Two pathways to understanding electron transfer in reaction centers from photosynthetic bacteria: A comparison of Rhodobacter sphaeroides and Rhodobacter capsulatus mutants.

The rates, yields, mechanisms and directionality of electron transfer (ET) are explored in twelve pairs of Rhodobacter (R.) sphaeroides and R. capsulatus mutant RCs designed to defeat ET from the excited primary donor (P*) to the A-side cofactors and re-direct ET to the normally inactive mirror-image B-side cofactors. In general, the R. sphaeroides variants have larger P+HB- yields (up to ∼90%) than their R. capsulatus analogs (up to ∼60%), where HB is the B-side bacteriopheophytin. Substitution of Tyr for Phe at L-polypeptide position L181 near BB primarily increases the contribution of fast P* â†’ P+BB- â†’ P+HB- two-step ET, where BB is the "bridging" B-side bacteriochlorophyll. The second step (∼6-8 ps) is slower than the first (∼3-4 ps), unlike A-side two-step ET (P* â†’ P+BA- â†’ P+HA-) where the second step (∼1 ps) is faster than the first (∼3-4 ps) in the native RC. Substitutions near HB, at L185 (Leu, Trp or Arg) and at M-polypeptide site M133/131 (Thr, Val or Glu), strongly affect the contribution of slower (20-50 ps) P* â†’ P+HB- one-step superexchange ET. Both ET mechanisms are effective in directing electrons "the wrong way" to HB and both compete with internal conversion of P* to the ground state (∼200 ps) and ET to the A-side cofactors. Collectively, the work demonstrates cooperative amino-acid control of rates, yields and mechanisms of ET in bacterial RCs and how A- vs. B-side charge separation can be tuned in both species.

Experimental and Theoretical Mutation of Exciton States on the Smallest Type-I Photosynthetic Reaction Center Complex of a Green Sulfur Bacterium Chlorobaclum tepidum.

The exciton states on the smallest type-I photosynthetic reaction center complex of a green sulfur bacterium Chlorobaculum tepidum (GsbRC) consisting of 26 bacteriochlorophylls a (BChl a) and four chlorophylls a (Chl a) located on the homodimer of two PscA reaction center polypeptides were investigated. This analysis involved the study of exciton states through a combination of theoretical modeling and the genetic removal of BChl a pigments at eight sites. (1) A theoretical model of the pigment assembly exciton state on GsbRC was constructed using Poisson TrESP (P-TrESP) and charge density coupling (CDC) methods based on structural information. The model reproduced the experimentally obtained absorption spectrum, circular dichroism spectrum, and excitation transfer dynamics, as well as explained the effects of mutation. (2) Eight BChl a molecules at different locations on the GsbRC were selectively removed by genetic exchange of the His residue, which ligates the central Mg atom of BChl a, with the Leu residue on either one or two PscAs in the RC. His locations are conserved among all type-I RC plant polypeptide, cyanobacteria, and bacteria amino acid sequences. (3) Purified mutant-GsbRCs demonstrated distinct absorption and fluorescence spectra at 77 K, which were different from each other, suggesting successful pigment removal. (4) The same mutations were applied to the constructed theoretical model to analyze the outcomes of these mutations. (5) The combination of theoretical predictions and experimental mutations based on structural information is a new tool for studying the function and evolution of photosynthetic reaction centers.

Spectral Properties of Chlorophyll f in the B800 Cavity of Light-harvesting Complex 2 from the Purple Photosynthetic Bacterium Rhodoblastus acidophilus.

