Heterogeneity of carotenoid content and composition in LH2 of the purple sulphur bacterium Allochromatium minutissimum grown under carotenoid-biosynthesis inhibition

The effects brought about by growing Allochromatium (Alc.) minutissimum in the presence of different concentrations of the carotenoid (Car) biosynthetic inhibitor diphenylamine (DPA) have been investigated. A decrease of Car content (from approximately 70% to >5%) in the membranes was accompanied by an increase of the percentage of (immature) Cars with reduced numbers of conjugated C=C bonds (from neurosporene to phytoene). Based on the obtained results and the analysis of literature data, the conclusion is reached that accumulation of phytoene during inhibition did not occur. Surprisingly, DPA inhibited phytoene synthase instead of phytoene desaturase as generally assumed. The distribution of Cars in peripheral antenna (LH2) complexes and their effect on the stability of LH2 has been investigated using absorption spectroscopy and HPLC analysis. Heterogeneity of Car composition and contents in the LH2 pool is revealed. The Car contents in LH2 varied widely from control levels to complete absence. According to common view, the assembly of LH2 occurs only in the presence of Cars. Here, we show that the LH2 can be assembled without any Cars. The presence of Cars, however, is important for structural stability of LH2 complexes.     

Evolution of low-light adapted peripheral light-harvesting complexes in strains of Rhodopseudomonas palustris

Purple bacteria have peripheral light-harvesting (PLH) complexes adapted to high-light (LH2) and low-light (LH3, LH4) growth conditions. The latter two have only been fully characterised in Rhodopseudomonas acidophila 7050 and Rhodopseudomonas palustris CGA009, respectively. It is known that LH4 complexes are expressed under the control of two light sensing bacteriophytochromes (BphPs). Recent genomic sequencing of a number of Rps. palustris strains has provided extensive information on PLH genes. We show that both LH3 and LH4 complexes are present in Rps. palustris and have evolved in the same operon controlled by the two adjacent BphPs. Two rare marker genes indicate that a gene cluster CL2, containing LH2 genes and the BphP RpBphP4, was internally transferred within the genome to form a new operon CL1. In CL1, RpBphP4 underwent gene duplication to RpBphP2 and RpBphP3, which evolved to sense light intensity rather than spectral red/far-red intensity ratio. We show that a second LH2 complex was acquired in CL1 belonging to a different PLH clade and these two PLH complexes co-evolved together into LH3 or LH4 complexes. The near-infrared spectra provide additional support for our conclusions on the evolution of PLH complexes based on genomic data.     

The LH2 complexes are assembled in the cells of purple sulfur bacterium Ectothiorhodospira haloalkaliphila with inhibition of carotenoid biosynthesis

The effect of the inhibitor of carotenoid (Car) biosynthesis, diphenylamine (DPA), on the cells of the purple sulfur bacterium Ectothiorhodospira (Ect.) haloalkaliphila has been studied. There occurs an inhibition of the biosynthesis of colored Cars (≥99 %) at 71 μM DPA. Considering “empty” Car pockets (Moskalenko and Makhneva 2012) the content of Cars in the DPA-treated samples is first calculated more correctly. The total content of the colored Cars in the sample at 71 μM DPA does not exceed 1 % of the wild type. In the DPA-treated cells (membranes) a complete set of pigment-protein complexes is retained. The LH2 complex at 71 μM DPA is isolated, which is identical to the LH2 complex of the wild type in near IR absorption spectra. This suggests that the principles for assembling this LH2 complex in vivo in the absence of colored Cars remain the same. These results are in full agreement with the data obtained earlier for Allochromatium (Alc.) minutissimum (Moskalenko and Makhneva 2012). They are as follows: (1) DPA almost entirely inhibits the biosynthesis of the colored Cars in Ect. haloalkaliphila cells. (2) In the DPA-treated samples non-colored Cars are detected at 53.25 μM DPA (as traces) and at 71 μM DPA. (3) DPA may affect both phytoene synthase (at ≤71 μM DPA) and phytoene desaturase (at ≥53.25 μM DPA). (4) The assembly of LH2 complex does occur without any colored Cars.     

