Minor complexes at work: light-harvesting by carotenoids in the photosystem II antenna complexes CP24 and CP26

Plant photosynthesis relies on the capacity of chlorophylls and carotenoids to absorb light. One of the roles of carotenoids is to harvest green-blue light and transfer the excitation energy to the chlorophylls. The corresponding dynamics were investigated here for the first time, to our knowledge, in the CP26 and CP24 minor antenna complexes. The results for the two complexes differ substantially. In CP26 fast transfer (80 fs) occurs from the carotenoid S₂ state to chlorophylls a absorbing at 675 and 678 nm, whereas transfer from the hot S₁ state to the lowest energy chlorophylls is observed in <1 ps. In CP24, energy transfer from the S₂ state leads in 80 fs to the population of chlorophylls b and high-energy chlorophylls a absorbing at 670 nm, whereas the low-energy chlorophylls a are populated only in several picoseconds. The results suggest that CP26 has a structural and functional organization similar to that of LHCII, whereas CP24 differs substantially from the other Lhc complexes, especially regarding the lutein L1 binding domain. No energy transfer from the carotenoid S₁ state to chlorophylls was observed in either complex, suggesting that this state is energetically below the chlorophyll Qy state and therefore may play a role in the quenching of chlorophyll excitations.     

The development of antenna complexes of barley (Hordeum vulgare cv. Akcent) under different light conditions as judged from the analysis of 77 K chlorophyll a fluorescence spectra

Using 77 K chlorophyll a (Chl a) fluorescence spectra in vivo, the development was studied of Photosystems II (PS II) and I (PS I) during greening of barley under intermittent light followed by continuous light at low (LI, 50 μmol m⁻² s⁻¹) and high (HI, 1000 μmol m⁻² s⁻¹) irradiances. The greening at HI intermittent light was accompanied with significantly reduced fluorescence intensity from Chl b excitation for both PS II (F685) and PS I (F743), in comparison with LI plants, indicating that assembly of light-harvesting complexes (LHC) of both photosystems was affected to a similar degree. During greening at continuous HI, a slower increase of emission from Chl b excitation in PS II as compared with PS I was observed, indicating a preferred reduction in the accumulation of LHC II. The following characteristics of 77 K Chl a fluorescence spectra documented the photoprotective function of an elevated content of carotenoids in HI leaves: (1) a pronounced suppression of Soret region of excitation spectra (410–450 nm) in comparison with the red region (670–690 nm) during the early stage of greening indicated a strongly reduced excitation energy transfer from carotenoids to the Chl a fluorescing forms within PS I and PS II; (2) changes in the shape of the excitation band of Chl b and carotenoids (460–490 nm) during greening under continuous light confirmed that the energy transfer from carotenoids to Chl a within PS II remained lower as compared with the LI plants. This revised version was published online in June 2006 with corrections to the Cover Date.     

Utilization of blue-green light by chlorosomes from the photosynthetic bacterium Chloroflexus aurantiacus: Ultrafast excitation energy conversion and transfer

Chlorosomes of photosynthetic green bacteria are unique molecular assemblies providing efficient light harvesting followed by multi-step transfer of excitation energy to reaction centers. In each chlorosome, 10⁴–10⁵ bacteriochlorophyll (BChl) c/d/e molecules are organized by self-assembly into high-ordered aggregates. We studied the early-time dynamics of the excitation energy flow and energy conversion in chlorosomes isolated from Chloroflexus (Cfx.) aurantiacus bacteria by pump-probe spectroscopy with 30-fs temporal resolution at room temperature. Both the S₂ state of carotenoids (Cars) and the Soret states of BChl c were excited at ~490 nm, and absorption changes were probed at 400–900 nm. A global analysis of spectroscopy data revealed that the excitation energy transfer (EET) from Cars to BChl c aggregates occurred within ~100 fs, and the Soret → Q energy conversion in BChl c occurred faster within ~40 fs. This conclusion was confirmed by a detailed comparison of the early exciton dynamics in chlorosomes with different content of Cars. These processes are accompanied by excitonic and vibrational relaxation within 100–270 fs. The well-known EET from BChl c to the baseplate BChl a proceeded on a ps time-scale. We showed that the S₁ state of Cars does not participate in EET. We discussed the possible presence (or absence) of an intermediate state that might mediates the Soret → Q_y internal conversion in chlorosomal BChl c. We discussed a possible relationship between the observed exciton dynamics and the structural heterogeneity of chlorosomes.     

