B800-850 Complex Research Articles

Inter- and Intraband Energy Transfer in LH2-Antenna Complexes of Purple Bacteria. A Fluorescence Line-Narrowing and Hole-Burning Study

High-resolution site-selection fluorescence- and hole-burning spectroscopy were used to study energy transfer in two LH2 light-harvesting complexes of purple bacteria: the B800-850 complex of isolated Rb. sphaeroides and the B800-820 complex of Rps. acidophila, at 1.2 K. Fluorescence spectra, hole widths, and hole depths were measured as a function of excitation wavelength [lambda][sub exc] within the B800 band. For [lambda][sub exc] [>=] 798 nm, fluorescence line-narrowing is observed and the energy-transfer times ([tau] = 2.5 and 2.0 ps for B800-850 and B800-820, respectively) are independent of [lambda][sub exc]. In this spectral region only interband B800 [yields] B850 (B820) energy transfer takes place. For 780 nm [<=] [lambda][sub exc] [<=] 798 nm, the fluorescence bands are broad and the transfer time, obtained from hole widths extrapolated to zero burning-fluorence density, decreases toward the blue side of B800. In this wavelength region competition occurs between B800 [yields] B850 (B820) and B800 [yields] B800 [open quotes]downhill[close quotes] energy transfer. 36 refs., 5 figs.

The Q gene of Rhodobacter sphaeroides: its role in puf operon expression and spectral complex assembly

The Q gene of the facultative photoheterotroph Rhodobacter sphaeroides, localized immediately upstream of the oxygen- and light-regulated puf operon, encodes a 77-amino-acid polypeptide. The 5' and 3' ends of the 561-bp Q transcript were determined. To gain insight into the role of the Q gene product, a number of Q mutations were constructed by oligonucleotide-directed mutagenesis and subsequent substitution of the mutated form of the gene in single copy for the chromosomal copy via homologous recombination. The resulting mutants can grow photosynthetically, with the exception of QSTART, in which the initiation codon for the Q protein was altered. Spectral analysis of the intracytoplasmic membranes showed that one of the missense mutants (QdA) was deficient in the formation of detectable B875 light-harvesting complex (LHC), whereas deletion of the stem-loop structure (Qloop) failed to form B800-850 LHC when grown anaerobically either in the dark or under light intensity of 100 W/m2. Other missense mutants (QuA and QuB) contained either more B800-850 LHC or more B875 LHC, respectively, than the wild type. Although the levels of puf and puc transcripts isolated from QSTART grown anaerobically on succinate-dimethyl sulfoxide in the dark were comparable to wild-type levels, no B875 spectral complex was detected and there was a greater than 90% reduction in the level of the B800-850 pigment-protein complex. It has also been confirmed that the ultimate cellular levels of either the B875 or B800-850 spectral complexes can vary over wide limits without any change in the level(s) of complex specific transcripts. When the wild-type Q gene was reintroduced in trans into the Q mutations, QSTART was able to grow photosynthetically and both B800-850 and B875 spectral complexes were formed in either QdA or Qloop. Finally, we demonstrated that the level of each puf-specific mRNA behaves independently of one another as well as independently of the level(s) of Q gene-specific mRNA. These results are compatible with the existence of regulatory sequences affecting the puf mRNA level(s) being localized within the Q structural gene. These results suggest that Q-specific expression is uncoupled from puf-specific transcription and that the Q protein is not involved in the regulation of transcription of the puf operon but is directly involved in the assembly of both the B875 and B800-850 pigment-protein complexes.

Presence and significance of minor antenna components in the energy transfer sequence of the green photosynthetic bacterium Chloroflexus aurantiacus

Antenna components in the energy transfer processes of a green photosynthetic bacterium Chloroflexus aurantiacus were spectrally investigated by time-resolved fluorescence spectroscopy at −196°C on intact cells. Besides major antenna components so far reported, three minor components were resolved; those were Bchl c located at 785 nm, the baseplate Bchl a at 819 nm and Bchl a in the B808-866 complex at 910 nm. The last component was assigned to a longer wavelength antenna closely associated with a reaction center. An additional Bchl c fluorescence component was kinetically suggested to be present, which can be an energy donor to a major Bchl c. Presence of these minor components was signified in terms of (1) increase in the spectral overlap integral and (2) adjustment of the direction of dipole moments in the energy transfer sequence of intact cells.

