Excitation energy transfer pathways in Lhca4

EET in reconstituted Lhca4, a peripheral light-harvesting complex from Photosystem I of Arabidopsis thaliana, containing 10 chlorophylls and 2 carotenoids, was studied at room temperature by femtosecond transient absorption spectroscopy. Two spectral forms of Lut were observed in the sites L1 and L2, characterized by significantly different interactions with nearby chlorophyll a molecules. A favorable interpretation of these differences is that the efficiency of EET to Chls is about two times lower from the "blue" Lut in the site L1 than from the "red" Lut in the site L2 due to fast IC in the former case. A major part of the energy absorbed by the "red" Lut, approximately 60%-70%, is transferred to Chls on a sub-100-fs timescale from the state S(2) but, in addition, minor EET from the hot S(1) state within 400-500 fs is also observed. EET from the S(1) state to chlorophylls occurs also within 2-3 ps and is ascribed to Vio and/or "blue" Lut. EET from Chl b to Chl a is biphasic and characterized by time constants of approximately 300 fs and 3.0 ps. These rates are ascribed to EET from Chl b spectral forms absorbing at approximately 644 nm and approximately 650 nm, respectively. About 25% of the excited Chls a decays very fast-within approximately 15 ps. This decay is proposed to be related to the presence of the interacting Chls A5 and B5 located next to the carotenoid in the site L2 and may imply some photoprotective role for Lhca4 in the photosystem I super-complex.     

The Origin of the Low-Energy Form of Photosystem I Light-Harvesting Complex Lhca4: Mixing of the Lowest Exciton with a Charge-Transfer State

The peripheral light-harvesting complex of photosystem I contains red chlorophylls (Chls) that, unlike the typical antenna Chls, absorb at lower energy than the primary electron donor P700. It has been shown that the red-most absorption band arises from two excitonically coupled Chls, although this interaction alone cannot explain the extreme red-shifted emission (25 nm, ∼480 cm⁻¹ for Lhca4 at 4 K) that the red Chls present. Here, we report the electric field-induced absorption changes (Stark effect) on the Q_y region of the Lhca4 complex. Two spectral forms, centered around 690 nm and 710 nm, were necessary to describe the absorption and Stark spectra. The analysis of the lowest energy transition yields a high value for the change in dipole moment, Δμ_710nm ≈ 8 Df⁻¹, between the ground and excited states as compared with monomeric, Δμ = 1 D, or dimeric, Δμ = 5 D, Chl a in solution. The high value of the Δμ demonstrates that the origin of the red-shifted emission is the mixing of the lowest exciton state with a charge-transfer state of the dimer. This energetic configuration, an excited state with charge-transfer character, is very favorable for the trapping and dissipation of excitations and could be involved in the photoprotective mechanism(s) of the photosystem I complex.     

Assignment of spectral substructures to pigment-binding sites in higher plant light-harvesting complex LHC-II

The trimeric main light-harvesting complex (LHC-II) is the only antenna complex of higher plants of which a high-resolution 3D structure has been obtained (Kühlbrandt, W., Wang, D., and Fujiyoshi, Y. (1994) Nature 367, 614-621) and which can be refolded in vitro from its components. Four different recombinant forms of LHC-II, each with a specific chlorophyll (Chl) binding site removed by site-directed mutagenesis, were refolded from heterologously overexpressed apoprotein, purified pigments, and lipid. Absorption spectra of mutant LHC-II were measured in the temperature range from 4 to 300 K and compared to likewise refolded wild-type complex and to native LHC-II isolated from pea chloroplasts. Chls at different binding sites have characteristic, well-defined absorption sub-bands. Mixed occupation of binding sites with Chls a and b is not observed. Temperature-dependent changes of the mutant absorption spectra reveal a consistent shift of the major difference bands but an irregular behavior of minor bands. A model of the spectral substructure of LHC-II is proposed which accounts for the different absorption properties of the 12 individual Chls in the complex, thus establishing a first consistent correlation between the 3D structure of LHC-II and its spectral properties. The spectral substructure is valid for recombinant and native LHC-II, indicating that both have the same spatial arrangement of Chls and that the refolded complex is fully functional.     

