Article pubs.acs.org/JPCB
Modeling of Optical Spectra of the Light-Harvesting CP29 Antenna Complex of Photosystem IIPart II Ximao Feng,† Adam Kell,† Jörg Pieper,‡ and Ryszard Jankowiak*,†,§,∥ †
Department of Chemistry, Kansas State University, Manhattan, Kansas 66506, United States Institute of Physics, University of Tartu, Tartu, Estonia § Faculty of Applied Physics and Mathematics, Gdańsk University of Technology, Gdańsk, Poland ‡
S Supporting Information *
ABSTRACT: Until recently, it was believed that the CP29 protein from higher plant photosystem II (PSII) contains 8 chlorophylls (Chl’s) per complex (Ahn et al. Science 2008, 320, 794−797; Bassi et al. Proc. Natl. Acad. Sci. U.S.A. 1999, 96, 10056−10061) in contrast to the 13 Chl’s revealed by the recent X-ray structure (Pan et al. Nat. Struct. Mol. Biol. 2011, 18, 309−315). This disagreement presents a constraint on the interpretation of the underlying electronic structure of this complex. To shed more light on the interpretation of various experimental optical spectra discussed in the accompanying paper (part I, DOI 10.1021/ jp4004328), we report here calculated low-temperature (5 K) absorption, fluorescence, hole-burned (HB), and 300 K circular dichroism (CD) spectra for CP29 complexes with a different number of pigments. We focus on excitonic structure and the nature of the lowenergy state using modeling based on the X-ray structure of CP29 and Redfield theory. We show that the lowest energy state is mostly contributed to by a612, a611, and a615 Chl’s. We suggest that in the previously studied CP29 complexes from spinach (Pieper et al. Photochem. Photobiol. 2000, 71, 574−589) two Chl’s could have been lost during the preparation/purification procedure, but it is unlikely that the spinach CP29 protein contains only eight Chl’s, as suggested by the sequence homology-based study (Bassi et al. Proc. Natl. Acad. Sci. U.S.A. 1999, 96, 10056−10061). The likely Chl’s missing in wild-type (WT) CP29 complexes studied previously (Pieper et al. Photochem. Photobiol. 2000, 71, 574−589) include a615 and b607. This is why the nonresonant HB spectra shown in that reference were ∼1 nm blue-shifted with the low-energy state mostly localized on about one Chl a (i.e., a612) molecule. Pigment composition of CP29 is discussed in the context of light-harvesting and excitation energy transfer.
1. INTRODUCTION The primary reactions of photosynthesis in higher plants and green algae are initiated by light-harvesting of the antenna systems in photosystem II (PSII). The PSII supercomplex contains a highly efficient light-harvesting system consisting of core and peripheral antennas.1,2 CP47 and CP43 are referred to as core antenna, since they are located just outside of the reaction center (RC) in PSII. These two core complexes are responsible for lightharvesting and transfer the excitation energy from the peripheral antennas to the RC. The peripheral antennas, situated outside of the core antenna, include LHCII (Lhcb1−3),3−5 CP29 (Lhcb4),6 CP26 (Lhcb5), and CP24 (Lhcb6)7 complexes. While the major complex LHCII exists in a trimeric form in Nature, the three minor antenna complexes are usually monomeric. The CP29 complex, of interest to this work, is located between the outmost antenna LHCII and the inner antenna CP47.2 Until recently,6 the consensus was that the CP29 complex contained only eight pigments, which was based on studies of the extracted and purified CP29 (assuming two carotenoid molecules per complex),8−10 which was subsequently compared with the reconstituted proteins.11 However, recent X-ray structural studies of the CP29 protein from spinach6 demonstrated that each CP29 © XXXX American Chemical Society
monomer has 13 chlorophylls (Chl’s), i.e., eight Chl a, four Chl b (and likely one mixed Chl a/Chl b site), and three carotenoids. It appears that the 13 Chl molecules form four major weakly interacting domains of Chl’s: domain 1 contains a602, a603, a608, b609, a/b610, a611, a612, and a615 Chl’s; domains 2 and 2′ each have one Chl, i.e., a613 and b614, respectively; and domain 3 contains three pigments, a604, b606, and b607 Chl’s.6 Domains 2/2′ and 3 are located on the luminal side with the remaining pigments (domain 1) being on the stromal side. The couplings between the cofactors on the stromal and luminal sides, as well as between domains 1, 2/2′, and 3, are weak (see Table SI5 in the Supporting Information). A cutoff energy Vc of 10 cm−1 was used in the definition of domains; i.e., if the coupling between pigments is below Vc, delocalization of exciton states between these pigments is not allowed.12,13 As shown in Figure 1, the four Chl b molecules are located at three different positions, with three of them on the luminal side. Mixed pigment composition ensures absorption of light energy in a broad spectral Received: January 14, 2013 Revised: May 10, 2013
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transfer (EET) processes in CP29. We focus on the following questions: (1) which pigments, if any, were missing in the CP29 complexes previously studied by HB spectroscopy;18 (2) what is the composition of the low-energy excitonic state(s); (3) which pigments contribute to the lowest-energy state and possibly the exit in energy transfer pathways; and (4) which cluster of pigments could be involved in the excess energy quenching.
