Giant circular dichroism of chlorosomes fromChloroflexus aurantiacus treated with 1-hexanol and proteolytic enzymes

The circular dichroism (CD) spectrum of isolated chlorosomes fromChloroflexus aurantiacus showed a conservative, S-shaped signal with a negative maximum at 723 nm, a positive maximum at 750 nm and a zero-crossing at 740 nm. Proteolytic treatment of chlorosomes with trypsin at 37°C did not change the CD signal or the absorption spectrum in contrast to treatment with proteinase K, where a twofold increase in rotational strength and a slight decrease of the absorption band at 740 nm were observed. Treatment with saturating 1-hexanol concentrations resulted in a blue shift of the absorption band at 740 nm as well as in changes of the CD spectrum. These changes reversed when the sample was diluted to half the saturating 1-hexanol concentration. In contrast to that, we observed an irreversible formation of a giant CD signal using the combination of 1-hexanol and proteinase K treatment. Electron micrographs of chlorosomes treated with both 1-hexanol and proteinase K showed large aggregates of multiple chlorosome size. By comparison of proteinase K induced effects with trypsin effects it appeared that the 5.7 kDa polypeptide has a structural role in the organisation of BChlc in the chlorosome.     

Characterization of the chlorosome antenna of the filamentous anoxygenic phototrophic bacterium<Emphasis Type="Italic"> Chloronema</Emphasis> sp. strain UdG9001

The absorption and fluorescence properties of chlorosomes of the filamentous anoxygenic phototrophic bacterium Chloronema sp. strain UdG9001 were analyzed. The chlorosome antenna of Chloronema consists of bacteriochlorophyll (BChl) d and BChl c together with -carotene as the main carotenoid. HPLC analysis combined with APCI LC-MS/MS showed that the chlorosomal BChls comprise a highly diverse array of homologues that differ in both the degree of alkylation of the macrocycle at C-8 and/or C-12 and the alcohol moiety esterified to the propionic acid group at C-17. BChl c and BChl d from Chloronema were mainly esterified with geranylgeraniol (33% of the total), heptadecanol (24%), octadecenol (19%), octadecanol (14%), and hexadecenol (9%). Despite this pigment heterogeneity, fluorescence emission of the chlorosomes showed a single peak centered at 765 nm upon excitation at wavelengths ranging from 710 to 740 nm. This single emission, assigned to BChl c, indicates an energy transfer from BChl d to BChl c within the same chlorosome. Likewise, incubation of chlorosomes under reducing conditions caused a weak increase in fluorescence emission, which indicates a small redox-dependent fluorescence. Finally, protein analysis of Chloronema chlorosomes using SDS-PAGE and MALDI-TOF-MS revealed the presence of a chlorosomal polypeptide with a molecular mass of 5.7 kDa, resembling the CsmA protein found in Chloroflexus aurantiacus and Chlorobium tepidum chlorosomes. Several minor polypeptides were also detected but not identified. These results indicate that, compared with other members of filamentous anoxygenic phototrophic bacteria and green sulfur bacteria, Chloronema possesses an antenna system with novel features that may be of interest for further investigations.Abbreviations APCI LC-MS/MS Atmospheric pressure chemical ionization liquid chromatography mass spectrometry - BChl Bacteriochlorophyll - Chl. Chlorobium - Cfl. Chloroflexus - MALDI-TOF-MS Matrix assisted laser desorption/ionization time-of-flight mass spectrometry - [Et] Ethyl - [i-Bu] Isobutyl - [Me] Methyl - [neo-Pent] Neopentyl - [n-Pr] Propyl - t _R Retention time     

Structures of chlorosomes and aggregated BChlc inChlorobium tepidum from solid state high resolution CP/MAS13C NMR

Cross polarization/magic angle spinning (CP/MAS)¹³C (solid state high resolution) NMR spectra were observed for chlorosomes and BChlc aggregates. Similarity of both kinds of spectra (except for some signals assignable to proteins and lipids in chlorosomes) indicates that BChlc's in chlorosomes are present just as in synthetic BChlc aggregates. Chemical shifts for C13¹ carbonyl and C3¹ hydroxylethyl carbons indicate hydrogen bonding between them. Comparison of solution and solid state¹³C NMR chemical shifts shows the five coordinated nature of BChlc aggregates. Some chemical shift differences were attributable to ring currents shifts. Their comparisons with calculated ring current shift values predicted structures for the aggregates. Cross polarization dynamics of the CP/MAS¹³C NMR signals explored dynamic and structural nature of the BChlc aggregates.     