The origin of the long-wavelength fluorescence emission band (77 degrees K) from photosystem I

Isolated photosystem I (PSI)-110 particles, prepared using a minimal concentration of Triton X-100 [J. E. Mullet, J. J. Burke, and C. J. Arntzen (1980) Plant Physiol. 65, 814-822] and further subjected to short-term solubilization with sodium dodecyl sulfate (SDS), were resolved into four pigment-containing bands on polyacrylamide gel electrophoresis (PAGE). We have identified these in order of increasing electrophoretic mobility as being (a) CPIa, (b) CPI, (c) the light-harvesting complex of photosystem I (LHC-I), and (d) a free pigment-zone. LHC-I had an absorption maximum in the red at 668-669 nm and a shoulder at 650 nm, which was resolved by its first-derivative spectrum to indicate the presence of chlorophyll b. LHC-I exhibited a 77 degrees K fluorescence emission maximum at 729-730 nm. The 77 degrees K fluorescence emission maxima of CPIa and CPI, excised from the gel, were at 729 and 722 nm, respectively. The LHC-I band, excised from the gel and rerun on dissociating SDS-PAGE, was resolved into two polypeptide doublets of 24-22.5 and 21-20.5 kDa. The CPIa band under similar conditions was resolved into polypeptides of 68, 24, 22.5, 21, 20.5, 19, 15, and 14 kDa; on the contrary, CPI contained only the 68-kDa polypeptide. When intact thylakoids were subjected to "nondenaturing" SDS-PAGE, LHC-I comigrated with an oligomeric form (dimer) of the light-harvesting chlorophyll a/b pigment-protein that preferentially serves photosystem II (LHCP-II). When this combined LHC-I/LHCP-II pigment-protein band was prepared by SDS-PAGE from isolated stroma lamellae, it exhibited a long-wavelength fluorescence band near 730 nm at 77 degrees K. When a similar preparation was obtained from sucrose density gradients containing SDS [J. Argyroudi-Akoyunoglou and H. Thomou (1981) FEBS Lett. 135, 171-181], it was found to be enriched in a 21-kDa polypeptide. The data suggest that the 21-kDa polypeptide of LHC-I is the chlorophyll-containing polypeptide responsible for the long-wavelength fluorescence of LHC-I; other polypeptides in the complex (20.5, 22.5, and 24 kDa) presumably bind chlorophyll and also serve an antennae function.     

Electronic spectra of PS I mutants: the peripheral subunits do not bind red chlorophylls in Synechocystis sp. PCC 6803

Steady-state fluorescence and absorption spectra have been obtained in the Qy spectral region (690-780 nm and 600-750 nm, respectively) for several subunit-deficient photosystem I mutants from the cyanobacterium Synechocystis sp. PCC 6803. The 77 K fluorescence spectra of the wild-type and subunit-deficient mutant photosystem I particles are all very similar, peaking at approximately 720 nm with essentially the same excitation spectrum. Because emission from far-red chlorophylls absorbing near 708 nm dominates low-temperature fluorescence in Synechocystis sp., these pigments are not coordinated to any the subunits PsaF, Psa I, PsaJ, PsaK, PsaL, or psaM. The room temperature (wild-type-mutant) absorption difference spectra for trimeric mutants lacking the PsaF/J, PsaK, and PsaM subunits suggest that these mutants are deficient in core antenna chlorophylls (Chls) absorbing near 685, 670, 675, and 700 nm, respectively. The absorption difference spectrum for the PsaF/J/I/L-deficient photosystem I complexes at 5 K reveals considerably more structure than the room-temperature spectrum. The integrated absorbance difference spectra (when normalized to the total PS I Qy spectral area) are comparable to the fractions of Chls bound by the respective (groups of) subunits, according to the 4-A density map of PS I from Synechococcus elongatus. The spectrum of the monomeric PsaL-deficient mutant suggests that this subunit may bind pigments absorbing near 700 nm.     