Comparison of the fluorescence kinetics of detergent-solubilized and membrane-reconstituted LH2 complexes from Rps. acidophila and Rb. sphaeroides

Picosecond time-resolved fluorescence spectroscopy has been used in order to compare the fluorescence kinetics of detergent-solubilized and membrane-reconstituted light-harvesting 2 (LH2) complexes from the purple bacteria Rhodopseudomonas (Rps.) acidophila and Rhodobacter (Rb.) sphaeroides. LH2 complexes were reconstituted in phospholipid model membranes at different lipid:protein-ratios and all samples were studied exciting with a wide range of excitation densities. While the detergent-solubilized LH2 complexes from Rps. acidophila showed monoexponential decay kinetics (τ_f = 980 ps) for excitation densities of up to 3·10¹³ photons/(pulse·cm²), the membrane-reconstituted LH2 complexes showed multiexponential kinetics even at low excitation densities and high lipid:protein-ratios. The latter finding indicates an efficient clustering of LH2 complexes in the phospholipid membranes. Similar results were obtained for the LH2 complexes from Rb. sphaeroides. Guest editor: Dr. Conrad Mullineaux.     

Conjugation-length dependence of the T1 lifetimes of carotenoids free in solution and incorporated into the LH2, LH1, RC, and RC-LH1 complexes: possible mechanisms of triplet-energy dissipation

In addition to the roles of antioxidant and spacer, carotenoids (Cars) in purple photosynthetic bacteria pursue two physiological functions, i.e., light harvesting and photoprotection. To reveal the mechanisms of the photoprotective function, i.e., quenching triplet bacteriochlorophyll to prevent the sensitized generation of singlet oxygen, the triplet absorption spectra were recorded for Cars, where the number of conjugated double bonds (n) is in the region of 9-13, to determine the dependence on n of the triplet lifetime. The Cars examined include those in (a) solution; (b) the reconstituted LH1 complexes; (c) the native LH2 complexes from Rba. sphaeroides G1C, Rba. sphaeroides 2.4.1, Rsp. molischianum, and Rps. acidophila 10050; (d) the RCs from Rba. sphaeroides G1C, Rba. sphaeroides 2.4.1, and Rsp. rubrum S1; and (e) the RC-LH1 complexes from Rba. sphaeroides G1C, Rba. sphaeroides 2.4.1, Rsp. molischianum, Rps. acidophila 10050, and Rsp. rubrum S1. The results lead us to propose the following mechanisms: (i) A substantial shift of the linear dependence to shorter lifetimes on going from solution to the LH2 complex was ascribed to the twisting of the Car conjugated chain. (ii) A substantial decrease in the slope of the linear dependence on going from the reconstituted LH1 to the LH1 component of the RC-LH1 complex was ascribed to the minor-component Car forming a leak channel of triplet energy. (iii) The loss of conjugation-length dependence on going from the isolated RC to the RC component of the RC-LH1 complex was ascribed to the presence of a triplet-energy reservoir consisting of bacteriochlorophylls in the RC component.     

体外组装类胡萝卜素对外周捕光复合体(LH2)能量传递活性的影响

【目的】探求不产氧光合细菌(APB)外周捕光复合体(LH2)中类胡萝卜素(Car)结构和能量传递效率的关系和规律。【方法】通过二苯胺(DPA)抑制Car合成的方法从固氮红细菌134K20中获得部分缺失Car的LH2 (LC-LH2);采用TLC和HPLC法从3种APB中制备球形烯(SE)、玫红品(RP)和奥氏酮(OK) 3种Car;在含0.1%十二烷基二甲基胺氧化物(LDAO)的10 mmol/L Tris-HCl (pH 8.0)缓冲液中采用超声孵育法分别将这3种Car与LC-LH2体外组装,采用吸收光谱法、拉曼光谱法和荧光光谱法对组装LH2进行结构与功能分析。【结果】制备的部分缺失Car的LH2中,SE缺失率约为64.7%。这3种共轭长度、取代基的极性不同的Car均能与这种部分缺失SE的LH2自组装,Car组装率约在24.0%?29.4%之间,其中SE和OK的组装率高于RP。与部分缺失Car LH2中原有SE构象一致,重组的Car在LH2中也呈现较为伸展的平面构象。LH2中重组Car到细菌叶绿素(BChl)的能量传递效率由高到低的顺序依次为SE-LH2>RP-LH2>OK-LH2,与Car共轭体系大小的关系一致,而与Car极性大小没有明显的关系。【结论】在组装的LH2中Car采用平面构象与脱辅基蛋白结合,Car共轭长度仍是决定和影响LH2中Car-BChl能量传递效率的主要因素,而Car的取代基和极性影响较小。     