Self quenching of chlorosome chlorophylls in water and hexanol-saturated water

The optical properties of a methyl ester homolog of bacteriochlorophylld (BChld _M ) and bacteriochlorophyllc (BChlc) in H₂O, hexanol-saturated H₂O and methanol were studied by absorption, fluorescence emission, and circular dichroism (CD). In H₂O, BChld _M spontaneously forms an aggregate similar to that formed in hexane, with absorption maximum at 730 nm and fluorescence emission at 748 nm. For the pigment sample in hexanol-saturated H₂O, while the absorption peaks at 661 nm, only slightly red-shifted compared to the monomer, the fluorescence emission is highly quenched. When diluted 2–3 fold with H₂O, the absorption returns to around 720 nm, characteristic of an aggregate. The CD spectrum of the H₂O aggregate exhibits a derivative-shaped feature with positive and negative peaks, while the amplitude is lower than that of chlorosomes. The Fourier transform infrared spectra of BChld _M aggregates in H₂O and hexane were measured. A 1644 cm^–1 band, indicative of a bonded 13¹-keto group, is detected for both samples. A marker band for 5-coordinated Mg was observed at 1611 cm^–1 for the two samples as well. To study the kinetic behavior of the samples, both single-photon counting (SPC) fluorescence and transient absorption difference spectroscopic measurements were performed. For BChld _M in hexanol-saturated H₂O, a fast decay component with a lifetime of 10 to 14 ps was detected using the two different techniques. The fast decay could be explained by the concentration quenching phenomenon due to a high local pigment concentration. For the pigment sample in H₂O, SPC gave a 16 ps component, whereas global analysis of transient absorption data generated two fast components: 3.5 and 26 ps. The difference may arise from the different excitation intensities. With a much higher excitation in the latter measurements, other quenching processes, e.g. annihilation, might be introduced, giving the 3.5 ps component. Finally, atomic force microscopy was used to examine the ultrastructure of BChld _M in H₂O and hexanol-saturated H₂O. Pigment clusters with diameters ranging from 15 to 45 nm were observed in both samples.     

The effect of detergent on the structure and composition of chlorosomes isolated from Chloroflexus aurantiacus

Isolated chlorosomes, treated with the detergent lithium dodecyl sulfate (LDS), can be separated into two green fractions by agarose gel electrophoresis. One fraction contains chlorosomes with a full complement of proteins and antenna BChl c absorbing at 740 nm, but with a more spherical form than the normal ellipsoid shape observed in control chlorosomes. The second fraction was completely devoid of proteins but had a similar absorption spectrum. Electron micrographs of the protein-free fraction indicated the presence of stain-excluding spheres with overall dimensions resembling those of intact chlorosomes (40–100 nm). These spheres are probably micelles of BChl c liberated from the chlorosomes during the detergent treatment, since similar structures could be produced when purified BChl c, dissolved in 1-hexanol, was dispersed in buffer, producing an aggregate absorbing at 742 nm. These results suggest that the chlorosome proteins are not required to produce an arrangement of BChl c chromophores which gives rise to a 740 nm absorption peak resembling that of intact chlorosomes. It seems probable, however, that proteins have a role in determining the overall shape of the chlorosome. Treatment with cross-linking reagents did not prevent the detergent-induced changes in chlorosome morphology.Abbreviations BChl bacteriochlorophyll - DSP dithiobis-succinimidyl-2-propionate - EM electron microscopy - LDS lithium dodecyl sulfate - MGDG monogalactosyl diacylglycerol - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis     

Molecular organization of bacteriochlorophyll in chlorosomes of the green photosynthetic bacteriumChloroflexus aurantiacus: Studies of fluorescence depolarization accompanied by energy transfer processes