The interactions of chlorophyll (Chl) and bacteriochlorophyll (BChl) pigments with the polypeptides in photosynthetic light-harvesting proteins are responsible for controlling the absorption energy of (B)Chls in protein matrixes. The binding pocket of B800 BChl a in LH2 proteins, which are peripheral light-harvesting proteins in purple photosynthetic bacteria, is useful for studying such structure-property relationships. We report the reconstitution of Chl f, which has the formyl group at the 2-position, in the B800 cavity of LH2 from the purple bacterium Rhodoblastus acidophilus. The Qy absorption band of Chl f in the B800 cavity was shifted by 14 nm to longer wavelength compared to that of the corresponding five-coordinated monomer in acetone. This redshift was larger than that of Chl a and Chl b. Resonance Raman spectroscopy indicated hydrogen bonding between the 2-formyl group of Chl f and the LH2 polypeptide. These results suggest that this hydrogen bonding contributes to the Qy redshift of Chl f. Furthermore, the Qy redshift of Chl f in the B800 cavity was smaller than that of Chl d. This may have arisen from the different patterns of hydrogen bonding between Chl f and Chl d and/or from the steric hindrance of the 3-vinyl group in Chl f.

Cryo-EM structure of the monomeric Rhodobacter sphaeroides RC-LH1 core complex at 2.5†Å.

Reaction centre light-harvesting 1 (RC-LH1) complexes are the essential components of bacterial photosynthesis. The membrane-intrinsic LH1 complex absorbs light and the energy migrates to an enclosed RC where a succession of electron and proton transfers conserves the energy as a quinol, which is exported to the cytochrome bc1 complex. In some RC-LH1 variants quinols can diffuse through small pores in a fully circular, 16-subunit LH1 ring, while in others missing LH1 subunits create a gap for quinol export. We used cryogenic electron microscopy to obtain a 2.5 Šresolution structure of one such RC-LH1, a monomeric complex from Rhodobacter sphaeroides. The structure shows that the RC is partly enclosed by a 14-subunit LH1 ring in which each αß heterodimer binds two bacteriochlorophylls and, unusually for currently reported complexes, two carotenoids rather than one. Although the extra carotenoids confer an advantage in terms of photoprotection and light harvesting, they could impede passage of quinones through small, transient pores in the LH1 ring, necessitating a mechanism to create a dedicated quinone channel. The structure shows that two transmembrane proteins play a part in stabilising an open ring structure; one of these components, the PufX polypeptide, is augmented by a hitherto undescribed protein subunit we designate as protein-Y, which lies against the transmembrane regions of the thirteenth and fourteenth LH1α polypeptides. Protein-Y prevents LH1 subunits 11-14 adjacent to the RC QB site from bending inwards towards the RC and, with PufX preventing complete encirclement of the RC, this pair of polypeptides ensures unhindered quinone diffusion.

Cryo-EM structure of the Rhodospirillum rubrum RC-LH1 complex at 2.5†Å.

The reaction centre light-harvesting 1 (RC-LH1) complex is the core functional component of bacterial photosynthesis. We determined the cryo-electron microscopy (cryo-EM) structure of the RC-LH1 complex from Rhodospirillum rubrum at 2.5 Šresolution, which reveals a unique monomeric bacteriochlorophyll with a phospholipid ligand in the gap between the RC and LH1 complexes. The LH1 complex comprises a circular array of 16 αß-polypeptide subunits that completely surrounds the RC, with a preferential binding site for a quinone, designated QP, on the inner face of the encircling LH1 complex. Quinols, initially generated at the RC QB site, are proposed to transiently occupy the QP site prior to traversing the LH1 barrier and diffusing to the cytochrome bc1 complex. Thus, the QP site, which is analogous to other such sites in recent cryo-EM structures of RC-LH1 complexes, likely reflects a general mechanism for exporting quinols from the RC-LH1 complex.

Circular dichroism and resonance Raman spectroscopies of bacteriochlorophyll b-containing LH1-RC complexes.