Triplet exciton formation as a novel photoprotection mechanism in chlorosomes of Chlorobium tepidum

Chlorosomes comprise thousands of bacteriochlorophylls (BChl c, d, or e) in a closely packed structure surrounded by a lipid-protein envelope and additionally contain considerable amounts of carotenoids, quinones, and BChl a. It has been suggested that carotenoids in chlorosomes provide photoprotection by rapidly quenching triplet excited states of BChl via a triplet-triplet energy transfer mechanism that prevents energy transfer to oxygen and the formation of harmful singlet oxygen. In this work we studied triplet energy transfer kinetics and photodegradation of chlorosomes isolated from wild-type Chlorobium tepidum and from genetically modified species with different types of carotenoids and from a carotenoid-free mutant. Supporting a photoprotective function of carotenoids, carotenoid-free chlorosomes photodegrade approximately 3 times faster than wild-type chlorosomes. However, a significant fraction of the BChls forms a long-lived, triplet-like state that does not interact with carotenoids or with oxygen. We propose that these states are triplet excitons that form due to triplet-triplet interaction between the closely packed BChls. Numerical exciton simulations predict that the energy of these triplet excitons may fall below that of singlet oxygen and triplet carotenoids; this would prevent energy transfer from triplet BChl. Thus, the formation of triplet excitons in chlorosomes serves as an alternative photoprotection mechanism.     

Altering the exciton landscape by removal of specific chlorophylls in monomeric LHCII provides information on the sites of triplet formation and quenching by means of ODMR and EPR spectroscopies

The triplet states populated under illumination in the monomeric light-harvesting complex II (LHCII) were analyzed by EPR and Optically Detected Magnetic Resonance (ODMR) in order to fully characterize the perturbations introduced by site-directed mutations leading to the removal of key chlorophylls. We considered the A2 and A5 mutants, lacking Chls a612(a611) and Chl a603 respectively, since these Chls have been proposed as the sites of formation of triplet states which are subsequently quenched by the luteins. Chls a612 and Chl a603 belong to the two clusters determining the low energy exciton states in the complex. Their removal is expected to significantly alter the excitation energy transfer pathways. On the basis of the TR- and pulse EPR triplet spectra, the two symmetrically related pairs constituted by Chl a612/Lut620 and Chl a603/Lut621 were both possible candidate for triplet-triplet energy transfer (TTET). However, the ODMR results clearly show that only Lut620 is involved in triplet quenching. In the A5 mutant, the Chl a612/Lut620 pair retains this pivotal photoprotective role, while the A2 mutant was found to activate an alternative pathway involving the Chl a603/Lut621pair. These results shows that LHCII is characterized by a robust photoprotective mechanism, able to adapt to the removal of individual chromophores while maintaining a remarkable degree of Chl triplet quenching. Small amounts of unquenched Chl triplet states were also detected. The analysis of the results allowed us to assign the sites of “unquenched” chlorophyll triplets to Chl a610 and Chl a602.     

Carotenoids and fatty alcohols as non-ionic hydrophobic inhibitors of photochemical activities of chioroplasts

Carotenoids and fatty alcohols were added to intact spinachchloroplasts in suspension, and the effects of these reagentson the absorption spectrum and photochemical activities of chloroplastswere examined. The addition of lutein, ß-caroteneand neoxanthin transformed a chlorophyll a form (C684) havingan absorption maximum at 684 nm into C666, and the activityof photosystem II decreased parallel with this transformation.The activity of photosystem I was completely unaffected by theeffect of added carotenoids, whereas both cyclic and non-cyclicphotophosphorylations were inhibited by the additions. Theseeffects were reproducibly observed only with a "petroleum ether-soluble"fraction of each carotenoid, monomeric and less polymerizedforms, which are soluble in petroleum ether at –25°C;the "insoluble" fraction of more polymerized forms was ineffective.Comparative experiments with lutein, DCMU and CCCP as inhibitorsand the dependence of inhibition on light intensity suggestedthat the site of inhibition by carotenoids is on the water sideof photosystem II between its reaction center and the CCCP-inhibitingsite. Fatty alcohols with carbon atoms between 8 and 12 producedstronger but less specific effects on the spectrum and activities.At low concentrations the uncoupling of photophosphorylationoccurred and, in a middle concentration range, the activityof photosystem II decreased with the transformation of C684into C666 and with a change in carotenoid bands. At higher concentrations,where the enhanced activity of photosystem I decreased froma maximum to zero, the red band of chlorophyll a further changedin a complex manner and the absorption around 503 nm decreased.We, thus, deduced that C684 and some endogeneous carotenoidsare associated with photosystem II on its water side and thatadded carotenoids and fatty alcohols at lower concentrationschange their states, resulting in the inhibition of this photosystem. (Received December 20, 1971; )     