Examination was made of changes in fluorescence polarization plane by energy transfer in the chlorosomes of the green photosynthetic bacterium,Chloroflexus aurantiacus. Fluorescence anisotropy in the picosecond (ps) time region was analyzed using chlorosomes suspended in solution as well as those oriented in a polyacrylamide gel. When the main component of BChlc was preferentially excited, the decay of fluorescence anisotropy was found to depend on wavelength. In the chlorosome suspension, the anisotropy ratio of BChlc changed from 0.31 to 0.24 within 100 ps following excitation. In the baseplate BChla region, this ratio decreased to a negative value (–0.09) from the initial 0.14. In oriented samples, the degree of polarization remained at 0.68 for BChlc, and changed from 0.25 to –0.40 for the baseplate BChla by excitation light whose electric vector was parallel to the longest axis of chlorosomes. In the latter case, there was a shift from 0.30 to –0.55 by excitation perpendicular to the longest axis. Time-resolved fluorescence polarization spectra clearly indicated extensive changes in polarization plane accompanied by energy transfer. The directions of polarization plane of emission from oriented samples were mostly dependent on chlorosome orientation in the gel but not on that of the polarization plane of excitation light. Orientations of the dipole moment of fluorescence components was consistent with that of absorption components as determined by the linear dichroism (Matsuura et al. (1993) Photochem. Photobiol. 57: 92–97). A model for molecular organization of BChlc anda in chlorosomes is proposed based on anisotropic optical properties.     

Tubular exciton models for BChl c antennae in chlorosomes from green photosynthetic bacteria

Exciton calculations on tubular pigment aggregates similar to recently proposed models for BChl c/d/e antennae in light-harvesting chlorosomes from green photosynthetic bacteria yield electronic absorption spectra that are super-impositions of linear J-aggregate spectra. While the electronic spectroscopy of such antennae differs considerably from that of linear J-aggregates, tubular exciton models (which may be viewed as cross-coupled J-aggregates) may be constructed to yield spectra that resemble that of the BChl c antenna in the green bacterium Chloroflexus aurantiacus. Highly symmetric tubular models yield absorption spectra with dipole strength distributions essentially identical to that of a J-aggregate; strong symmetry-breaking is needed to simulate the absorption spectrum of the BChl c antenna.Abbreviations BChl bacteriochlorophyll - [E,M] BChl c _S bacteriochlorophyll c with ethyl and methyl substituents in the 8- and 12-positions, and with stearol as the esterifying alcohol     

Incorporation of exogenous long-chain alcohols into bacteriochlorophyll c homologs by Chloroflexus aurantiacus

Chloroflexus aurantiacus grown in batch culture took up exogenous alcohols and incorporated these into bacteriochlorophyll c as the esterifying alcohol. It was possible to change the distribution of the naturally occurring homologs of bacteriochlorophyll c esterified with phytol, hexadecanol, and octadecanol by adding the appropriate alcohol. The corresponding homolog then made up at least 60% of the cellular bacteriochlorophyll c. It was also possible to obtain novel bacteriochlorophyll homologs not found in detectable amounts in control cells by adding fatty alcohols with short chains (C10, C12) or long chains (C20). These changes in bacteriochlorophyll composition had no detectable effects on the spectral properties of the chlorosomes.Abbreviation BChl Bacteriochlorophyll     

Temperature shift effect on the Chlorobaculum tepidum chlorosomes

Chlorobaculum [Cba.] tepidum is known to grow optimally at 48–52 °C and can also be cultured at ambient temperatures. In this paper, we prepared constant temperature, temperature shift, and temperature shift followed by backshift cultures and investigated the intrinsic properties and spectral features of chlorosomes from those cultures using various approaches, including temperature-dependent measurements on circular dichroism (CD), UV–visible, and dynamic light scattering. Our studies indicate that (1) chlorosomes from constant temperature cultures at 50 and 30 °C exhibited more resistance to heat relative to temperature shift cultures; (2) as temperature increases bacteriochlorophyll c (BChl c) in chlorosomes is prone to demetalation, which forms bacteriopheophytin c, and degradation under aerobic conditions. Some BChl c aggregates inside reduced chlorosomes prepared in low-oxygen environments can reform after heat treatments; (3) temperature shift cultures synthesize and incorporate more BChl c homologs with a smaller substituent at C-8 on the chlorin ring and less BChl c homologs with a larger long-chain alcohol at C-17³ versus constant-temperature cultures. We hypothesize that the long-chain alcohol at C-17³ (and perhaps together with the substituent at C-8) may account for thermal stability of chlorosomes and the substituent at C-8 may assist self-assembling BChls; and (4) while almost identical absorption spectra are detected, chlorosomes from different growth conditions exhibited differences in the rotational length of the CD signal, and aerobic and reduced chlorosomes also display different Q_y CD intensities. Further, chlorosomes exhibited changes of CD features in response to temperature increases. Additionally, we compare temperature-dependent studies for the Cba. tepidum chlorosomes and previous studies for the Chloroflexus aurantiacus chlorosomes. Together, our work provides useful and novel insights on the properties and organization of chlorosomes.     