The core light-harvesting complexes (LH1) in bacteriochlorophyll (BChl) b-containing purple phototrophic bacteria are characterized by a near-infrared absorption maximum around 1010 nm. The determinative cause for this ultra-redshift remains unclear. Here, we present results of circular dichroism (CD) and resonance Raman measurements on the purified LH1 complexes in a reaction center-associated form from a mesophilic and a thermophilic Blastochloris species. Both the LH1 complexes displayed purely positive CD signals for their Qy transitions, in contrast to those of BChl a-containing LH1 complexes. This may reflect differences in the conjugation system of the bacteriochlorin between BChl b and BChl a and/or the differences in the pigment organization between the BChl b- and BChl a-containing LH1 complexes. Resonance Raman spectroscopy revealed remarkably large redshifts of the Raman bands for the BChl b C3-acetyl group, indicating unusually strong hydrogen bonds formed with LH1 polypeptides, results that were verified by a published structure. A linear correlation was found between the redshift of the Raman band for the BChl C3-acetyl group and the change in LH1-Qy transition for all native BChl a- and BChl b-containing LH1 complexes examined. The strong hydrogen bonding and π-π interactions between BChl b and nearby aromatic residues in the LH1 polypeptides, along with the CD results, provide crucial insights into the spectral and structural origins for the ultra-redshift of the long-wavelength absorption maximum of BChl b-containing phototrophs.

Mg-protoporphyrin IX monomethyl ester cyclase from Rhodobacter capsulatus: radical SAM-dependent synthesis of the isocyclic ring of bacteriochlorophylls.

During bacteriochlorophyll a biosynthesis, the oxygen-independent conversion of Mg-protoporphyrin IX monomethyl ester (Mg-PME) to protochlorophyllide (Pchlide) is catalyzed by the anaerobic Mg-PME cyclase termed BchE. Bioinformatics analyses in combination with pigment studies of cobalamin-requiring Rhodobacter capsulatus mutants indicated an unusual radical S-adenosylmethionine (SAM) and cobalamin-dependent BchE catalysis. However, in vitro biosynthesis of the isocyclic ring moiety of bacteriochlorophyll using purified recombinant BchE has never been demonstrated. We established a spectroscopic in vitro activity assay which was subsequently validated by HPLC analyses and H218O isotope label transfer onto the carbonyl-group (C-131-oxo) of the isocyclic ring of Pchlide. The reaction product was further converted to chlorophyllide in the presence of light-dependent Pchlide reductase. BchE activity was stimulated by increasing concentrations of NADPH or SAM, and inhibited by S-adenosylhomocysteine. Subcellular fractionation experiments revealed that membrane-localized BchE requires an additional, heat-sensitive cytosolic component for activity. BchE catalysis was not sustained in chimeric experiments when a cytosolic extract from E. coli was used as a substitute. Size-fractionation of the soluble R. capsulatus fraction indicated that enzymatic activity relies on a specific component with an estimated molecular mass between 3 and 10 kDa. A structure guided site-directed mutagenesis approach was performed on the basis of a three-dimensional homology model of BchE. A newly established in vivo complementation assay was used to investigate 24 BchE mutant proteins. Potential ligands of the [4Fe-4S] cluster (Cys204, Cys208, Cys211), of SAM (Phe210, Glu308 and Lys320) and of the proposed cobalamin cofactor (Asp248, Glu249, Leu29, Thr71, Val97) were identified.

Metal-based photosensitizers for photodynamic therapy: the future of multimodal oncology?

Photodynamic therapy (PDT) is an approved medical technique to treat certain forms of cancer. It has been used to complement traditional anticancer modalities such as surgery, chemotherapy or radiotherapy, and in certain cases, to replace these treatments. One critical parameter of PDT is the photosensitizer (PS); historically, a purely organic macrocyclic tetrapyrrole-based structure. This short review surveys two recent clinical examples of metal complexes, namely TOOKAD®-Soluble and TLD-1433, which have ideal photophysical properties to act as PDT PSs. We highlight the important role played by the metal ions in the PS for PDT activity.

Unusual features in the photosynthetic machinery of Halorhodospira halochloris DSM 1059 revealed by complete genome sequencing.