Characterization of a D1-D2-cyt b-559 complex containing 4 chlorophyll a/2 pheophytin a isolated with the use of MgSO4

A D1-D2-cyt b-559 complex containing 4 chlorophyll a, 1 β-carotene and 1 cytochrome b-559 per 2 pheophytin a has been isolated from spinach with 30% yield using a Q-Sepharose Fast-Flow anion-exchange column equilibrated with 0.1% Triton X-100, 10 mM MgSO4 and 50 mM Tris-HCl (pH 7.2). The preparation was then stabilized with 0.1% dodecyl-β-d-maltoside. This method gave a yield 10 times higher than that using a Fractogel TSK-DEAE 650(S) column equilibrated with 0.1% Triton X-100, 30 mM NaCl and 50 mM Tris-HCl (pH 7.2). The PS II RC complex was characterized using absorption and fluorescence spectroscopy at 277 and 77 K. A selective reversible bleaching under reducing conditions with maximum at 682 nm, associated with pheophytin a reduction, and light-induced absorption differences with a lifetime of 1.0 ms, ascribed to the triplet state of P680 were measured and indicated that the isolated Dl-D2-cyt b-559 complex is active in charge separation. The results are compared with the data obtained for a PS II RC preparation containing 6 chlorophyll a, 2 β-carotene and 1 cyt b-559 per 2 pheophytin a.

Spectral and thermodynamic properties of the two hemes of the D1D2cytochrome b-559 complex of spinach

In agreement with previous work [Shuvalov, Heber and Schreiber (1988) FEBS Lett. 258, 27-31] two hemes (low potential (LP) and extra low potential (XLP )) per two pheophytins were found in isolated D1D2Cyt b-559 complexes. Reductive and oxidative redox titrations demonstrate that the E m of the LP form is at about +150 mV. It is independent of pH between pH 7.2 and 9.4. The XLP heme is autoxidizable at pH 7.2 and displays, at this pH, an E m of − 45 mV. Both the LP and XLP hemes show absorption peaks at 559 nm. They are proposed to have bis-histidine ligation of the heme iron. At pH 9.4, the XLP heme splits into two forms. One of them has an E m of +40 mV, and absorption peaks at 559 nm showing the bis-histidine ligation. The other displays an E m of −220 mV and the peak is shifted to 562 nm. This last form is proposed to be due to the incorporation of OH − which occupies the 6th coordination position of the heme Fe(III) at high pH. The p K value for the conversion of the XLP heme is close to 7.7. In a structure simulation of the α-helices of α- and β-polypeptides, the β-polypeptide, but not the α-polypeptide, reveals a distance between the histidine N and the heme Fe which permits stable N-Fe coordination. In the α-polypeptide, OH − can be incorporated between N and Fe. The functional role of the two hemes of cyt b-559 is briefly discussed with respect to water oxidation and cyclic electron transfer.

The effect of growth conditions on the light-harvesting apparatus in Rhodopseudomonas acidophila

The detailed effect on the light-harvesting apparatus of three different wild-type strains of Rhodopseudomonas acidophila in response to changes in both light-intensity and temperature have been investigated. In all three strains at high light-intensities (160 μmol s m(2) and above) the only LH2 antenna complex synthesised is the B800-850 complex. In strains 7050 and 7750 as the light-intensity is lowered the B800-850 complex is gradually replaced by another type of LH2 the B800-820 complex. However, at no light-intensities studied is this changeover complete when the cells are grown at 30°C. If however, the light-intensity is lowered at temperatures below 25°C with strain 7750 there is a complete replacement of the B800-850 complex by the B800-820 complex. At all light-intensities and temperatures tested, strain 10050 only synthesised the B800-850 complex. Strain 7050 also responded to changes in light-intensity by altering its carotenoid composition. At high light-intensity the major carotenoids were rhodopin and rhodopin-glucoside, while at low light-intensities the major ones were rhodopinal and rhodopinal-glucoside. This change in carotenoid content started to occur at rather higher light-intensities than the switchover from B800-850 to B800-820.