Bacteriochlorophyllc formation and chlorosome development inChloroflexus aurantiacus

The dependence of chlorosome development on bacteriochlorophyll (BChl)c synthesis was studied with the phototrophic green bacteriumChloroflexus aurantiacus. By selecting defined culture conditions, three possibilities could be identified. Upon addition of 5-aminolevulinic acid, cells of resting cultures increased their specific BChlc contents as well as the volumes of already existing chlorosomes. The number of chlorosomes, however, remained constant. Serine-limited chemostat cultures grown under steady state conditions exhibited constant rates of synthesis of both BChlc as well as of chlorosomes. The volume of the latter remained constant, as well. Upon addition of ALA to chemostat cultures, chlorosomes were synthesized at the same rate as before but their volumes increased as a consequence of increased BChlc incorporation. In chlorosomes isolated from resting cultures supplied with ALA the amounts of all of the polypeptides increased only slightly, if at all. Moreover, the ratio of all of the chlorosomal polypeptides remained largely constant. These results show that chlorosomes may incorporate newly synthesized BChlc without concomitant formation of chlorosomal polypeptides. This means that there was no obvious coordination of polypeptide and BChlc synthesis. On this basis, it appears unlikely that one of the chlorosomal polypeptides functions as an apoprotein of a presumed BChlc holochrome complex.     

Rearrangement of light harvesting bacteriochlorophyll homologues as a response of green sulfur bacteria to low light intensities

The pigment composition of two species of green-colored BChl c-containing green sulfur bacteria (Chlorobium limicola and C. chlorovibrioides) and two species of brown-colored BChl e-containing ones (C. phaeobacteroides and C. phaeovibrioides) incubated at different light intensities have been studied. All species responded to the reduction of light intensity from 50 to 1 Einstein(E) m^–2 s^–1 by an increase in the specific content of light harvesting pigments, bacteriochlorophylls and carotenoids. At critical light intensities (0.5 to 0.1 E m^–2 s^–1) only brown-colored chlorobia were able to grow, though at low specific rates (0.002 days^–1 mg prot^–1). High variations in the relative content of farnesyl-bacteriochlorophyll homologues were found, in particular BChl e ₁ and BChl e ₄, which were tentatively identified as [M, E] and [I, E] BChl_F e, respectively. The former was almost completely lost upon reduction of light intensity from 50 to 0.1 E m^–2 s^–1, whereas the latter increased from 7.2 to 38.4% and from 13.6 to 42.0% in C. phaeobacteroides and C. phaeovibrioides, respectively. This increase in the content of highly alkylated pigment molecules inside the chlorosomes of brown species is interpreted as a physiological mechanism to improve the efficiency of energy transfer towards the reaction center. This study provides some clues for understanding the physiological basis of the adaptation of brown species to extremely low light intensities.Abbreviations BChl bacteriochlorophyll - [M, E] BChl_F e 8-methyl, 12-ethyl BChl e, esterified with farnesol (F). Analogously: I - isobutyl - Pr propyl - Car carotenoids - Chlb chlorobactene - HPLC high performance liquid chromatography - Isr isorenieratene - LHP light harvesting pigments - PDA photodiode array detector - RC reaction center - RCH relative content of homologues     

Excitation energy transfer in the green photosynthetic bacterium Chloroflexus aurantiacus: A specific effect of 1-hexanol on the optical properties of baseplate and energy transfer processes

The effect of 1-hexanol on spectral properties and the processes of energy transfer of the green gliding photosynthetic bacterium Chloroflexus aurantiacus was investigated with reference to the baseplate region. On addition of 1-hexanol to a cell suspension in a concentration of one-fourth saturation, a specific change in the baseplate region was induced: that is, a bleach of the 793-nm component, and an increase in absorption of the 813-nm component. This result was also confirmed by fluorescence spectra of whole cells and isolated chlorosomes. The processes of energy transfer were affected in the overall transfer efficiency but not kinetically, indicating that 1-hexanol suppressed the flux of energy flow from the baseplate to the B806-866 complexes in the cytoplasmic membranes. The fluorescence excitation spectrum suggests a specific site of interaction between bacteriochlorophyll (BChl) c with a maximum at 771 nm in the rod elements and BChl a with a maximum at 793 nm in the baseplate, which is a funnel for a fast transfer of energy to the B806-866 complexes in the membranes. The absorption spectrum of chlorosomes was resolved to components consistently on the basis, including circular dichroism and magnetic circular dichroism spectra; besides two major BChl c forms, bands corresponding to tetramer, dimer, and monomer were also discernible, which are supposed to be intermediary components for a higher order structure. A tentative model for the antenna system of C. aurantiacus is proposed.Abbreviations A670 a component whose absorption maximum is located at 670 nm - (B)Chl (bacterio)chlorophyll - CD circular dichroism - F675 a component whose emission maximum is located at 675 nm - FMO protein Fenna-Mathews-Olson protein - LD linear dichroism - LH light-harvesting - McD magnetic circular dichroism - PS photosystem - RC reaction center     