Halorhodospira halochloris is an anaerobic, halophilic, purple photosynthetic bacterium belonging to γ-Proteobacteria. H. halochloris is also characteristic as a thermophilic phototrophic isolate producing bacteriochlorophyll (BChl) b. Here, we report the complete genome sequence of H. halochloris DSM 1059. The genetic arrangement for this bacterium's photosynthetic apparatus is of particular interest; its genome contains two sets of puf operons encoding the reaction center and core light-harvesting 1 (LH1) complexes having almost identical nucleotide sequences (e.g., 98.8-99.9% of nucleotide identities between two sets of pufLM genes, but 100% of deduced amino acid sequence identities). This duplication of photosynthetic genes may provide a glimpse at natural selection in action. The ß-polypeptides of the LH1 complex in purple bacteria usually contain two histidine residues to bind BChl a; however, those of H. halochloris were revealed to have four histidine residues, indicating unusual pigment organization in the LH1 complex of this species. Like in other BChl b-producing phototrophs, the genome of H. halochloris lacks the divinyl reductase genes bciA and bciB. The phylogeny of chlorophyllide a oxidoreductase, which catalyzes committed steps in the synthesis of BChl a and BChl b, indicates that evolution toward BChl b production is convergent. Geranylgeranyl reductase (BchP) of H. halochloris has an insertion region in its primary structure, which could be important for its unusual sequential reduction reactions.

Cryo-EM structure of the Blastochloris viridis LH1-RC complex at 2.9 Å.

The light-harvesting 1-reaction centre (LH1-RC) complex is a key functional component of bacterial photosynthesis. Here we present a 2.9 Å resolution cryo-electron microscopy structure of the bacteriochlorophyll b-based LH1-RC complex from Blastochloris viridis that reveals the structural basis for absorption of infrared light and the molecular mechanism of quinone migration across the LH1 complex. The triple-ring LH1 complex comprises a circular array of 17 ß-polypeptides sandwiched between 17 α- and 16 γ-polypeptides. Tight packing of the γ-apoproteins between ß-polypeptides collectively interlocks and stabilizes the LH1 structure; this, together with the short Mg-Mg distances of bacteriochlorophyll b pairs, contributes to the large redshift of bacteriochlorophyll b absorption. The 'missing' 17th γ-polypeptide creates a pore in the LH1 ring, and an adjacent binding pocket provides a folding template for a quinone, Q P, which adopts a compact, export-ready conformation before passage through the pore and eventual diffusion to the cytochrome bc 1 complex.

Proposal for verifying dipole properties of light-harvesting antennas.

For light harvesters with a reaction center complex (LH1-RC complex) of three types, we propose an experiment to verify our analysis based upon antenna theories that automatically include the required structural information. Our analysis conforms to the current understanding of light-harvesting antennas in that we can explain known properties of these complexes. We provide an explanation for the functional roles of the notch at the light harvester, a functional role of the polypeptide called PufX or W at the opening, a functional role of the special pair, a reason that the cross section of the light harvester must not be circular, a reason that the light harvester must not be spherical, reasons for the use of dielectric bacteriochlorophylls instead of conductors to make the light harvester, a mechanism to prevent damage from excess sunlight, an advantage of the dimeric form, and reasons for the modular design of nature. Based upon our analysis we provide a mechanism for dimerization. We predict that the dimeric form of light-harvesting complexes is favored under intense sunlight. We further comment upon the classification of the dimeric or S-shape complexes. The S-shape complexes should not be considered as the third type of light harvester but simply as a composite form.

Structure of a symmetric photosynthetic reaction center-photosystem.

Reaction centers are pigment-protein complexes that drive photosynthesis by converting light into chemical energy. It is believed that they arose once from a homodimeric protein. The symmetry of a homodimer is broken in heterodimeric reaction-center structures, such as those reported previously. The 2.2-angstrom resolution x-ray structure of the homodimeric reaction center-photosystem from the phototroph Heliobacterium modesticaldum exhibits perfect C2 symmetry. The core polypeptide dimer and two small subunits coordinate 54 bacteriochlorophylls and 2 carotenoids that capture and transfer energy to the electron transfer chain at the center, which performs charge separation and consists of 6 (bacterio)chlorophylls and an iron-sulfur cluster; unlike other reaction centers, it lacks a bound quinone. This structure preserves characteristics of the ancestral reaction center, providing insight into the evolution of photosynthesis.