ALTERED SPECTRAL PROPERTIES OF THE B875 LIGHT‐HARVESTING PIGMENT‐PROTEIN COMPLEX IN A Rhodobacter sphaeroides MUTANT LACKING pufX

Abstract— Mutants of Rhodobacler sphac, roicies lacking the pufX gene are unable to grow photosynthetically and have been reported to show impaired cyclic electron flow and elevated B875 levels when grown at low oxygen tension. An examination of the low‐temperature optical properties of chromatophores from a pufx‐ strain, obtained by complementation of a pufLMX mutant in trans with pufQBALM, encoding polypeptides of the B875 antenna and reaction center complexes, revealed that the absorption and fluorescence emission maxima and the fluorescence polarization rise over the B875‐Qy band were shifted to lower energies. These results suggest that the pufX gene product may limit the aggregation state of the B875 complex, assuring the proper functional arrangement of antenna and reaction ccntcr components within the cores of the photosynthetic units.

Picosecond dynamics of excitations in light-harvesting complex B800-850 from Chromatium minutissimum studied using fluorescence spectrochronography

The picosecond dynamics of excitations in the isolated B800-850 light-harvesting complex of the purple sulfur bacterium Chromatium minutissimum has been studied using picosecond fluorescence spectrochronography. A short-lived component of about 20 ps lifetime has been found at 77K at the short wavelength part of the B850 fluorescence spectrum similar to that previously described for the core antenna bacteriochlorophyll band B880 of Rhodospirillum rubrum. Evidence has been presented indicating that this component is likely to reflect excitation energy relaxation step(s) involving both photoexcited bacteriochlorophyll and the protein environment. A new kinetic scheme of excitation transfer from the peripheral antenna to the photoreceptor units in purple bacteria is suggested which takes into account these findings.

FT-IR studies on the triplet state of P680 in the photosystem II reaction center: triplet equilibrium within a chlorophyll dimer.

The structure and molecular interactions of the primary donor (P680) in the reaction center (D1-D2-cytochrome b-559 complex) of photosystem II (PS II) have been investigated by detecting light-induced FT-IR difference spectra upon the formation of its triplet state (3P680). The 3P680/P680 spectrum obtained was analyzed by comparing it with difference spectra between the ground and lowest triplet states of purified chlorophyll a (Chl) in organic solvents. The negative peaks at 1669 and 1707 cm-1 accompanied by the positive peaks at 1627 and 1659 cm-1 in the 3P680/P680 spectrum were assigned to the keto C = O stretching mode, and the appearance of these two pairs of bands indicated that P680 has a dimeric structure analogous to that of the bacterial primary donor. From the band positions of the keto and carbomethoxy C = O stretches, the hydrogen-bonding properties of these two Chl molecules were found to be asymmetrical; in one Chl molecule both the keto and carbomethoxy C = O groups form hydrogen bonds, while in the other Chl molecule the keto C = O is not hydrogen-bonded whereas the carbomethoxy C = O probably is hydrogen-bonded. The temperature dependence of the intensity ratios of the keto C = O bands revealed that the triplet state is equilibrated between the two Chl molecules with an energy gap of 8.4 +/- 0.7 meV. Most of the triplet population was found to be localized on one Chl molecule (86% at 80 K), in which both of the two C = O groups are hydrogen-bonded, that is probably attached to the D1 subunit. Considering the structure of the bacterial reaction center determined by X-ray crystallography and the sequence homology between the D1 and D2 subunits of PS II and the L and M subunits of bacteria, a model of the P680 structure and its interactions with apoproteins has been proposed.