Chemical composition of the lipopolysaccharides of Rhodobacter sulfidophilus,Rhodopseudomonas acidophila,and Rhodopseudomonas blastica

The lipopolysaccharides of Rhodobacter sulfidophilus and the two budding species Rhodopseudomonas acidophila and Rhodopseudomonas blastica were isolated and chemically analyzed. The all have a lipid A backbone structure with glucosamine as the only amino sugar. The lipid A's of Rb. sulfidophilus and Rps. blastica contain phosphate, their fatty acids are characterized by ester-linked, unsubstituted 3-OH-10:0 and amide-linked 3-OH-14:0 (Rb. sulfidophilus) or 3-oxo-14:0 (Rps. blastica). Lipid A of Rps. acidophila is free of phosphate and contains the rare 3-OH-16:0 fatty acid in amide linkage.The lipopolysaccharides of all three species contain 2-keto-3-deoxy-octonate (KDO) but are devoid of heptoses. Neutral sugars with the exception of glucose are lacking in the lipopolysaccharide of Rb. sulfidophilus. This shows a high galacturonic acid content. The lipopolysaccharides of Rps. acidophila and Rps. blastica have neutral sugar spectra indicative for typical O-chains (rhamnose, mannose, galactose, glucose in both species, and in Rps. blastica additionally 2-O-methyl-6-deoxy-hexose). The taxonomic value of the data is discussed.This paper is dedicated to Prof. Dr. Norbert Pfennig on the occasion of his 60th birthday     

Identification of protein W,the elusive sixth subunit of the Rhodopseudomonas palustris reaction center-light harvesting 1 core complex

The X-ray crystal structure of the Rhodopseudomonas (Rps.) palustris reaction center-light harvesting 1 (RC-LH1) core complex revealed the presence of a sixth protein component, variably referred to in the literature as helix W, subunit W or protein W. The position of this protein prevents closure of the LH1 ring, possibly to allow diffusion of ubiquinone/ubiquinol between the RC and the cytochrome bc₁ complex in analogous fashion to the well-studied PufX protein from Rhodobacter sphaeroides. The identity and function of helix W have remained unknown for over 13 years; here we use a combination of biochemistry, mass spectrometry, molecular genetics and electron microscopy to identify this protein as RPA4402 in Rps. palustris CGA009. Protein W shares key conserved sequence features with PufX homologs, and although a deletion mutant was able to grow under photosynthetic conditions with no discernible phenotype, we show that a tagged version of protein W pulls down the RC-LH1 complex. Protein W is not encoded in the photosynthesis gene cluster and our data indicate that only approximately 10% of wild-type Rps. palustris core complexes contain this non-essential subunit; functional and evolutionary consequences of this observation are discussed. The ability to purify uniform RC-LH1 and RC-LH1-protein W preparations will also be beneficial for future structural studies of these bacterial core complexes.     