Optical spectroscopic studies of light-harvesting by pigment-reconstituted peridinin-chlorophyll-proteins at cryogenic temperatures

Low temperature, steady-state, optical spectroscopic methods were used to study the spectral features of peridinin-chlorophyll-protein (PCP) complexes in which recombinant apoprotein has been refolded in the presence of peridinin and either chlorophyll a (Chl a), chlorophyll b (Chl b), chlorophyll d (Chl d), 3-acetyl-chlorophyll a (3-acetyl-Chl a) or bacteriochlorophyll a (BChl a). Absorption spectra taken at 10 K provide better resolution of the spectroscopic bands than seen at room temperature and reveal specific pigment–protein interactions responsible for the positions of the Q_y bands of the chlorophylls. The study reveals that the functional groups attached to Ring I of the two protein-bound chlorophylls modulate the Q_y and Soret transition energies. Fluorescence excitation spectra were used to compute energy transfer efficiencies of the various complexes at room temperature and these were correlated with previously reported ultrafast, time-resolved optical spectroscopic dynamics data. The results illustrate the robust nature and value of the PCP complex, which maintains a high efficiency of antenna function even in the presence of non-native chlorophyll species, as an effective tool for elucidating the molecular details of photosynthetic light-harvesting.     

Direct evidence for excitonically coupled chlorophylls a and b in LHC II of higher plants by nonlinear polarization spectroscopy in the frequency domain

Occurrence of excitonic interactions in light-harvesting complex II (LHC II) was investigated by nonlinear polarization spectroscopy in the frequency domain (NLPF) at room temperature. NLPF spectra were obtained upon probing in the chlorophyll (Chl) a/b Soret region and pumping in the Q_y region. The lowest energy Chl a absorbing at 678 nm is strongly excitonically coupled to Chl b.     

Structural-functional organization of the main light harvesting complex and photosystem 2 of higher plants

The interrelation between spectral and structural–functional properties of LhcIIb was studied. The dipole strength of the main Q_y bands of chlorophylls (Chl a 30.8 D²; Chl b 18.5 D²) and chlorophyll a/b ratio (Chl a/Chl b = 7 : 6) were determined for LhcIIb. The Chl a/Chl b value shows that the subunit of this complex contains seven Chl a and six Chl b molecules. Individual bands of chlorophylls (bands in stokes and anti-Stokes parts at 77 K were Lorentzian and Gaussian, respectively) were resolved using synchronized deconvolution of absorption, CD, and LD bands of chlorophylls. Seven of these bands belonged to Chl a. Parameters of absorption bands of Chl a indicate that seven molecules represent a united cluster (heptamer) with exciton interactions, determining the spectrum of LhcIIb in the Chl a absorption region. Parameters of absorption bands of Chl b show the existence of three clusters: monomer (639.6 nm), dimer (645.2 and 647.4 nm), and trimer (649.8 and 654.1 nm). These clusters and their properties agree with the well-known structure of porphyrin groups of the LhcIIb subunit (Kuhlbrandt, 1994). A distorted ring of seven porphyrins in the stromal range of the subunit corresponds to Chl a heptamer; a separately located molecule near the N-terminal domain on the stromal side of the subunit corresponds to Chl b monomer; a dimer and a trimer of porphyrins in the lumenal range of the subunit correspond to the dimer and trimer of Chl b, respectively. The calculated lifetimes of the excitation energy (exciton) transfer in subunit and trimer of LhcIIb confirm this location of pigments. The geometry of the Chl a heptamer (mutual orientation of transition dipole moments) was determined by the steady-state Kasha–Tinoco approximation using parameters of individual bands of exciton splitting. The calculated parameters of mutual orientation of Chl a dipoles agree with the topography of the stromal porphyrins found by electron crystallography (Kuhlbrandt, 1994). A structural model of the granal multicentral macrocomplex of PSII (MPSII) is suggested. The lifetimes of the exciton migration between the main pigment–protein compartments of MPSII were calculated. The results of calculation are consistent with the structural model of the photosystem. The location of pigments provides for fast exciton hopping between Chl a clusters of neighboring proteins in the MPSII along the stromal surface within the membrane (5-25 psec) and between stacked membranes (40 psec) of chloroplast grana.     

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.     

