxidoreductase (COR), a nitrogenase-like enzyme, showed distinct substrate recognition and catalytic reaction between BChl a- and b-producing proteobacteria. COR from BChl b-producing Blastochloris viridis synthesized the C8-ethylidene group from 8-vinyl-chlorophyllide a. In contrast, despite the highly conserved primary structures, COR from BChl a-producing Rhodobacter capsulatus catalyzes the C8-vinyl reduction as well as the previously known reaction of the C7 = C8 double bond reduction on 8-vinyl-chlorophyllide a. The present data indicate that the plasticity of the nitrogenase-like enzyme caused the branched pathways of BChls a and b biosynthesis, ultimately leading to ecologically different niches of BChl a- and b-based photosynthesis differentiated by more than 150 nm wavelength.

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The discovery of new chlorophyllous pigments would provide greater understanding of the mechanisms and evolution of photosynthesis. Bacteriochlorophyll f has never been observed in nature, although this name was proposed ~40 years ago based on structurally related compounds. We constructed a bacteriochlorophyll f–accumulating mutant of the green sulfur bacterium Chlorobaculum limnaeum, which originally produced bacteriochlorophyll e, by knocking out the bchU gene encoding C-20 methyltransferase based on natural transformation. This novel pigment self-aggregates in an in vivo light-harvesting antenna, the chlorosome, and exhibits a Qy peak of 705 nm, more blue-shifted than any other chlorosome reported so far; the peak overlaps the maximum (~700 nm) of the solar photon flux spectrum. Bacteriochlorophyll f chlorosomes can transfer light energy from core aggregated pigments to another bacteriochlorophyll in the chlorosomal envelope across an energy gap of ~100 nm, and is thus a promising material for development of new bioenergy applications.

"Candidatus Chloracidobacterium thermophilum" is a recently discovered chlorophototroph from the bacterial phylum Acidobacteria, which synthesizes bacteriochlorophyll (BChl) c and chlorosomes like members of the green sulfur bacteria (GSB) and the green filamentous anoxygenic phototrophs (FAPs). The pigments (BChl c homologs and carotenoids), quinones, lipids, and hopanoids of cells and chlorosomes of this new chlorophototroph were characterized in this study. "Ca. Chloracidobacterium thermophilum" methylates its antenna BChls at the C-8 2 and C-12 1 positions like GSB, but these BChls were esterified with a variety of isoprenoid and straight-chain alkyl alcohols as in FAPs. Unlike the chlorosomes of other green bacteria, "Ca. Chloracidobacterium thermophilum" chlorosomes contained two major xanthophyll carotenoids, echinenone and canthaxanthin. These carotenoids may confer enhanced protection against reactive oxygen species and could represent a specific adaptation to the highly oxic natural environment in which "Ca. Chloracidobacterium thermophilum" occurs. Dihydrogenated menaquinone-8 [menaquinone-8(H 2 )], which probably acts as a quencher of energy transfer under oxic conditions, was an abundant component of both cells and chlorosomes of "Ca. Chloracidobacterium thermophilum." The betaine lipid diacylglycerylhydroxymethyl-N,N,N-trimethyl-␤-alanine, esterified with 13-methyl-tetradecanoic (isopentadecanoic) acid, was a prominent polar lipid in the membranes of both "Ca. Chloracidobacterium thermophilum" cells and chlorosomes. This lipid may represent a specific adaptive response to chronic phosphorus limitation in the mats. Finally, three hopanoids, diploptene, bacteriohopanetetrol, and bacteriohopanetetrol cyclitol ether, which may help to stabilize membranes during diel shifts in pH and other physicochemical conditions in the mats, were detected in the membranes of "Ca. Chloracidobacterium thermophilum."

We report results on circular dichroism (CD) measured on single immobilized chlorosomes of a triple mutant of green sulfur bacterium Chlorobaculum tepidum . The CD signal is measured by monitoring chlorosomal bacteriochlorphyll c fluorescence excited by alternate left and right circularly polarized laser light with a fixed wavelength of 733 nm. The excitation wavelength is close to a maximum of the negative CD signal of a bulk solution of the same chlorosomes. The average CD dissymmetry parameter obtained from an ensemble of individual chlorosomes was gs = -0.025, with an intrinsic standard deviation (due to variations between individual chlorosomes) of 0.006. The dissymmetry value is about 2.5 times larger than that obtained at the same wavelength in the bulk solution. The difference can be satisfactorily explained by taking into account the orientation factor in the single-chlorosome experiments. The observed distribution of the dissymmetry parameter reflects the well-ordered nature of the mutant chlorosomes.

The PscD subunit in the homodimeric "type I" photosynthetic reaction center (RC) complex of the green sulfur bacterium Chlorobium tepidum was disrupted by insertional mutagenesis of its relevant pscD gene. This is the first report on the use of the direct mutagenic approach into the RC-related genes in green sulfur bacteria. The RC complex of C. tepidum is supposed to form a homodimer of two identical PscA subunits together with three other subunits: PscB (F A /F B -containing protein), PscC (cytochrome c z ), and PscD. PscD shows a relatively low but significant similarity in its amino acid sequence to PsaD in the photosystem I of plants and cyanobacteria. We studied the biochemical and spectroscopic properties of a mutant lacking PscD in order to elucidate its unknown function. 1) The RC complex isolated from the mutant cells showed no band corresponding to PscD on SDS-PAGE analysis. 2) The growth rate of the PscD-less mutant was slower than that of the wildtype cells at low light intensities. 3) Time-resolved fluorescence spectra at 77 K revealed prolonged decay times of the fluorescence from bacteriochlorophyll c on the antenna chlorosome and from bacteriochlorophyll a on the Fenna-Matthews-Olson antenna protein in the mutant cells. The loss of PscD led to a much slower energy transfer from the antenna pigments to the special pair bacteriochlorophyll a (P840). 4) The mutant strain exhibited slightly less activity of ferredoxin-mediated NADP ؉ photoreduction compared with that in the wild-type strain. The extent of suppression, however, was less significant than that reported in the PsaD-less mutants of cyanobacterial photosystem I. The evolutionary relationship between PscD and PsaD was also discussed based on a structural homology modeling of the former.Photosynthetic organisms convert light energy into electrochemical free energy by carrying out a series of light-driven electron transfer reactions. This process, which is fundamental for life, is mediated by reaction center (RC) 1 complexes. The RCs are primarily grouped into two types based on their terminal electron acceptors, type I (FeS-type) RCs and type II (quinone-type) RCs. Purple photosynthetic bacteria contain only type II RCs, which do not evolve oxygen, whereas oxygenic cyanobacteria and plants utilize both type I (photosystem I) and type II (photosystem II) RCs, which are connected in-line through the b 6 f complex. Green sulfur bacteria and heliobacteria have unique type I RCs, so-called "homodimeric" RCs, which are made of two identical core polypeptides.The "heterodimeric" type I and II RCs, which are found in all photosynthetic organisms other than green sulfur bacteria and heliobacteria, consist of a set of two partially different core polypeptides that produce a slightly asymmetric arrangement of cofactors. A three-dimensional structure of the heterodimeric RC was first obtained from the type II RC of the purple bacterium Blastochloris viridis in 1985 (1). Recently, those of photosystem I and II RCs from the thermophilic cyanobacterium Sy...