Membrane Curvature Induced by Aggregates of LH2s and Monomeric LH1s

The photosynthetic apparatus of purple bacteria is contained within organelles called chromatophores, which form as extensions of the cytoplasmic membrane. The shape of these chromatophores can be spherical (as in Rhodobacter sphaeroides), lamellar (as in Rhodopseudomonas acidophila and Phaeospirillum molischianum), or tubular (as in certain Rb. sphaeroides mutants). Chromatophore shape is thought to be influenced by the integral membrane proteins Light Harvesting Complexes I and II (LH1 and LH2), which pack tightly together in the chromatophore. It has been suggested that the shape of LH2, together with its close packing in the membrane, induces membrane curvature. The mechanism of LH2-induced curvature is explored via molecular dynamics simulations of multiple LH2 complexes in a membrane patch. LH2s from three species—Rb. sphaeroides, Rps. acidophila, and Phsp. molischianum—were simulated in different packing arrangements. In each case, the LH2s pack together and tilt with respect to neighboring LH2s in a way that produces an overall curvature. This curvature appears to be driven by a combination of LH2's shape and electrostatic forces that are modulated by the presence of well-conserved cytoplasmic charged residues, the removal of which inhibits LH2 curvature. The interaction of LH2s and an LH1 monomer is also explored, and it suggests that curvature is diminished by the presence of LH1 monomers. The implications of our results for chromatophore shape are discussed.     

Triplet Excitation Transfer between Carotenoids in the LH2 Complex from Photosynthetic Bacterium Rhodopseudomonas palustris

We have studied, by means of sub-microsecond time-resolved absorption spectroscopy, the triplet-excited state dynamics of carotenoids (Cars) in the intermediate-light adapted LH2 complex (ML-LH2) from Rhodopseudomonas palustris containing Cars with different numbers of conjugated double bonds. Following pulsed photo-excitation at 590 nm at room temperature, rapid spectral equilibration was observed either as a red shift of the isosbestic wavelength on a time scale of 0.6-1.0 mus, or as a fast decay in the shorter-wavelength side of the T(n)<--T(1) absorption of Cars with a time constant of 0.5-0.8 mus. Two major spectral components assignable to Cars with 11 and 12 conjugated double bonds were identified. The equilibration was not observed in the ML-LH2 at 77 K, or in the LH2 complex from Rhodobacter sphaeroides G1C containing a single type of Car. The unique spectral equilibration was ascribed to temperature-dependent triplet excitation transfer among different Car compositions. The results suggest that Cars of 11 and 12 conjugated bonds, both in close proximity of BChls, may coexist in an alpha,beta-subunit of the ML-LH2 complex.     

Sulfide utilization by purple nonsulfur bacteria

Summary The purple nonsulfur bacteria Rhodospirillum rubrum SMG 107, Rhodopseudomonas capsulata SMG 155, Rps. sphaeroides SMG 158 and Rps. palustris SMG 124 were tested for a possible utilization of sulfide. The first three strains were found to oxidize sulfide to extracellular elemental sulfur only, whereas Rps. palustris SMG 124 converted sulfide into sulfate without intermediate accumulation of elemental sulfur. Growth ceased at lower sulfide concentrations than usually found with purple sulfur bacteria. In consequence of the low sulfide tolerance information on the specific growth rates obtainable with sulfide as photosynthetic electron donor could not be provided by cultivation in batch cultures. Sulfide-limited chemostat cultures of Rps. capsulata SMG 155 showed that the maximum specific growth rate was close to 0.14 h^-1 (doubling time 5 h). Sulfide was converted into extracellular elemental sulfur at all dilution rates tested. The maximum specific growth rate of Rps. palustris SMG 124 was found to be much lower (less than 0.03 h^-1). Sulfate was the only product of the conversion of sulfide.These data show that at least some purple nonsulfur bacteria may play a role in the dissimilatory sulfur cycle in nature. Taxonomic implications of our results are discussed.Abbreviation SMG Sammlung für Mikroorganismen, Göttingen     

Single-Molecule Spectroscopy Unmasks the Lowest Exciton State of the B850 Assembly in LH2 from Rps. acidophila

We have recorded fluorescence-excitation and emission spectra from single LH2 complexes from Rhodopseudomonas (Rps.) acidophila. Both types of spectra show strong temporal spectral fluctuations that can be visualized as spectral diffusion plots. Comparison of the excitation and emission spectra reveals that for most of the complexes the lowest exciton transition is not observable in the excitation spectra due to the cutoff of the detection filter characteristics. However, from the spectral diffusion plots we have the full spectral and temporal information at hand and can select those complexes for which the excitation spectra are complete. Correlating the red most spectral feature of the excitation spectrum with the blue most spectral feature of the emission spectrum allows an unambiguous assignment of the lowest exciton state. Hence, application of fluorescence-excitation and emission spectroscopy on the same individual LH2 complex allows us to decipher spectral subtleties that are usually hidden in traditional ensemble spectroscopy.     