Red pool chlorophylls of photosystem I of the cyanobacterium Thermosynechococcus elongatus: a single-molecule study

Photosystem I reaction centers of the cyanobacterium Thermosynechococcus elongatus have been investigated using single-molecule spectroscopy. Single-molecule fluorescence emission spectra reveal a new fluorescence band located at 745 nm. Fluorescence polarization spectroscopy and fluorescence autocorrelation analysis show that only a few chlorophylls are responsible for the photoemission from the Photosystem I trimer at low temperature. Intersystem crossing parameters of the red pool chlorophylls have been determined via fluorescence autocorrelation measurements. The triplet yield of the red chlorophylls is strongly reduced in comparison to chlorophyll a in solution. Strong quenching of the triplet state indicates that the red chlorophylls are located in close contact to carotenoids.     

Structural modeling of the Lhca4 Subunit of LHCI-730 peripheral antenna in photosystem I based on similarity with LHCII

Peripheral chlorophyll a/b binding antenna of photosystem I (LHCI) from green algae and higher plants binds specific low energy absorbing chlorophylls (red pigments) that give rise to a unique red-shifted emission. A three-dimensional structural model of the Lhca4 polypeptide from the LHCI from higher plants was constructed on the basis of comparative sequence analysis, secondary structure prediction, and homology modeling using LHCII as a template. The obtained model of Lhca4 helps to visualize protein ligands to nine chlorophylls (Chls) and three potential His residues to extra Chls. Central domain of the Lhca4 comprising the first (A) and the third (C) transmembrane (TM) helices that binds 6 Chl molecules and two carotenoids is conserved structurally, whereas the interface between the first and the second TM helices and the outer surface of the second TM helix differ significantly among the LHCI and LHCII polypeptides. The model of Lhca4 predicts a histidine residue in the second TM helix, a potential binding site for extra Chl in close proximity to Chls a5 and b5 (labeling by Kühlbrandt). The interpigment interactions in the formed pigment cluster are suggested to cause a red spectral shift in absorption and emission. Modeling of the LHCI-730 heterodimer based on the model structures of Lhca1 and Lhca4 allowed us to suggest potential sites of pigment-pigment interactions that might be formed upon heterodimerization or docking of the LHCI dimers to the surface of PSI.     

Photosynthetic activity of far-red light in green plants

We have found that long-wavelength quanta up to 780 nm support oxygen evolution from the leaves of sunflower and bean. The far-red light excitations are supporting the photochemical activity of photosystem II, as is indicated by the increased chlorophyll fluorescence in response to the reduction of the photosystem II primary electron acceptor, Q(A). The results also demonstrate that the far-red photosystem II excitations are susceptible to non-photochemical quenching, although less than the red excitations. Uphill activation energies of 9.8+/-0.5 kJ mol(-1) and 12.5+/-0.7 kJ mol(-1) have been revealed in sunflower leaves for the 716 and 740 nm illumination, respectively, from the temperature dependencies of quantum yields, comparable to the corresponding energy gaps of 8.8 and 14.3 kJ mol(-1) between the 716 and 680 nm, and the 740 and 680 nm light quanta. Similarly, the non-photochemical quenching of far-red excitations is facilitated by temperature confirming thermal activation of the far-red quanta to the photosystem II core. The observations are discussed in terms of as yet undisclosed far-red forms of chlorophyll in the photosystem II antenna, reversed (uphill) spill-over of excitation from photosystem I antenna to the photosystem II antenna, as well as absorption from thermally populated vibrational sub-levels of photosystem II chlorophylls in the ground electronic state. From these three interpretations, our analysis favours the first one, i.e., the presence in intact plant leaves of a small number of far-red chlorophylls of photosystem II. Based on analogy with the well-known far-red spectral forms in photosystem I, it is likely that some kind of strongly coupled chlorophyll dimers/aggregates are involved. The similarity of the result for sunflower and bean proves that both the extreme long-wavelength oxygen evolution and the local quantum yield maximum are general properties of the plants.     