Unexpected similarities of the B800-850 light-harvesting complex from Rhodospirillum molischianum to the B870 light-harvesting complexes from other purple photosynthetic bacteria

The B800-850 light-harvesting complex (also called LH2) was isolated from photosynthetic membranes of Rhodospirillum molischianum DSM 119 using molecular sieve and ion-exchange chromatography. Its two bacteriochlorophyll a-binding polypeptides (alpha-subunit and beta-subunit) were purified with a reverse-phase HPLC system. The complete amino acid sequences of both subunits have been determined. The alpha- and beta-subunits consist of 56 and 45 amino acids, respectively, corresponding to molecular weights of 5939 and 5133. In contrast to the B800-850 complexes from other photosynthetic bacteria, the native B800-850 complex from Rs. molischianum is most likely an octamer of monomers with a stoichiometry of three bacteriochlorophyll a and 1.5 lycopenes per alpha,beta-subunit. Resonance Raman spectra provide evidence for a 5-coordinated Mg2+ in the BChl, and a carotenoid mainly in the all-trans configuration. A comparison between resonance Raman data from different photosynthetic bacteria indicates that the BChl a-binding site of the B800-850 complex from Rs. molischianum is more similar to the B870 complexes (also called LH1) than to the B800-850 complexes of other photosynthetic bacteria. Sequence similarities especially between the beta-subunits of the B800-850 complex of Rs. molischianum and the B870 and B800-850 complexes of other photosynthetic bacteria agree with this result and provide information on the mode of pigment binding in bacterial antenna complexes.

5-Aminolevulinic acid availability and control of spectral complex formation in hemA and hemT mutants of Rhodobacter sphaeroides.

In the photosynthetic bacterium Rhodobacter sphaeroides, two genes, hemA and hemT, each encode a distinct 5-aminolevulinic acid (ALA) synthase isozyme (E. L. Neidle and S. Kaplan, J. Bacteriol. 175:2292-2303, 1993). This enzyme catalyzes the first and rate-limiting step in a branched pathway for tetrapyrrole formation, leading to the biosynthesis of hemes, bacteriochlorophylls, and corrinoids. In an attempt to determine the functions of hemA and hemT, mutant strains were constructed with specific chromosomal disruptions. These chromosomal disruption allowed hemA and hemT to be precisely localized on the larger and smaller of two R. sphaeroides chromosomes, respectively. Mutants carrying a single hemA or hemT disruption grew well without the addition of ALA, whereas a mutant, HemAT1, in which hemA and hemT had both been inactivated required exogenous ALA for growth. The growth rates, ALA synthase enzyme levels, and the amounts of bacteriochlorophyll-containing intracytoplasmic membrane spectral complexes of all strains were compared. Under photosynthetic growth conditions, the levels of bacteriochlorophyll, carotenoids, and B800-850 and B875 light-harvesting complexes were significantly lower in the Hem mutants than in the wild type. In the mutant strains, available bacteriochlorophyll appeared to be preferentially targeted to the B875 light-harvesting complex relative to the B800-850 complex. In strain HemAT1, the amount of B800-850 complex varied with the concentration of ALA added to the growth medium, and under conditions of ALA limitation, no B800-850 complexes could be detected. In the Hem mutants, there were aberrant transcript levels corresponding to the puc and puf operons encoding structural polypeptides of the B800-850 and B875 complexes. These results suggest that hemA and hemT expression is coupled to the genetic control of the R. sphaeroides photosynthetic apparatus.