Excitation energy pathways in the photosynthetic units of reaction center LM- and H-subunit deletion mutants of <Emphasis Type="Italic">Rhodospirillum rubrum</Emphasis>

Light-induced reaction dynamics of isolated photosynthetic membranes obtained from wild-type (WT) and reaction center (RC)-subunit deletion strains SPUHK1 (an H-subunit deletion mutant) and SKΔLM (an (L+M) deletion mutant) of the purple non-sulphur bacterium Rhodospirillum rubrum have been investigated by femtosecond transient absorption spectroscopy. Upon excitation of the spirilloxanthin (Spx) S₂ state at 546 nm, of the bacteriochlorophyll Soret band at 388 nm and probing spectral regions, which are characteristic for carotenoids, similar dynamics in the SPUHK1, SKΔLM and WT strains could be observed. The excitation of Spx S₂ is followed by the simultaneous population of the lower singlet excited states S₁ and S* which decay with lifetimes of 1.4 and 5 ps, respectively for the mutants, and 1.4 and 4 ps, respectively, for the wild-type. The excitation of the BChl Soret band is followed by relaxation into BChl lower excited states which compete with excitation energy transfer BChl-to-Spx. The deexcitation pathway BChl(Soret) → Spx(S₂) → Spx(S₁) occurs with the same transition rate for all investigated samples (WT, SPUHK1 and SKΔLM). The kinetic traces measured for the Spx S₁ → S_N transition display similar behaviour for all samples showing a positive signal which increases within the first 400 fs (i.e. the time needed for the excitation energy to reach the Spx S₁ excited state) and decays with a lifetime of about 1.5 ps. This suggests that the Spx excited state dynamics in the investigated complexes do not differ significantly. Moreover, a longer excited state lifetime of BChl for SPUHK1 in comparison to WT was observed, consistent with a photochemical quenching channel present in the presence of RC. For long delay times, photobleaching of the RC special pair and an electrochromic blue shift of the monomeric BChl a can be observed only for the WT but not for the mutants. The close similarity of the excited state decay processes of all strains indicates that the pigment geometry of the LH1 complex in native membranes is unaffected by the presence of an RC and allows us to draw a model representation of the WT, SKΔLM and SPUHK1 PSU complexes.     

The back and forth of energy transfer between carotenoids and chlorophylls and its role in the regulation of light harvesting

Many aspects in the regulation of photosynthetic light-harvesting of plants are still quite poorly understood. For example, it is still a matter of debate which physical mechanism(s) results in the regulation and dissipation of excess energy in high light. Many researchers agree that electronic interactions between chlorophylls (Chl) and certain states of carotenoids are involved in these mechanisms. However, in particular, the role of the first excited state of carotenoids (Car S₁) is not easily revealed, because of its optical forbidden character. The use of two-photon excitation is an elegant approach to address directly this state and to investigate the energy transfer in the direction Car S₁ → Chl. Meanwhile, it has been applied to a large variety of systems starting from simple carotenoid–tetrapyrrole model compounds up to entire plants. Here, we present a systematic summary of the observations obtained by two-photon excitation about Car S₁ → Chl energy transfer in systems with increasing complexity and the correlation to fluorescence quenching. We compare these observations directly with the energy transfer in the opposite direction, Chl → Car S₁, for the same systems as obtained in pump-probe studies. We discuss what surprising aspects of this comparison led us to the suggestion that quenching excitonic Car–Chl interactions could contribute to the regulation of light harvesting, and how this suggestion can be connected to other models proposed.     

Three photosynthetic antenna porphyrins in a primitive green alga

Fluorescence excitation spectra (between 400–500 and 610–700 nm) for chlorophyll emission from particles and detergent extracts of the primitive green microalga, Mantoniella, were measured. The results showed that the prophyrin, magnesium 2,4-divinylpheoporphyrin a₅, which this alga accumulates in addition to Chl b, also can transfer excitation energy to Chl a, and therefore act as antenna for photosynthesis. Evidence was found that magnesium 2,4-divinylpheoporphyrin a₅ has a Soret band near 450 nm in vivo which further increases the light-harvesting capacity of these algae growing deep in the open ocean.     