2. METHODS 2.1. Structural Data and Coupling Constants. Isolation/ purification procedures of CP29 complexes and details of experimental setup to measure low-temperature optical spectra discussed in this work are described in the accompanying paper (part I, DOI 10.1021/jp4004328). The crystal structure of Pan et al. (PDB ID: 3PL9) was used to calculate the coupling matrix elements, Vnm, which were obtained using the TrEsp approach of Madjet et al.23 For example, in the a612−a611− a615 cluster (domain 1), a611 has very strong coupling with both a612 and a615 (86 and 96 cm−1, respectively). In the second trimer, a603−a609−b608, the strongest coupling is between Chl a603 and a609 (95 cm−1). In domain 3 (a604− b606−b607), the coupling matrix elements between a604−b607 and b607−b606 are moderate (i.e., 33 and 30 cm−1, respectively); however, the coupling between a604−b606 is very strong (91 cm−1). An effective dipole moment of 4.3 and 3.6 D was used for all Chl a and Chl b, respectively.24 The Qy transition dipole moments were calculated using the new structure and were assumed to extend from ND to NB. The center of the Chl molecule was taken to be at the averaged position of the four chlorin ring nitrogen atoms, rather than at the central magnesium atom. The coupling constants used in modeling studies described below are summarized in Table SI5 in the Supporting Information. 2.2. Modeling Studies of Optical Spectra. In the present work, optical spectra are modeled using the density matrix description of Renger and Marcus25 in order to more accurately reflect the influences of electron−phonon (el−ph) coupling and lifetime broadening on the calculated optical spectra. A detailed description of this theory has already been presented in refs 23 and 26, and some practical comments on its numerical application and the simulation method used to calculate HB spectra are summarized in the Supporting Information of ref 26. In excitonic calculations, we considered CP29 complexes with different numbers of pigments, including division into four domains. Below, however, only several relevant results are discussed in detail: (1) all 13 Chl’s (as revealed by X-ray structure) are placed in one domain; (2) Chl’s are separated into the four domains identified above; and (3) various combinations of selected pigments are assumed to be lost. A situation where CP29 contains only eight pigments (see Figure 1) as specified by the sequence-homology model8 is briefly discussed in the Supporting Information of this work. We hasten to add that separation into domains of pigments is not critical for the steady-state spectra discussed in this work but should be considered in theoretical descriptions of time-resolved spectra, i.e., while theoretically considering energy transfer dynamics (to be published elsewhere). Nevertheless, in this paper, we focus on case 2 where CP29 is divided into four domains, as partially coherent energy relaxation is expected within the strongly coupled domains (1 and 3), while incoherent Förster-type EET occurs between the low-energy states of each domain. That is, one can assume that, if the couplings, Vnm, within a certain group of pigments are large and the couplings to other pigments in the complex are small (smaller than the differences in site energies
Figure 1. Arrangement of Chl’s in the CP29 complex; the view is along the membrane plane (numbering is adopted from ref 6). Chl’s are represented by the central magnesium atom. Chl a/b molecules are indicated by a starting letter “a”/“b”. The center-to-center distances (in Å) are labeled with black numbers (see text).