Structural characterization of the B800–850 and B875 light-harvesting antenna complexes from Rhodobacter sphaeroides by electron microscopy

The structure and aggregation behavior of B800–850 (LHII) and B875 (LHI) antenna complexes of Rhodobacter sphaeroides were studied by electron microscopy. Single molecular projections (top views and side views) of isolated particles were analyzed. The B800–850 complexes, isolated as 150 kDa particles, are cylindrical with a diameter of 8.5 nm and with a height of 6.5 nm. If corrected for attached detergent, the actual diameter in the plane of the membrane would be about 5.1 nm. Stain accumulation in the center of the structure indicates a small indentation, therefore a ring-shaped structure is expected. The size, shape and estimated mass give evidence for a B800–850 complex structure consisting of 4–6 αβ heterodimers, if in an α n β n ring-shaped configuration. The B875 complexes, isolated as 360 kDa particles, are dimeric units, but the two monomers are only loosely connected. A single B875 unit has a corrected diameter of about 5.2 nm and a height of 6.2 nm, similar to the B800–850 unit. Therefore, the B875 complex most likely has the same α n β n configuration. Image analysis of B800–850 projections points to a 3-or 6-fold symmetry in the top views, rather than a 4- or 5-fold symmetry. Therefore, our results are most compatible with an α 6 β 6 configuration.

Chemical crosslinking studies of the isolated light-harvesting B800-850 complex of Chromatium minutissimum

The spatial relationship of polypeptides comprising the light-harvesting B800-850 complex of Chromatium minutissimum has been studied by means of chemical crosslinking of the isolated complex with cleavable, 1.2 nm-long dithiobis(succinimidyl propionate). The samples were analyzed by different types of electrophoresis and spectrophotometrically. No difference was shown between crosslinking of the B800-850 complex either solubilized or incorporated into proteoliposomes. It was found that two main polypeptides form only one type of heterodimer. The crosslinked complex was more thermostable. This crosslinkage restricted the conformational transitions causing the shift of the long wavelength band in the near infrared region. A structure of the complex is discussed.

Precise location of the Cu(II)-inhibitory binding site in higher plant and bacterial photosynthetic reaction centers as probed by light-induced absorption changes.

Light-dependent absorption change at 325 nm, ascribed to QA activity, was strongly reduced in the presence of Cu(II) in oxygen-evolving core complex. This change was much less affected in the presence of the herbicide 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), indicating that the Cu(II)-binding site is different from that of the DCMU and that Cu(II) blocks QA reduction. Cu(II) did not eliminate the absorption change at 545 nm, ascribed to pheophytin reduction, in Na2S2O4-treated oxygen-evolving core and D1-D2-cytochrome b559 complexes. This indicates that Cu(II) does not affect the electron transport between P680 and pheophytin. Moreover, the activity of the bacterial reaction center probed by the absorption change at 790 nm was inhibited by Cu(II), but the signal at 530 nm, associated to the reduction of bacteriopheophytin in Na2S2O4-treated reaction center, was not inhibited. We conclude that Cu(II) impaired the photosynthetic electron transport between pheophytin and QA in both higher plants and photosynthetic bacteria. Cu(II) would bind to an amino acid(s) highly conserved in non-oxygenic and oxygenic reaction centers, which is(are) necessary for the electron transfer between pheophytin and QA. Based on the atomic structure of the bacterial reaction center several schemes of possible Cu(II) binding are shown.

Investigation of the neighbour relationships between photosystem II polypeptides in the two types of isolated reaction centres (D1/D2/cyt b559 and CP47/D1/D2/cyt b559 complexes)

The nearest neighbour relationships within the D1/D2/cyt b 559 complex (PSIIRC) and the CP47/D1/D2/cyt b 559 complex (RC-CP47) were investgated by using different length bifunctional crosslinking agents. The crosslinking products were identified by immunoblotting with polyclonal antibodies and by two-dimensional gel electrophoresis. Seven products (CP47/D2, D1/D2/α, D1/D2, D2/α, D1/α, α/αa, α/β) have been revealed in both complexes. The crosslinking of both complexes does not increase their photostability. The photocrosslinking products (D1/α and D2/α) appeared under illumination of complexes with light of high intensity.