Effect of detergent on aggregation of the light-harvesting chlorophyll a/b complex of photosystem 2 and its impact for carotenoid function and fluorescence quenching

Spectroscopy was used to investigate the fluorescence quenching mechanism in light-harvesting complex 2 (LHC2). The 77 K fluorescence excitation spectroscopy was performed for detection of aggregation state of LHC2 treated with different concentrations of octylphenol poly(ethyleneglycol ether)₁₀ (TX-100). Resonance Raman (RR) spectra excited with 488, 496, and 514 nm provided molecular configuration of neoxanthin, lutein 1, and lutein 2, respectively. At increased concentration of TX-100, the RR signals of xanthophylls were enhanced in the four frequency regions, which was accompanied with increase of fluorescence of chlorophyll (Chl) a. Thus the absorption of the three xanthophyll molecules was inclined to excitation wavelength, which proved that functional configurations of xanthophyll molecules in LHC2 were vital for fast transfer of excitation energy to Chl a molecules. Changes in the v4 region (C-H out-of-plane bending modes, at ∼960 cm⁻¹ in RR spectra) demonstrated that the twist feature of neoxanthin, lutein 1, and lutein 2 molecules existed in LHC2 trimers, however, it was lost in the LHC2 macro-aggregates. In the second derivative absorption spectra of LHC2, neoxanthin absorption was not detected in LHC2 macro-aggregates, while evident absorption was found in LHC2 trimers and this absorption decreased obviously when TX-100 concentration was higher than 1 mM. Hence the neoxanthin molecule had a structural role in formation of LHC2 trimers. The RR and absorption spectra also implied that carotenoid molecules constructed the functional LHC2 trimers via their intrinsic configuration features, which enabled energy transfer to Chl a efficiently and led to lower fluorescence quenching efficiency. In contrast, these intrinsic twist configurations were lost in LHC2 macro-aggregates and led to lower energy transfer efficiency and higher fluorescence quenching efficiency.     

Energy transfer pathways in the CP24 and CP26 antenna complexes of higher plant photosystem II: a comparative study

Antenna complexes are key components of plant photosynthesis, the process that converts sunlight, CO₂, and water into oxygen and sugars. We report the first (to our knowledge) femtosecond transient absorption study on the light-harvesting pigment-protein complexes CP26 (Lhcb5) and CP24 (Lhcb6) of Photosystem II. The complexes are excited at three different wavelengths in the chlorophyll (Chl) Qy region. Both complexes show a single subpicosecond Chl b to Chl a transfer process. In addition, a reduction in the population of the intermediate states (in the 660-670 nm range) as compared to light-harvesting complex II is correlated in CP26 to the absence of both Chls a604 and b605. However, Chl forms around 670 nm are still present in the Chl a Qy range, which undergoes relaxation with slow rates (10-15 ps). This reduction in intermediate-state amplitude CP24 shows a distinctive narrow band at 670 nm connected with Chls b and decaying to the low-energy Chl a states in 3-5 ps. This 670 nm band, which is fully populated in 0.6 ps together with the Chl a low-energy states, is proposed to originate from Chl 602 or 603. In this study, we monitored the energy flow within two minor complexes, and our results may help elucidate these structures in the future.     

Determination of the primary charge separation rate in Photosystem II reaction centers at 15 K

We have measured the rate constant for the formation of the oxidized chlorophyll a electron donor (P680⁺) and the reduced electron acceptor pheophytin a ^– (Pheo a ^–) following excitation of isolated Photosystem II reaction centers (PS II RC) at 15 K. This PS II RC complex consists of D₁, D₂, and cytochrome b-559 proteins and was prepared by a procedure which stabilizes the protein complex. Transient absorption difference spectra were measured from 450–840 nm as a function of time with 500fs resolution following 610 nm laser excitation. The formation of P680⁺-Pheo a ^– is indicated by the appearance of a band due to P680⁺ at 820 nm and corresponding absorbance changes at 490, 515 and 546 nm due to the formation of Pheo a ^–. The appearance of the 490 nm and 820 nm bands is monoexponenital with =1.4±0.2 ps. Treatment of the PS II RC with sodium dithionite and methyl viologen followed by exposure to laser excitation results in accumulation of Pheo a ^–. Laser excitation of these prereduced RCs at 15 K results in formation of a transient absorption spectrum assigned to ^1*P680. We observe wavelength-dependent kinetics for the recovery of the transient bleach of the Q_y absorption bands of the pigments in both untreated and pre-reduced PS II RCs at 15K. This result is attributed to an energy transfer process within the PS II RC at low temperature that is not connected with charge separation.Abbreviations PS I Photosystem I - PS II Photosystem II - RC reaction center - P680 primary electron donor in Photosystem II - Chl a chlorophyll a - Pheo a pheophytin a     