Role of the surface charges D72 and K8 in the function and structural stability of the cytochrome c from Nostoc sp. PCC 7119

We investigated the role of electrostatic charges at positions D72 and K8 in the function and structural stability of cytochrome c6 from Nostoc sp. PCC 7119 (cyt c6). A series of mutant forms was generated to span the possible combinations of charge neutralization (by mutation to alanine) and charge inversion (by mutation to lysine and aspartate, respectively) in these positions. All forms of cyt c6 were functionally characterized by laser flash absorption spectroscopy, and their stability was probed by urea-induced folding equilibrium relaxation experiments and differential scanning calorimetry. Neutralization or inversion of the positive charge at position K8 reduced the efficiency of electron transfer to photosystem I. This effect could not be reversed by compensating for the change in global charge that had been introduced by the mutation, indicating a specific role for K8 in the formation of the electron transfer complex between cyt c6 and photosystem I. Replacement of D72 by asparagine or lysine increased the efficiency of electron transfer to photosystem I, but destabilized the protein. D72 apparently participates in electrostatic interactions that stabilize the structure of cyt c6. The destabilizing effect was reduced when aspartate was replaced by the small amino acid alanine. Complementing the mutation D72A with a charge neutralization or inversion at position K8 led to mutant forms of cyt c6 that were more stable than the wild-type under all tested conditions.     

The Lhca antenna complexes of higher plants photosystem I

The Lhca antenna complexes of photosystem I (PSI) have been characterized by comparison of native and recombinant preparations. Eight Lhca polypeptides have been found to be all organized as dimers in the PSI-LHCI complex. The red emission fluorescence is associated not only with Lhca1-4 heterodimer, but also with dimers containing Lhca2 and/or Lhca3 complexes. Reconstitution of Lhca1 and Lhca4 monomers as well as of the Lhca1-4 dimer in vitro was obtained. The biochemical and spectroscopic features of these three complexes are reported. The monomers Lhca1 and Lhca4 bind 10 Chls each, while the Chl a/b ratio is lower in Lhca4 as compared to Lhca1. Three carotenoid binding sites have been found in Lhca1, while only two are present in Lhca4. Both complexes contain lutein and violaxanthin while beta-carotene is selectively bound to the Lhca1-4 dimer in substoichiometric amounts upon dimerization. Spectral analysis revealed the presence of low energy absorption forms in Lhca1 previously thought to be exclusively associated with Lhca4. It is shown that the process of dimerization changes the spectroscopic properties of some chromophores and increases the amplitude of the red absorption tail of the complexes. The origin of these spectroscopic features is discussed.     

Photothermal Spectra of Thylakoids Isolated from Cucumber Cotyledons at Various Stages of Greening

The character of interaction between carotenoids (Cars) and chlorophylls (Chls) in thylakoids isolated from cucumber cotyledons at three stages of greening (3, 6, and 24 h of irradiation with 120 µmol m^–2 s^–1) was studied. The shapes of the steady state photoacoustic spectra were changed with the change in time of greening and with the frequency of radiation modulation. The shapes show that changes not only in the contents of various pigments but also in pigment interactions with surrounding occur and that processes of thermal deactivation characterised by different kinetics take place. Slow processes of thermal deactivation are in most cases due to deactivation of triplet states. Long living triplet states are very often engaged in photochemical reactions that can destroy the tissue. Analysis of the time-resolved photothermal spectra shows that at later stage of greening, the chlorophyll (Chl) molecules are better shielded against photo-destruction because Cars more efficiently quench their triplet states. The yield of formation of the pigment triplet states measured by the time resolved photothermal method, always at the same energy absorbed by pigment mixture, declined during sample greening. The decay time of the slow component of pigment thermal deactivation, due predominantly to deactivation of the triplet state of Chl, decreases with the increase of time of greening from 6.2 µs for the 3-h sample to 1.5 µs for the 24 h sample. The energy taken by Cars from Chls is dissipated into heat, therefore the steady state and quick thermal deactivation values increased during the greening process. The Cars/Chls ratio in the thylakoids decreased during greening approximately 2 fold. Hence at a later phase of greening the Cars can quench the triplet states of Chls more efficiently than at an earlier phase of greening.     