Manipulation of the bacteriochlorophyll c homolog distribution in the green sulfur bacterium Chlorobium tepidum

We have shown that the green sulfur bacterium Chlorobium tepidum can be grown in batch culture supplemented with potentially toxic fatty alcohols without a major effect on the growth rate if the concentration of the alcohols is kept low either by programmed addition or by adding the alcohol as an inclusion complex with -cyclodextrin. HPLC and GC analysis of pigment extracts from the supplemented cells showed that the fatty alcohols were incorporated into bacteriochlorophyll c as the esterifying alcohol. It was possible to change up to 43% of the naturally occurring farnesyl ester of bacteriochlorophyll c with the added alcohol. This change in the homolog composition had no effect on the spectral properties of the cells when farnesol was partially replaced by stearol, phytol or geranylgeraniol. However, with dodecanol we obtained a blue-shift of 6 nm of the Q_y band of the bacteriochlorophyll c and a concomitant change in the fluorescence emission was observed. The possible significance of these findings is discussed in the light of current ideas about bacteriochlorophyll organization in the chlorosomes.Abbreviations -CD -cyclodextrin - BChl bacteriochlorophyll - BChl c _H bacteriochlorophyllide c - [E,M] BChl c _F 8-ethyl, 12-methyl, farnesyl BChl c - [E,E] BChl c _F 8-ethyl, 12-ethyl, farnesyl BChl c - [P,E] BChl c _F 8-propyl, 12-ethyl, farnesyl BChl c - [I,E] BChl c _F 8-isobutyl, 12-ethyl, farnesyl BChl c - Car carotenoids     

Chlorosomes of green sulfur bacteria: Pigment composition and energy transfer

The pigment composition and energy transfer pathways in isolated chlorosomes ofChlorobium phaeovibrioides andChlorobium vibrioforme were studied by means of high performance liquid chromatography (HPLC) and picosecond absorbance difference spectroscopy. Analysis of pigment extracts of the chlorosomes revealed that they contain small amounts of bacteriochlorophyll (BChl)a esterified with phytol, whereas the BChlsc, d ande are predominantly esterified with farnesol. The chlorosomal BChla content inC. phaeovibrioides andC. vibrioforme was found to be 1.5% and 0.9%, respectively. The time resolved absorbance difference spectra showed a bleaching shifted to longer wavelengths as compared to the Q_y absorption maxima and in chlorosomes ofC. vibrioforme also an absorbance increase at shorter wavelengths was observed. These spectral features were ascribed to excitation of oligomers of BChle and BChlc/d, respectively. One-color and two-color pump-probe kinetics ofC. phaeovibrioides showed rapid energy transfer to long-wavelength absorbing BChle oligomers, followed by trapping of excitations by BChla with a time constant of about 60 ps. Time resolved anisotropy measurements inC. vibrioforme showed randomization of excitations among BChla molecules with a time constant of about 20 ps, indicating that BChla in the baseplate is organized in clusters. One-color and two-color pump-probe measurements inC. vibrioforme showed rapid energy transfer from short-wavelength to long-wavelength absorbing oligomers with a time constant of about 11 ps. Trapping of excitations by BChla in this species could not be resolved unambiguously due to annihilation processes in the BChla clusters, but may occur with time constants of 15, 70 and 200 ps.     