Singlet and triplet excited carotenoid and antenna bacteriochlorophyll of the photosynthetic purple bacterium Rhodospirillum rubrum as studied by picosecond absorbance difference spectroscopy

Picosecond absorbance difference spectra at a number of delay times after a 35 ps excitation pulse and kinetics of absorbance changes were measured in chromatophores of the photosynthetic purple bacterium Rhodospirillum rubrum after chemical oxidation of the primary electron donor P-875. Kinetics and spectra were measured of the excited singlet states of carotenoid and bacteriochlorophyll a and also of the triplet state of the carotenoid. The excited singlet state of carotenoid, produced by direct excitation at 532 nm, is characterized by a bleaching of the ground state absorption bands in the region 450–490 nm and by an absorbance increase with a maximum near 570 nm. Its lifetime was calculated to be 0.6 ± 0.1 ps in vitro and less than 1 ps in vivo. The triplet state of carotenoid in vivo is formed within 100 ps after direct carotenoid excitation via a pathway that does not involve excited states of bacteriochlorophyll. Singlet excitation of a bacteriochlorophyll a molecule causes the bleaching of its Q_x and Q_y absorption bands, and is probably associated with blue shifts of the Q_y absorption band of about six neighboring bacteriochlorophyll molecules. Upon increasing the excitation density, the average lifetime of the singlet excitations on bacteriochlorophyll decreased from about 350 ps to about 10 ps or less. The results are in quantitative agreement with the known effect of singlet-singlet annihilation upon the fluorescence yield, and furthermore show that no bacteriochlorophyll or carotenoid triplet formation is associated with this annihilation.     

Excitation relaxation dynamics and energy transfer in fucoxanthin–chlorophyll a/c-protein complexes,probed by time-resolved fluorescence

In algae, light-harvesting complexes contain specific chlorophylls (Chls) and keto-carotenoids; Chl a, Chl c, and fucoxanthin (Fx) in diatoms and brown algae; Chl a, Chl c, and peridinin in photosynthetic dinoflagellates; and Chl a, Chl b, and siphonaxanthin in green algae. The Fx–Chl a/c-protein (FCP) complex from the diatom Chaetoceros gracilis contains Chl c₁, Chl c₂, and the keto-carotenoid, Fx, as antenna pigments, in addition to Chl a. In the present study, we investigated energy transfer in the FCP complex associated with photosystem II (FCPII) of C. gracilis. For these investigations, we analyzed time-resolved fluorescence spectra, fluorescence rise and decay curves, and time-resolved fluorescence anisotropy data. Chl a exhibited different energy forms with fluorescence peaks ranging from 677 nm to 688 nm. Fx transferred excitation energy to lower-energy Chl a with a time constant of 300 fs. Chl c transferred excitation energy to Chl a with time constants of 500–600 fs (intra-complex transfer), 600–700 fs (intra-complex transfer), and 4–6 ps (inter-complex transfer). The latter process made a greater contribution to total Chl c-to-Chl a transfer in intact cells of C. gracilis than in the isolated FCPII complexes. The lower-energy Chl a received excitation energy from Fx and transferred the energy to higher-energy Chl a. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: Keys to Produce Clean Energy.     

What can we still learn from the electrochromic band-shifts in Photosystem II?

Electrochromic band-shifts have been investigated in Photosystem II (PSII) from Thermosynechoccocus elongatus. Firstly, by using Mn-depleted PsbA1-PSII and PsbA3-PSII in which the Q_X absorption of Phe_D1 differs, a band-shift in the Q_X region of Phe_D2 centered at ~ 544 nm has been identified upon the oxidation, at pH 8.6, of Tyr_D. In contrast, a band-shift due to the formation of either Q_A^•- or Tyr_Z^• is observed in PsbA3-PSII at ~ 546 nm, as expected with E130 H-bonded to Phe_D1 and at ~ 544 nm as expected with Q130 H-bonded to Phe_D1. Secondly, electrochromic band-shifts in the Chla Soret region have been measured in O₂-evolving PSII in PsbA3-PSII, in the PsbA3/H198Q mutant in which the Soret band of P_D1 is blue shifted and in the PsbA3/T179H mutant. Upon Tyr_Z^•Q_A^•- formation the Soret band of P_D1 is red shifted and the Soret band of Chl_D1 is blue shifted. In contrast, only P_D1 undergoes a detectable S-state dependent electrochromism. Thirdly, the time resolved S-state dependent electrochromism attributed to P_D1 is biphasic for all the S-state transitions except for S₁ to S₂, and shows that: i) the proton release in S₀ to S₁ occurs after the electron transfer and ii) the proton release and the electron transfer kinetics in S₂ to S₃, in T. elongatus, are significantly faster than often considered. The nature of S₂Tyr_Z^• is discussed in view of the models in the literature involving intermediate states in the S₂ to S₃ transition.     