Single-Molecule Spectroscopy Unmasks the Lowest Exciton State of the B850 Assembly in LH2 from Rps. acidophila

We have recorded fluorescence-excitation and emission spectra from single LH2 complexes from Rhodopseudomonas (Rps.) acidophila. Both types of spectra show strong temporal spectral fluctuations that can be visualized as spectral diffusion plots. Comparison of the excitation and emission spectra reveals that for most of the complexes the lowest exciton transition is not observable in the excitation spectra due to the cutoff of the detection filter characteristics. However, from the spectral diffusion plots we have the full spectral and temporal information at hand and can select those complexes for which the excitation spectra are complete. Correlating the red most spectral feature of the excitation spectrum with the blue most spectral feature of the emission spectrum allows an unambiguous assignment of the lowest exciton state. Hence, application of fluorescence-excitation and emission spectroscopy on the same individual LH2 complex allows us to decipher spectral subtleties that are usually hidden in traditional ensemble spectroscopy.     

Intrinsic curvature properties of photosynthetic proteins in chromatophores

In purple bacteria, photosynthesis is carried out on large indentations of the bacterial plasma membrane termed chromatophores. Acting as primitive organelles, chromatophores are densely packed with the membrane proteins necessary for photosynthesis, including light harvesting complexes LH1 and LH2, reaction center (RC), and cytochrome bc₁. The shape of chromatophores is primarily dependent on species, and is typically spherical or flat. How these shapes arise from the protein-protein and protein-membrane interactions is still unknown. Now, using molecular dynamics simulations, we have observed the dynamic curvature of membranes caused by proteins in the chromatophore. A membrane-embedded array of LH2s was found to relax to a curved state, both for LH2 from Rps. acidophila and a homology-modeled LH2 from Rb. sphaeroides. A modeled LH1-RC-PufX dimer was found to develop a bend at the dimerizing interface resulting in a curved shape as well. In contrast, the bc₁ complex, which has not been imaged yet in native chromatophores, did not induce a preferred membrane curvature in simulation. Based on these results, a model for how the different photosynthetic proteins influence chromatophore shape is presented.     

Energy transfer in purple bacterial photosynthetic units from cells grown in various light intensities

Three photosynthetic membranes, called intra-cytoplasmic membranes (ICMs), from wild-type and the ?pucBA_abce mutant of the purple phototrophic bacterium Rps. palustris were investigated using optical spectroscopy. The ICMs contain identical light-harvesting complex 1–reaction centers (LH1–RC) but have various spectral forms of light-harvesting complex 2 (LH2). Spectroscopic studies involving steady-state absorption, fluorescence, and femtosecond time-resolved absorption at room temperature and at 77 K focused on inter-protein excitation energy transfer. The studies investigated how energy transfer is affected by altered spectral features of the LH2 complexes as those develop under growth at different light conditions. The study shows that LH1 → LH2 excitation energy transfer is strongly affected if the LH2 complex alters its spectroscopic signature. The LH1 → LH2 excitation energy transfer rate modeled with the Förster mechanism and kinetic simulations of transient absorption of the ICMs demonstrated that the transfer rate will be 2–3 times larger for ICMs accumulating LH2 complexes with the classical B800–850 spectral signature (grown in high light) compared to the ICMs from the same strain grown in low light. For the ICMs from the ?pucBA_abce mutant, in which the B850 band of the LH2 complex is blue-shifted and almost degenerate with the B800 band, the LH1 → LH2 excitation energy transfer was not observed nor predicted by calculations.     