Circular dichroism of green bacterial chlorosomes

Positive and negative bands in previously measured circular dichroism (CD) spectra of Chlorobium limicola chlorosomes appeared to be sign-reversed relative to those of Chloroflexus aurantiacus chlorosomes in the 740–750 nm spectral region where bacteriochlorophyll (BChl) c absorbs maximally. It was not clear, however, whether this difference was intrinsic to the chlorosomes or was due to differences in the procedures used to prepare them. We therefore repeated the CD measurements using chlorosomes isolated from both Cb. limicola f. thiosulfatophilum and Cf. aurantiacus using the method of Gerola and Olson (1986, Biochim. Biophys. Acta 848: 69–76). Contrary to the earlier results, both types of chlorosomes had very similar CD spectra, suggesting that both have similar arrangements of BChl c molecules. The previously reported difference between the CD spectra of Chlorobium and Chloroflexus chlorosomes is due to the instability of Chlorobium chlorosomes, which can undergo a hypsochromic shift in their near infrared absorption maximum accompanied by an apparent inversion in their near infrared CD spectrum during isolation. Treating isolated chlorosomes with the strong ionic detergent sodium dodecylsulfate, which removes BChl a, does not alter the arrangement of BChl c molecules in either Chloroflexus or Chlorobium chlorosomes, as indicated by the lack of an effect on their CD spectra.Abbreviations BChl bacteriochlorophyll - Cb. Chlorobium - CD circular dichroism - Cf. Chloroflexus - NIR near infrared     

Aggregation of 8,12-diethyl farnesyl bacteriochlorophyll c at low temperature

The effect of temperature on the aggregation of 3^lR-8,12-diethyl farnesyl bacteriochlorophyll c in a mixture of n-pentane and methylcyclohexane (1/1, v/v) was studied by means of absorption, circular dichroism and fluorescence spectroscopy. At room temperature essentially only two aggregate species, absorbing at 702 nm (A-702) and 719 nm (A-719), were present. Upon cooling to 219 K, A-702 was quantitatively converted to A-719. Further lowering of the temperature led to the stepwise formation of larger aggregates by the conversion of A-719 to aggregate species absorbing at 743 nm (A-743) and 755 nm (A-755). All absorption changes were reversible. A-719 was highly fluorescent (maximum at 192 K: 744 nm), while A-743 and especially A-755 were weakly fluorescent. Below 130 K the mixture solidified, and no major changes in the absorption spectrum were observed upon further cooling. At 45 K, however, a relatively strong emission at 775 nm was observed. Below 200 K, the absorption, fluorescence and circular dichroism spectra resembled that of the chlorosome. These results open up the possibility to study higher aggregates of BChl c as models for the chlorosome by various methods at low temperature, thus avoiding interference by thermal processes.Abbreviations A-680, A-702, A-719, A-743 and A-755- BChl c aggregates absorbing at the wavelengths indicated - BChl- bacteriochlorophyll - R[E,E] BChl c _F- the 3¹ R isomer of 8,12-diethyl BChl c esterified with farnesol (F), analogously - M- methyl - Pr- propyl - S- stearol (see Smith 1994) - CD- circular dichroism     

Effects of illumination intensity on bacteriochlorophyllc homolog distribution inChloroflexus aurantiacus grown under controlled conditions

Green photosynthetic bacteria contain a mixture of stereoisomers and homologs of their major light harvesting pigment, bacteriochlorophyll (BChl)c. We have determined the distribution of photosynthetic pigments in the green filamentous bacteriumChloroflexus aurantiacus grown in turbidostat culture under light-limited conditions at 5 different illumination intensities. Pigments were extracted from isolated cells, analyzed by HPLC, and the homologs of BChlc identified by their mass spectra. The ratio between BChlc, BChla and carotenoid remained constant at low illumination intensities; at higher intensities BChla and carotenoid increased in parallel compared to BChlc. The BChlc homolog distribution changed even under conditions where the ratio of the total amount to the other pigments was unchanged, but there were no evidence for a constant stoichiometric ratio between any pair of homologs.     

Aggregation of bacteriochlorophyll c homologs to dimers,tetramers, and polymers in water-saturated carbon tetrachloride