Energy transfer pathways in the CAC light-harvesting complex of Rhodomonas salina

Photosynthetic organisms had to evolve diverse mechanisms of light-harvesting to supply photosynthetic apparatus with enough energy. Cryptophytes represent one of the groups of photosynthetic organisms combining external and internal antenna systems. They contain one type of immobile phycobiliprotein located at the lumenal side of the thylakoid membrane, together with membrane-bound chlorophyll a/c antenna (CAC). Here we employ femtosecond transient absorption spectroscopy to study energy transfer pathways in the CAC proteins of cryptophyte Rhodomonas salina. The major CAC carotenoid, alloxanthin, is a cryptophyte-specific carotenoid, and it is the only naturally-occurring carotenoid with two triple bonds in its structure. In order to explore the energy transfer pathways within the CAC complex, three excitation wavelengths (505, 590, and 640 nm) were chosen to excite pigments in the CAC antenna. The excitation of Chl c at either 590 or 640 nm proves efficient energy transfer between Chl c and Chl a. The excitation of alloxanthin at 505 nm shows an active pathway from the S₂ state with efficiency around 50%, feeding both Chl a and Chl c with approximately 1:1 branching ratio, yet, the S₁-route is rather inefficient. The 57 ps energy transfer time to Chl a gives ~25% efficiency of the S₁ channel. The low efficiency of the S₁ route renders the overall carotenoid-Chl energy transfer efficiency low, pointing to the regulatory role of alloxanthin in the CAC antenna.     

Energy transfer in reconstituted peridinin-chlorophyll-protein complexes: ensemble and single-molecule spectroscopy studies

We combine ensemble and single-molecule spectroscopy to gain insight into the energy transfer between chlorophylls (Chls) in peridinin-chlorophyll-protein (PCP) complexes reconstituted with Chl a, Chl b, as well as both Chl a and Chl b. The main focus is the heterochlorophyllous system (Chl a/b-N-PCP), and reference information essential to interpret experimental observations is obtained from homochlorophyllous complexes. Energy transfer between Chls in Chl a/b-N-PCP takes place from Chl b to Chl a and also from Chl a to Chl b with comparable Förster energy transfer rates of 0.0324 and 0.0215 ps⁻¹, respectively. Monte Carlo simulations yield the ratio of 39:61 for the excitation distribution between Chl a and Chl b, which is larger than the equilibrium distribution of 34:66. An average Chl a/Chl b fluorescence intensity ratio of 66:34 is measured, however, for single Chl a/b-N-PCP complexes excited into the peridinin (Per) absorption. This difference is attributed to almost three times more efficient energy transfer from Per to Chl a than to Chl b. The results indicate also that due to bilateral energy transfer, the Chl system equilibrates only partially during the excited state lifetimes.     

Fluorescence properties of the allenic carotenoid fucoxanthin: Implication for energy transfer in photosynthetic pigment systems

The fluorescence spectrum of an allenic carotenoid, all-trans-fucoxanthin isolated from a brown alga, has been reported for the first time. This carotenoid is known to function efficiently as a primary photosynthetic antenna pigment in marine algae. The emission bands were located around 630, 685 and 750 nm in CS₂ at 20°C, absorption bands being located at 448, 476 and 505 nm. The energy difference between the 0-0 bands of absorption and emission spectra was about 3900 cm^-1 and location of the emission maximum was less sensitive to the polarizability of solvents than that of the absorption maximum. These clearly indicate that the emission originates from the optically forbidden singlet state (2A_g). This is in contrast to other carotenoids whose emission is assigned to 1B_u state, probably due to the symmetric structure of the conjugated double bond responsible for the absorption in the visible region. A rapid internal conversion from 1B_u to 2A_g state might be facilitated by distorted structure of the conjugated double bond of fucoxanthin. The energy level responsible for the emission is almost identical to the Q_y level of the acceptor molecule (Chl a), thus we propose an energy transfer pathway from the optically forbidden 2A_g state of the carotenoid to the Q_y transition of Chl a in algal pigment systems.