Construction of hybrid photosynthetic units using peripheral and core antennae from two different species of photosynthetic bacteria: detection of the energy transfer from bacteriochlorophyll a in LH2 to bacteriochlorophyll b in LH1

Typical purple bacterial photosynthetic units consist of supra-molecular arrays of peripheral (LH2) and core (LH1-RC) antenna complexes. Recent atomic force microscopy pictures of photosynthetic units in intact membranes have revealed that the architecture of these units is variable (Scheuring et al. (2005) Biochim Bhiophys Acta 1712:109–127). In this study, we describe methods for the construction of heterologous photosynthetic units in lipid-bilayers from mixtures of purified LH2 (from Rhodopseudomonas acidophila) and LH1-RC (from Rhodopseudomonas viridis) core complexes. The architecture of these reconstituted photosynthetic units can be varied by controlling ratio of added LH2 to core complexes. The arrangement of the complexes was visualized by electron-microscopy in combination with Fourier analysis. The regular trigonal array of the core complexes seen in the native photosynthetic membrane could be regenerated in the reconstituted membranes by temperature cycling. In the presence of added LH2 complexes, this trigonal symmetry was replaced with orthorhombic symmetry. The small lattice lengths for the latter suggest that the constituent unit of the orthorhombic lattice is the LH2. Fluorescence and fluorescence-excitation spectroscopy was applied to the set of the reconstituted membranes prepared with various proportions of LH2 to core complexes. Remarkably, even though the LH2 complexes contain bacteriochlorophyll a, and the core complexes contain bacteriochlorophyll b, it was possible to demonstrate energy transfer from LH2 to the core complexes. These experiments provide a first step along the path toward investigating how changing the architecture of purple bacterial photosynthetic units affects the overall efficiency of light-harvesting.     

In silico predictions of LH2 ring sizes from the crystal structure of a single subunit using molecular dynamics simulations

Most of the currently known light‐harvesting complexes 2 (LH2) rings are formed by 8 or 9 subunits. As of now, questions like “what factors govern the LH2 ring size?” and “are there other ring sizes possible?” remain largely unanswered. Here, we investigate by means of molecular dynamics (MD) simulations and stochastic modeling the possibility of predicting the size of an LH2 ring from the sole knowledge of the high resolution crystal structure of a single subunit. Starting with single subunits of two LH2 rings with known size, that is, an 8‐ring from Rs. moliscianum (MOLI) and a 9‐ring from Rps. acidophila (ACI), and one with unknown size (referred to as X), we build atomic models of subunit dimers corresponding to assumed 8‐, 9‐, and 10‐ring geometries. After inserting each of the dimers into a lipid‐water environment, we determine the preferred angle between the corresponding subunits by three methods: (1) energy minimization, (2) free MD simulations, and (3) potential of mean force calculations. We find that the results from all three methods are consistent with each other, and when taken together, it allows one to predict with reasonable level of confidence the sizes of the corresponding ring structures. One finds that X and ACI very likely form a 9‐ring, while MOLI is more likely to form an 8‐ring than a 9‐ring. Finally, we discuss both the merits and limitations of all three prediction methods. Proteins 2011; © 2011 Wiley‐Liss, Inc.     

Progress toward a biomimetic leaf: 4,000 h of hydrogen production by coating‐stabilized nongrowing photosynthetic Rhodopseudomonas palustris