Immunological Quantification of Proteins Related to Light-Harvesting Chlorophyll a/b Protein Complexes of the Two Photosystems in Rice Mutants Totally and Partially Deficient in Chlorophyll b

Ten rice chlorina mutants of Type I, which totally lack chlorophyllb and hence are unable to synthesize light-harvesting chlorophylla/b protein complexes of photosystem II (LHC-II), containedmRNA for proteins related to LHC-II. Immunoblotting with anantiserum, which had been raised against the 24 and 25 kDa apoproteinsof LHC-II and found to cross-react with the 26 kDa protein ofLHC-II and the 20 and 21 kDa apoproteins of light-harvestingchlorophyll a/b protein complexes of photosystem I (LHC-I),revealed that all the five proteins related to LHC-Iand LHC-IIwere present in reduced amounts in the Type I mutants. ThreeType HA mutants, which have a chlorophyll a/b ratio of 10, weremore abundant in the apoproteins, while three Type IIB mutantswith the ratio of 15 were heterogeneous in terms of the apoproteincontent. All the chlorina mutants contained less P700 comparedwith the wild type rice, but were relatively more abundant inthe LHC-I proteins than the LHC-II proteins. The results showthat all the rice chlorina strains are mutants of chlorophyllb synthesis and the deficiency of chlorophyll b differentlyaffects accumulation of the apoproteins of LHC-I and LHC-II.To balance light absorption between the two photosystem, lossof LHC-II is partly counter-balanced by a decrease in the numberof PSI complexes in the mutants. (Received January 21, 1988; Accepted April 28, 1988)     

A picosecond pulse train study of exciton dynamics in photosynthetic membranes.

The fluorescence decay time of spinach chloroplasts at 77 degrees K was determined at 735 nm (corresponding to the photosystem I emission) using a train of 10-ps laser pulses spaced 10 ns apart. The fluorescence lifetime is constant at congruent to 1.5 ns for up to the fourth pulse, but then decreases with increasing pulse number within the pulse train. This quenching is attributed to triplet excited states, and it is concluded that triplet excitons exhibit a time lag of about 50 ns in diffusing from light harvesting antenna pigments to photosystem I pigments. The diffusion coefficient of triplet excitons is a least 300--400 times slower than the diffusion coefficient of singlet excitons in chloroplast membranes.     

The Lhca antenna complexes of higher plants photosystem I

The Lhca antenna complexes of photosystem I (PSI) have been characterized by comparison of native and recombinant preparations. Eight Lhca polypeptides have been found to be all organized as dimers in the PSI-LHCI complex. The red emission fluorescence is associated not only with Lhca1-4 heterodimer, but also with dimers containing Lhca2 and/or Lhca3 complexes. Reconstitution of Lhca1 and Lhca4 monomers as well as of the Lhca1-4 dimer in vitro was obtained. The biochemical and spectroscopic features of these three complexes are reported. The monomers Lhca1 and Lhca4 bind 10 Chls each, while the Chl a/b ratio is lower in Lhca4 as compared to Lhca1. Three carotenoid binding sites have been found in Lhca1, while only two are present in Lhca4. Both complexes contain lutein and violaxanthin while β-carotene is selectively bound to the Lhca1-4 dimer in substoichiometric amounts upon dimerization. Spectral analysis revealed the presence of low energy absorption forms in Lhca1 previously thought to be exclusively associated with Lhca4. It is shown that the process of dimerization changes the spectroscopic properties of some chromophores and increases the amplitude of the red absorption tail of the complexes. The origin of these spectroscopic features is discussed.     

Time-resolved fluorescence emission measurements of photosystem I particles of various cyanobacteria: a unified compartmental model.