Three homologs of BChl c, 2-(R)-(1-hydroxyethyl)-4-n-propyl-5-ethyl-farnesyl BChl c (PEF-BChl c), 2-(R)-(1-hydroxyethyl)-4-ethyl-5-ethyl-farnesyl BChl c (EEF-BChl c), and 2-(S)-(1-hydroxyethyl)-4-isobutyl-5-methyl/ethyl-farnesyl BChl c (iBM/EF-BChl c), formed aggregates in water-saturated carbon tetrachloride (H₂O-satd CCl₄). The water content was about 100 times higher than that of the dried CCl₄ previously used. Absorption spectra were recorded for 8 concentrations for the three homologs of BChl c and were deconvoluted in terms of standard spectra of monomer, dimer, tetramer and polymer (747-nm aggregate, Olson and Pedersen (1990) Photosynthe Res 25: 25). PEF- and EEF-BChl c formed dimers (680 nm maximum) and tetramers (705–710 nm maximum), but iBM/EF-BChl c formed polymers. Inhibition of dimer formation by water faciliated the study of the initial stages of the polymerization of BChl c. When the logarithm of polymer concentration was plotted versus the logarithm of the monomer concentration for iBM/EF-BChl c, the initial slope was 30±10 and indicated the cooperation of 20–40 BChl c molecules to form a polymer from a seed. Circular dichroism spectra of the polymers with positive and negative bands at 743 and 760 nm, respectively, were similar to those for chlorosomes (Brune et al. (1990) Photosynth Res 24: 253).Abbreviations BChl bacteriochlorophyll - CD circular dichroism - EEF 4-ethyl-5-ethyl farnesyl - iBM/EF 4-isobutyl-5-methyl/ethyl farnesyl - H₂O-satd CCl₄ water saturated carbon tetrachloride - PEF 4-n-propyl-5-ethyl farnesyl     

Investigation on chlorosomal antenna geometries: tube,lamella and spiral-type self-aggregates

Molecular mechanics calculations and exciton theory have been used to study pigment organization in chlorosomes of green bacteria. Single and double rod, multiple concentric rod, lamella, and Archimedean spiral macrostructures of bacteriochlorophyll c molecules were created and their spectral properties evaluated. The effects of length, width, diameter, and curvature of the macrostructures as well as orientations of monomeric transition dipole moment vectors on the spectral properties of the aggregates were studied. Calculated absorption, linear dichroism, and polarization dependent fluorescence-excitation spectra of the studied long macrostructures were practically identical, but circular dichroism spectra turned out to be very sensitive to geometry and monomeric transition dipole moment orientations of the aggregates. The simulations for long multiple rod and spiral-type macrostructures, observed in recent high-resolution electron microscopy images (Oostergetel et al., FEBS Lett 581:5435–5439, 2007) gave shapes of circular dichroism spectra observed experimentally for chlorosomes. It was shown that the ratio of total circular dichroism intensity to integrated absorption of the Q _y transition is a good measure of degree of tubular structures in the chlorosomes. Calculations suggest that the broad Q _y line width of chlorosomes of sulfur bacteria could be due to (1) different orientations of the transition moment vectors in multi-walled rod structures or (2) a variety of Bchl-aggregate structures in the chlorosomes. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Bacteriochlorophyll c monomers,dimers, and higher aggregates in dichloromethane,chloroform, and carbon tetrachloride

Bacteriochlorophyll c in vivo is a mixture of at least 5 homologs, all of which form aggregates in CH₂Cl₂, CHCl₃ and CCl₄. Three homologs exist mainly in the 2-R-(1-hydroxyethyl) configuration, whereas the other two homologs, 4-isobutyl-5-ethyl and 4-isobutyl-5-methyl farnesyl bacteriochlorophyll c, exist mainly in the 2-S-(1-hydroxyethyl) configuration (Smith KM, Craig GW, Kehres LA and Pfennig N (1983) J. Chromatograph. 281: 209–223). In CCl₄ the S-homologs form an aggregate of 2–3 molecules whose absorption (747 nm maximum) and circular dichroism spectra resemble those of the chlorosome. In CH₂Cl₂, CHCl₃ and CCl₄ the 4-n-propyl homolog (R-configuration) forms dimers absorbing at ca. 680 nm and higher aggregates absorbing at 705–710 nm. In CCl₄ the dimerization constant is approx. 10 µM^–1 (1000 times that for chlorophyll a). The difference between the types of aggregates formed by the 4-n-propyl and 4-isobutyl homologs is attributed to the difference between the R- and S-configurations of the 2-(1-hydroxyethyl) groups in each chlorophyll.Abbreviations BChl bacteriochlorophyll - CD circular dichroism - Chl chlorophyll - DNS data not shown - EEF 4-ethyl-5-ethyl farnesyl - iBM/EF 4-isobutyl-5-methyl/ethyl farnesyl - MEF 4-methyl-5-ethyl farnesyl - PEP 4-n-propyl-5-ethyl farnesyl