Quenching Capabilities of Long-Chain Carotenoids in Light-Harvesting-2 Complexes from Rhodobacter sphaeroides with an Engineered Carotenoid Synthesis Pathway.

Six light-harvesting-2 complexes (LH2) from genetically modified strains of the purple photosynthetic bacterium Rhodobacter (Rb.) sphaeroides were studied using static and ultrafast optical methods and resonance Raman spectroscopy. These strains were engineered to incorporate carotenoids for which the number of conjugated groups (N = NC═C + NC═O) varies from 9 to 15. The Rb. sphaeroides strains incorporate their native carotenoids spheroidene (N = 10) and spheroidenone (N = 11), as well as longer-chain analogues including spirilloxanthin (N = 13) and diketospirilloxantion (N = 15) normally found in Rhodospirillum rubrum. Measurements of the properties of the carotenoid first singlet excited state (S1) in antennas from the Rb. sphaeroides set show that carotenoid-bacteriochlorophyll a (BChl a) interactions are similar to those in LH2 complexes from various other bacterial species and thus are not significantly impacted by differences in polypeptide composition. Instead, variations in carotenoid-to-BChl a energy transfer are primarily regulated by the N-determined energy of the carotenoid S1 excited state, which for long-chain (N ≥ 13) carotenoids is not involved in energy transfer. Furthermore, the role of the long-chain carotenoids switches from a light-harvesting supporter (via energy transfer to BChl a) to a quencher of the BChl a S1 excited state B850*. This quenching is manifested as a substantial (∼2-fold) reduction of the B850* lifetime and the B850* fluorescence quantum yield for LH2 housing the longest carotenoids.

Perturbation of bacteriochlorophyll molecules in Fenna-Matthews-Olson protein complexes through mutagenesis of cysteine residues.

The Fenna-Matthews-Olson (FMO) pigment-protein complex in green sulfur bacteria transfers excitation energy from the chlorosome antenna complex to the reaction center. In understanding energy transfer in the FMO protein, the individual contributions of the bacteriochlorophyll pigments to the FMO complex's absorption spectrum could provide detailed information with which molecular and energetic models can be constructed. The absorption properties of the pigments, however, are such that their spectra overlap significantly. To overcome this, we used site-directed mutagenesis to construct a series of mutant FMO complexes in the model green sulfur bacterium Chlorobaculum tepidum (formerly Chlorobium tepidum). Two cysteines at positions 49 and 353 in the C. tepidum FMO complex, which reside near hydrogen bonds between BChls 2 and 3, and their amino acid binding partner serine 73 and tyrosine 15, respectively, were changed to alanine residues. The resulting C49A, C353A, and C49A C353A double mutants were analyzed with a combination of optical absorption and circular dichroism (CD) spectroscopies. Our results revealed changes in the absorption properties of several underlying spectral components in the FMO complex, as well as the redox behavior of the complex in response to the reductant sodium dithionite. A high-resolution X-ray structure of the C49A C353A double mutant reveals that these spectral changes appear to be independent of any major structural rearrangements in the FMO mutants. Our findings provide important tests for theoretical calculations of the C. tepidum FMO absorption spectrum, and additionally highlight a possible role for cysteine residues in the redox activity of the pigment-protein complex.

Optimizing multi-step B-side charge separation in photosynthetic reaction centers from Rhodobacter capsulatus.