Intact cells are the most stable form of nature's photosynthetic machinery. Coating‐immobilized microbes have the potential to revolutionize the design of photoabsorbers for conversion of sunlight into fuels. Multi‐layer adhesive polymer coatings could spatially combine photoreactive bacteria and algae (complementary biological irradiance spectra) creating high surface area, thin, flexible structures optimized for light trapping, and production of hydrogen (H₂) from water, lignin, pollutants, or waste organics. We report a model coating system which produced 2.08 ± 0.01 mmol H₂ m^?2 h^?1 for 4,000 h with nongrowing Rhodopseudomonas palustris, a purple nonsulfur photosynthetic bacterium. This adhesive, flexible, nanoporous Rps. palustris latex coating produced 8.24 ± 0.03 mol H₂ m^?2 in an argon atmosphere when supplied with acetate and light. A simple low‐pressure hydrogen production and trapping system was tested using a 100 cm² coating. Rps. palustris CGA009 was combined in a bilayer coating with a carotenoid‐less mutant of Rps. palustris (CrtI^?) deficient in peripheral light harvesting (LH2) function. Cryogenic field emission gun scanning electron microscopy (cryo‐FEG‐SEM) and high‐pressure freezing were used to visualize the microstructure of hydrated coatings. A light interaction and reactivity model was evaluated to predict optimal coating thickness for light absorption using the Kubelka‐Munk theory (KMT) of reflectance and absorptance. A two‐flux model predicted light saturation thickness with good agreement to observed H₂ evolution rate. A combined materials and modeling approach could be used for guiding cellular engineering of light trapping and reactivity to enhance overall photosynthetic efficiency per meter square of sunlight incident on photocatalysts. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010     

Excitation energy transfer in chlorosomes of Chlorobium phaeobacteroides strain CL1401: the role of carotenoids

The role of carotenoids in chlorosomes of the green sulfur bacterium Chlorobium phaeobacteroides, containing bacteriochlorophyll (BChl) e and the carotenoid (Car) isorenieratene as main pigments, was studied by steady-state fluorescence excitation, picosecond single-photon timing and femtosecond transient absorption (TA) spectroscopy. In order to obtain information about energy transfer from Cars in this photosynthetic light-harvesting antenna with high spectral overlap between Cars and BChls, Car-depleted chlorosomes, obtained by inhibition of Car biosynthesis by 2-hydroxybiphenyl, were employed in a comparative study with control chlorosomes. Excitation spectra measured at room temperature give an efficiency of 60–70% for the excitation energy transfer from Cars to BChls in control chlorosomes. Femtosecond TA measurements enabled an identification of the excited state absorption band of Cars and the lifetime of their S₁ state was determined to be 10 ps. Based on this lifetime, we concluded that the involvement of this state in energy transfer is unlikely. Furthermore, evidence was obtained for the presence of an ultrafast (>100 fs) energy transfer process from the S₂ state of Cars to BChls in control chlorosomes. Using two time-resolved techniques, we further found that the absence of Cars leads to overall slower decay kinetics probed within the Q_y band of BChl e aggregates, and that two time constants are generally required to describe energy transfer from aggregated BChl e to baseplate BChl a.This revised version was published online in October 2005 with corrections to the Cover Date.     

Selective binding of carotenoids with a shorter conjugated chain to the LH2 antenna complex and those with a longer conjugated chain to the reaction center from Rubrivivax gelatinosus

Rubrivivax gelatinosus having both the spheroidene and spirilloxanthin biosynthetic pathways produces carotenoids (Cars) with a variety of conjugated chains, which consist of different numbers of conjugated double bonds (n), including the C=C (m) and C=O (o) bonds. When grown under anaerobic conditions, the wild type produces Cars for which n = m = 9-13, whereas under semiaerobic conditions, it additionally produces Cars for which n = m + o = 10 + 1, 13 + 1, and 13 + 2. On the other hand, a mutant, in which the latter pathway is genetically blocked, produces only Cars for which n = 9 and 10 under anaerobic conditions and n = 9, 10, and 10 + 1 under semianaerobic conditions. Those Cars that were extracted from the LH2 complex (LH2) and the reaction center (RC), isolated from the wild-type and the mutant Rvi. gelatinosus, were analyzed by HPLC, and their structures were determined by mass spectrometry and 1H NMR spectroscopy. The selective binding of Cars to those pigment-protein complexes has been characterized as follows. (1) Cars with a shorter conjugated chain are selectively bound to LH2 whereas Cars with a longer conjugated chain to the RC. (2) Shorter chain Cars with a hydroxyl group are bound to LH2 almost exclusively. This rule holds either in the absence or in the presence of the keto group. The natural selection of shorter chain Cars by LH2 and longer chain Cars by the RC is discussed, on the basis of the results now available, in relation to the light-harvesting and photoprotective functions of Cars.