range. The strongly interacting Chl a trimer and dimer are a612− a611−a615 and a603−a609, respectively, located on the periphery (and different sides) of the CP29 monomer (see Figure 1). Compared to the previous model,8 a recent X-ray structure revealed five more pigments in CP29, i.e., Chl’s a604, b607, b608, a611, and a615, which are shaded with pink color in Figure 1. The last two Chl’s (a611 and a615) share glycerol-3-phosphate, a phospholipid, as their central ligand and could be lost during the isolation/purification procedure. Note that the CP29 structure6 is very similar to that of the major LHCII antenna complex of PSII,4 which has 14 chromophores and whose pigment labeling is also given in Table A1 (left column). Labeling of CP29 pigments based on ref 8 is shown for comparison (right column). Understanding of CP29 excitonic structure and dynamics is critical, as the CP29 complex plays an important physiological role in the PSII supercomplexes.2 That is, besides the main function of light-harvesting, CP29 stabilizes the whole PSII supercomplex2 and is responsible for transferring energy from LHCII and CP24 to the RC. Therefore, it was suggested that the CP29 complex is involved in regulation of the excitation energy flow; i.e., the CP29 complex is involved in photoprotection.14−16 However, it is not clear which pigment(s) contribute to its lowest energy state(s) and what is its orientation within the PSII supercomplex. On the basis of the LHCII and CP29 protein structures, the likely candidates that contribute strongly to the lowest energy state of CP29 are the a612−a611−a615 trimer6 or the a603− a609 dimer within the a603−a609−b608 cluster,2,6 although localization of the lowest energy state on a604 has been suggested in LHCII17 and the lowest energy state in the previously studied CP29 appeared to be localized on a single, unspecified Chl.18 On the basis of the comparison with LHCII, one could suggest that the a612−a611−a/b610 trimer (similar to the a612−a611−a610 trimer in LHCII19−22) could act as the energy exit in CP29. However, this suggestion has not been previously made, since pigments a611−a615 were absent in the sequencehomology CP29 model.8 In short, further work is required to identify a composition of the lowest energy state which could simultaneously describe various types of optical spectra obtained for the CP29 protein. Below we report various calculated lowtemperature spectra and room-temperature CD to provide more insight into possible excitonic structure and excitation energy B
dx.doi.org/10.1021/jp4004278 | J. Phys. Chem. B XXXX, XXX, XXX−XXX
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where S is the Huang−Rhys factor, ωc = 50 cm−1, and σ = 0.9. In the calculations, the effective S = 0.5 is in agreement with the experimental value obtained by ΔFLNS.29 We note that the spectral density could also be approximated by a typically used half-Gaussian and half-Lorentzian fit (with fwhmG = 20 cm−1 and fwhmL = 120 cm−1)29 or the spectral density of Renger and Marcus,25 which is often used in theoretical calculations.12,30 However, the former cannot be used in Redfield calculations due to continuously increasing reorganization energy as a function of frequency caused by the long tail of the Lorentzian curve (details will be published elsewhere31). The log-normal parameters presented in this work were obtained by fitting the ΔFLN spectrum (λex = 684 nm)29 and not the Gaussian−Lorentzian spectral density, as improper pseudophonon sideband contributions were included in ref 29. Note the good agreement between calculated and experimental data in Figure 2. Calculated spectra of about 1000 combinations of various site energies indicate that the pigments making up the strongly coupled cluster a612−a611−a615−a610 (with site energies 14910 cm −1 [670.7 nm], 15010 cm −1 [666.2 nm], 14960 cm−1 [668.4 nm], and 14970 cm−1 [668.0 nm], respectively) contribute to the lowest energy excitonic state. The discrepancy observed in the Chl b absorption region will be discussed in section 4.1. Figure 3 shows the three lowest energy excitonic states labeled as α = 1, 2, and 3 (averaged over 50 000 complexes). The left and right insets show the averaged occupation number of Chl’s contributing to the first and second lowest energy excitonic states (α = 1 and α = 2), respectively. Frame B shows the distribution functions, dn(ω), of Chl’s mostly contributing to the lowest energy state (i.e., Chl’s a612, a611, a615, and a610) plotted as a function of energy (in cm−1). The lowest energy excitonic state (with a maximum near 14725 cm−1 [679.1 nm]) has a relative oscillator strength of ∼1 Chl a, with largest contributions from a612 (29%), a611 (26%), a615 (11%), and a610 (11%). The second lowest energy state is mostly contributed to by the a603−a609 dimer (see right inset). The delocalization length (inverse participation ratio) of the lowest excitonic state is 2.4. Thus, it is likely that the fourpigment cluster (with a 77% contribution to the lowest energy exciton) constitutes the exit trap in energy-transfer pathways to the CP47 complex. 3.2. Calculated Absorption, Fluorescence, and HB Spectra (Model 2). Below we show calculated data obtained for the CP29 complex (13 Chl’s) divided into four domains (d = 1, 2, 2′, and 3) as defined above. Since the coupling between pigments belonging to different domains is weak (