Using high-throughput methods for mutagenesis, protein isolation and charge-separation functionality, we have assayed 40 Rhodobacter capsulatus reaction center (RC) mutants for their P(+)QB(-) yield (P is a dimer of bacteriochlorophylls and Q is a ubiquinone) as produced using the normally inactive B-side cofactors BB and HB (where B is a bacteriochlorophyll and H is a bacteriopheophytin). Two sets of mutants explore all possible residues at M131 (M polypeptide, native residue Val near HB) in tandem with either a fixed His or a fixed Asn at L181 (L polypeptide, native residue Phe near BB). A third set of mutants explores all possible residues at L181 with a fixed Glu at M131 that can form a hydrogen bond to HB. For each set of mutants, the results of a rapid millisecond screening assay that probes the yield of P(+)QB(-) are compared among that set and to the other mutants reported here or previously. For a subset of eight mutants, the rate constants and yields of the individual B-side electron transfer processes are determined via transient absorption measurements spanning 100 fs to 50 µs. The resulting ranking of mutants for their yield of P(+)QB(-) from ultrafast experiments is in good agreement with that obtained from the millisecond screening assay, further validating the efficient, high-throughput screen for B-side transmembrane charge separation. Results from mutants that individually show progress toward optimization of P(+)HB(-)→P(+)QB(-) electron transfer or initial P*→P(+)HB(-) conversion highlight unmet challenges of optimizing both processes simultaneously.

Recent Progress in Chemical Modifications of Chlorophylls and Bacteriochlorophylls for the Applications in Photodynamic Therapy.

Since photodynamic therapy emerged as a promising cancer treatment, the development of photosensitizers has gained great interest. In this context, the photosynthetic pigments, chlorophylls and bacteriochlorophylls, as excellent natural photosensitizers, attracted much attention. In effect, several (bacterio) chlorophyll-based phototherapeutic agents have been developed and (or are about to) enter the clinics. The aim of this review article is to give a survey of the advances in the synthetic chemistry of these pigments which have been made over the last decade, and which are pertinent to the application of their derivatives as photosensitizers for photodynamic therapy (PDT). The review focuses on the synthetic strategies undertaken to obtain novel derivatives of (bacterio)chlorophylls with both enhanced photosensitizing and tumorlocalizing properties, and also improved photo- and chemical stability. These include modifications of the C- 17-ester moiety, the isocyclic ring, the central binding pocket, and the derivatization of peripheral functionalities at the C-3 and C-7 positions with carbohydrate-, peptide-, and nanoparticle moieties or other residues. The effects of these modifications on essential features of the pigments are discussed, such as the efficiency of reactive oxygen species generation, photostability, phototoxicity and interactions with living organisms. The review is divided into several sections. In the first part, the principles of PDT and photosensitizer action are briefly described. Then the relevant photophysical features of (bacterio)chlorophylls and earlier approaches to their modification are summarized. Next, a more detailed overview of the progress in synthetic methods is given, followed by a discussion of the effects of these modifications on the photophysics of the pigments and on their biological activity.

All-atom structures and calcium binding sites of the bacterial photosynthetic LH1-RC core complex from Thermochromatium tepidum.

Computationally derived structures of the photosynthetic core complex composed of the light-harvesting (LH) system LH1 and the reaction center (RC) from a thermophilic purple sulfur bacterium Thermochromatium tepidum are reported providing first models of the LH1 system at atomic resolution. We used the known primary structure of α and ß polypeptides from this particular LH1 complex and the related bacterial LH templates to design the LH1 torus composed of 16 αß subunits trapping bacteriochlorophyll (BChl-a) dimers and carotenoid molecules. The macromolecule of RC was placed in the center of the ring and the LH1-RC complex was inserted inside the lipid bilayer to simulate the membrane environment. Since thermal stability of the LH1-RC complex is linked to Ca(2+) binding by the complex, location of trapping sites of calcium ions in the LH1 polypeptides is examined by using molecular dynamics simulations of the entire system solvated in water with CaCl2 molecules in the system. The newly predicted Ca(2+) trapping sites can be responsible for attractive interaction of neighboring αß subunits of LH1 with relevance to stability of the calcium-bound LH1-RC complex.

Versatile design of biohybrid light-harvesting architectures to tune location, density, and spectral coverage of attached synthetic chromophores for enhanced energy capture.