Analysis of the optical absorbance spectra of D1-D2-cytochrome b-559 complexes by absorbance-detected magnetic resonance : Structural properties of P680

Absorbance-detected magnetic resonance (ADMR) spectroscopy was applied to D1-D2-cyt b-559 complexes isolated by different isolation methods. The triplet of the primary donor ( 3P680) does not show a homogeneous response to the microwaves; the maximum of the triplet-induced long-wavelength bleaching varied from 678 to 685.5 nm, whereas the | D|-value changed from 0.0293 to 0.0281 cm −1 with increasing monitoring wavelength within the bleaching band. The | E|-value was 0.0043 cm −1 for all detection wavelengths. Triplet-minus-singlet (T — S) difference spectra monitored with ADMR did not show any indication for a dimeric structure of P680. Linear dichroic (LD) ADMR applied to P680 yielded angles between the optical Q y transition moments and the triplet x-, y- and z- axes of 49° ± 2°, 32° ± 4° and 65° ± 3°, respectively. From LD-ADMR applied to chlorophyll a (Chl a) in vitro corresponding angles of 55° ± 6° and 43° ± 7° for the triplet x- and y-axes, respectively, were obtained. With the value of 30° ± 3° for the angle between the plane spanned by the triplet x- and y-axes and the membrane plane obtained by Van Mieghem et al. (Van Mieghem, F.J.E., Satoh, K. and Rutherford, A.W. (1991) Biochim. Biophys. Acta 1058, 379–385), it follows that the Q y optical-transition moment of P680 makes an angle with the membrane plane of 36° ± 8°. This angle is quite different from the angles calculated from the crystal structure for the dimeric or monomeric bacteriochlorophyll Q y transition moments of bacterial reaction centers (RCs). From the close correspondence between the relative orientation of the triplet axes and the Q y transition moment for both 3P680 and 3Chl in vitro as determined with LD-ADMR and the shape of the T — S spectra, which reflect a monomeric Chl a triplet state, it is concluded that P680 has little, or no interaction with other pigments from the D1-D2-cyt b-559 complex. Our observations contrast strikingly with our previous ADMR experiments on RCs of purple bacteria (Lous E.J. and Hoff, A.J. (1988) Proc. Natl. Acad. Sci. USA 84, 6147–6151) and strongly suggest that the analogy between the D1-D2-cyt b-559 complex and the RC of purple bacteria observed for the biochemical components of the complex does not extend to the conformation of the pigments. An additional triplet ascribed to a pheophytin (Pheo) was observed, having | D| = 0.0338 cm − and | E| = 0.0021 cm −1. The bleaching due to the Pheo-triplet is maximal at 680.0 ± 0.2 nm; at least one Pheo a present in the complex has its absorption maximum at this wavelength. Double-resonance experiments showed that the formation of 3Pheo depends on the population of 3P680. Simulation of the T — S spectra of 3P680 with Gaussian bands revealed three types of P680, two of which could be due to freezing in slightly different variations of a basically similar conformation. One type differs considerably from the other two with respect to the width of the bleaching, the shape of the T — S spectrum and its influence on Pheo a-triplet formation and is likely to have a conformation different from the other two. The T — S spectrum of degraded samples, composed of a mixture of intact and degraded RCs characterised by an absorption maximum below 674 nm, does not differ from that of fresh samples, which, at room temperature, have an absorption maximum at 676 nm. Together with the observation that the triplet yield of degraded samples is much lower, this indicates that for degraded RCs the triplet formation is completely inhibited.

Distribution of energy absorbed by purple bacteria between various paths of de-excitation