Photosystem I (PS-I) contains a small fraction of chlorophylls (Chls) that absorb at wavelengths longer than the primary electron donor P700. The total number of these long wavelength Chls and their spectral distribution are strongly species dependent. In this contribution we present room temperature time-resolved fluorescence data of five PS-I core complexes that contain different amounts of these long wavelength Chls, i.e., monomeric and trimeric photosystem I particles of the cyanobacteria Synechocystis sp. PCC 6803, Synechococcus elongatus, and Spirulina platensis, which were obtained using a synchroscan streak camera. Global analysis of the data reveals considerable differences between the equilibration components (3.4-15 ps) and trapping components (23-50 ps) of the various PS-I complexes. We show that a relatively simple compartmental model can be used to reproduce all of the observed kinetics and demonstrate that the large kinetic differences are purely the result of differences in the long wavelength Chl content. This procedure not only offers rate constants of energy transfer between and of trapping from the compartments, but also well-defined room temperature emission spectra of the individual Chl pools. A pool of red shifted Chls absorbing around 702 nm and emitting around 712 nm was found to be a common feature of all studied PS-I particles. These red shifted Chls were found to be located neither very close to P700 nor very remote from P700. In Synechococcus trimeric and Spirulina monomeric PS-I cores, a second pool of red Chls was present which absorbs around 708 nm, and emits around 721 nm. In Spirulina trimeric PS-I cores an even more red shifted second pool of red Chls was found, absorbing around 715 nm and emitting at 730 nm.     

Excited states and primary charge separation in the pigment system of the green photosynthetic bacterium Prosthecochloris aestuarii as studied by picosecond absorbance difference spectroscopy

Picosecond absorbance difference spectra at a number of delay times after a 35 ps excitation flash and kinetics of absorbance changes were measured of the membrane vesicle preparation Complex I from the photosynthetic green sulfur bacterium Prosthecochloris aestuarii. After chemical oxidation of the primary donor the excitation pulse produced singlet and triplet excited states of carotenoid and bacteriochlorophyll a. With active reaction centers present also the flash-induced primary charge separation and subsequent electron transfer were observed. The singlet excited state of the carotenoid, formed by direct excitation at 532 nm, is characterized by an absorbance band peaking at 590 nm. Its average lifetime was calculated to be about 1 ps. Excited singlet states of bacteriochlorophyll a were characterized by a bleaching of their ground state Q_y absorption bands. Singlet excited states, localized on the so-called core complex, were produced by energy transfer from excited carotenoid. Their lifetime was about 70 ps. A decay component of about 280 ps was ascribed to singlet excited bacteriochlorophyll a in the bacteriochlorophyll a protein. These singlet excitations were partly converted to the triplet state. With active reaction centers, oxidation of the primary donor, P-840, characterized by the bleaching of its Q_y and Q_x absorption bands, was observed. This oxidation was accompanied by a bleaching between 650 and 680 nm and an absorbance increase between 680 and 750 nm. These changes, presumably due to reduction of bacteriopheophytin c (Van Bochove, A.C., Swarthoff, T., Kingma, H., Hof, R.M., Van Grondelle, R., Duysens, L.N.M. and Amesz, J. (1984) Biochim. Biophys. Acta 764, 343–346), were attributed to the reduction of the primary electron acceptor. Electron transfer to a secondary acceptor occurred with a time-constant of 550 ± 50 ps. Since no absorbance changes due to reduction of this acceptor were observed in the red or infrared region, we tentatively assume that this acceptor is an iron-sulfur center.     

Electrochromic shift of chlorophyll absorption in photosystem I from Synechocystis sp. PCC 6803: a probe of optical and dielectric properties around the secondary electron acceptor

Nanosecond absorption dynamics at approximately 685 nm after excitation of photosystem I (PS I) from Synechocystis sp. PCC 6803 is consistent with electrochromic shift of absorption bands of the Chl a pigments in the vicinity of the secondary electron acceptor A(1). Based on experimental optical data and structure-based simulations, the effective local dielectric constant has been estimated to be between 3 and 20, which suggests that electron transfer in PS I is accompanied by considerable protein relaxation. Similar effective dielectric constant values have been previously observed for the bacterial photosynthetic reaction center and indicate that protein reorganization leading to effective charge screening may be a necessary structural property of proteins that facilitate the charge transfer function. The data presented here also argue against attributing redmost absorption in PS I to closely spaced antenna chlorophylls (Chls) A38 and A39, and suggest that optical transitions of these Chls, along with that of connecting chlorophyll (A40) lie in the range 680-695 nm.