Biohybrid antennas built upon chromophore-polypeptide conjugates show promise for the design of efficient light-capturing modules for specific purposes. Three new designs, each of which employs analogs of the ß-polypeptide from Rhodobacter sphaeroides, have been investigated. In the first design, amino acids at seven different positions on the polypeptide were individually substituted with cysteine, to which a synthetic chromophore (bacteriochlorin or Oregon Green) was covalently attached. The polypeptide positions are at -2, -6, -10, -14, -17, -21, and -34 relative to the 0-position of the histidine that coordinates bacteriochlorophyll a (BChl a). All chromophore-polypeptides readily formed LH1-type complexes upon combination with the α-polypeptide and BChl a. Efficient energy transfer occurs from the attached chromophore to the circular array of 875 nm absorbing BChl a molecules (denoted B875). In the second design, use of two attachment sites (positions -10 and -21) on the polypeptide affords (1) double the density of chromophores per polypeptide and (2) a highly efficient energy-transfer relay from the chromophore at -21 to that at -10 and on to B875. In the third design, three spectrally distinct bacteriochlorin-polypeptides were prepared (each attached to cysteine at the -14 position) and combined in an ~1:1:1 mixture to form a heterogeneous mixture of LH1-type complexes with increased solar coverage and nearly quantitative energy transfer from each bacteriochlorin to B875. Collectively, the results illustrate the great latitude of the biohybrid approach for the design of diverse light-harvesting systems.

Bacteriochlorophyll homolog compositions in the bchU mutants of green sulfur bacteria.

Chlorosomes of the green sulfur bacterium Chlorobaculum limnaeum contain a large number of self-aggregated bacteriochlorophyll (BChl) e molecules. The ΔbchU mutant of this organism lacks BchU, a C20-methyltransferase, and therefore produces BChl f, which is the C20-unsubstituted form of BChl e. The BChl e homolog compositions, in terms of degrees of C8(2)-methylation, were not changed in the wild type during growth, while the BChl f homolog patterns in the mutant were significantly altered at various time periods of growth. BChl f with an isobutyl group at the C8 position was dominant at the early stage of growth, whereas the proportion of BChl f with the C8-ethyl group increased in the late exponential phase. We also constructed the ΔbchU mutant of C. tepidum which originally produces BChl c: the mutant therefore produces BChl d. BChl d homologs highly methylated at the C8(2) position also increased in the ΔbchU mutant of C. tedium compared to those in the wild type. These phenomena suggest that BchU interferes with the methylation ability of BchQ, a C8(2)-methyltransferase, and that the enzymes might compete in terms of obtaining S-adenosyl-methionine, the source of a methyl group. As a result, when grown to the late log phase, the ΔbchU mutant of C. limnaeum had similar heterogeneities of pigment homolog compositions compared to those in the wild type. Chlorosomes with a high proportion of C8-ethylated BChl homologs might be important for fine-tuning the light-harvesting or energy-transfer efficiency. Chlorosomes of the ΔbchU mutants at the various growth stages will be good materials for investigating effects of C8(2)-methylations on supramolecular structures of self-aggregated pigments.

Cyclopropane-ring formation in the acyl groups of chlorosome glycolipids is crucial for acid resistance of green bacterial antenna systems.

Green photosynthetic bacteria have unique light-harvesting antenna systems called chlorosomes. Chlorobaculum tepidum, a model organism of the bacteria, biosynthesized monogalactosyl- and rhamnosylgalactosyldiacylglycerides possessing a methylene-bridged palmitoleyl group characterized by a cis-substituted cyclopropane ring as the dominant glycolipids of its chlorosome surface. The formation of the cyclopropane ring was chemically inhibited by supplementation of sinefungin, an analog of S-adenosyl-L-methionine, into the bacterial cultivation. The presence of the cyclopropane ring reinforced acid resistance of the light-harvesting chlorosomes and suppressed acidic demetalation (pheophytinization) of bacteriochlorophyll-c pigments constructing the core part of chlorosomes. The ring-formation would represent direct and post-synthetic modifications of chlorosome membrane properties and was tolerant of acidic environments.