Polarized absorption, photoacoustic and fluorescence spectra of cells of the purple bacterium Rhodospirillum rubrum, embedded in isotropic and stretched polyvinyl alcohol films, were measured at 296 and 85 K. The existence of pools of bacteriochlorophyll molecules, characterized by various absorption and emission maxima and different orientations of their transition moments vdth respect to the membrane plane, was shown. The polarized spectra suggest that in addition to the light-harvesting LH1 B875 complexes, other antenna complexes (B806 and complexes absorbing in the long-wavelength region) are present in Rhodospirillum rubrum. The ratio between the yield of fluorescence polarized parallel and perpendicular to the direction of the electric vector of the exciting light (376 nm) and the film axis is about 1.8 for bacteriochlorophyll emission at 892 nm and about 0.14 for that at 928 nm. In the parallel component, 28% of the energy absorbed in the near-IR bands of bacteriochlorophyll is converted into heat and 72% is used for fluorescence emission and photochemistry, whereas in the perpendicular component, the corresponding values are about 60% and 40% respectively. We conclude that the mutually parallel bacteriochlorophyll antenna chromophores lose less energy by thermal dissipation than chromophores with a lower degree of orientation. This suggests a very efficient trapping in the reaction centres of that part of the excitation energy which migrates between chromophores having mutually parallel transition moments.

Investigation of spatial relationships and energy transfer between complexes B800-850 and B890-RC from Chromatium minutissimum reconstituted into liposomes

Spatial relationships between difterent pigment-protein complexes in the membranes of the purple photosynthetic bacterium, Chromatium minutissimum, have been studied. The possibility of restoring the function of efficient excitation energy transfer from bacteriochlorephyll molecules to the reaction centers in the system of soybean liposomes, reconstituted with pigment-protein complexes B800-850 and B890-RC from C. minutissimum, has been explored. The chemical cross-linking method, together with stationary and picosecond spectrally resolved fluoresence measurements were employed. It has been shown that after the incorporation of the complexes into the liposame membranes conditions for directed excitation energy transfer from the light-harvesting pigments to the reaction centers are created, which are less optimal, however, than those in the native state. Possible reasons are considered.

The reaction-centre associated cytochrome subunit of the purple bacterium Rhodocyclus gelatinosus

Isolated chromatophore membranes and purified reaction centre light-harvesting B875 complex from the purple bacterium Rhodocyclus gelatinosus have been studied by EPR spectroscopy. Four haems occurring in approximately equal stoichiometries could be distinguished in chromatophores. The position of their g z peaks, their orientations with respect to the membrane and their electrochemical properties have been characterized ( g z = 3.40, E m = 320 ± 30mV, 0°: g z = 3.40, E m = 70 ± 20mV, 0: g z = 3.30, E m = 130 ± 20mV, 90°; g z = 3.15, E m = 300 ± 20mV, 90°). At ambient potentials where only the high potential haems are reduced (> + 200mV), the stable photooxidation of the + 300 mV haem occurred in a fraction (40%) of centres. Chemical prereduction of the + 130 mV haem allows stable charge separation at 4 K in all of the centres. In the case where all four haems are reduced prior to illumination, a subsequent stable photooxidation (at 4 K) of another haem seems to occur, resulting in trapping of the reduced states of both the primary quinone acceptor Q A and bacteriopheophytin.

The reaction-centre associated cytochrome subunit of the purple bacterium Rhodocyclus gelatinosus

Isolated chromatophore membranes and purified reaction centre light-harvesting B875 complex from the purple bacterium Rhodocyclus gelatinosus have been studied by EPR spectroscopy. Four haems occurring in approximately equal stoichiometries could be distinguished in chromatophores. The position of their g z peaks, their orientations with respect to the membrane and their electrochemical properties have been characterized ( g z = 3.40, E m = 320 ± 30mV, 0°: g z = 3.40, E m = 70 ± 20mV, 0: g z = 3.30, E m = 130 ± 20mV, 90°; g z = 3.15, E m = 300 ± 20mV, 90°). At ambient potentials where only the high potential haems are reduced (> + 200mV), the stable photooxidation of the + 300 mV haem occurred in a fraction (40%) of centres. Chemical prereduction of the + 130 mV haem allows stable charge separation at 4 K in all of the centres. In the case where all four haems are reduced prior to illumination, a subsequent stable photooxidation (at 4 K) of another haem seems to occur, resulting in trapping of the reduced states of both the primary quinone acceptor Q A and bacteriopheophytin.