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Self-assembly and energy transfer in artificial light-harvesting complexes of bacteriochlorophyll c with astaxanthin


Chlorosomes, the light-harvesting antennae of green photosynthetic bacteria, are based on large aggregates of bacteriochlorophyll molecules. Aggregates with similar properties to those in chlorosomes can also be prepared in vitro. Several agents were shown to induce aggregation of bacteriochlorophyll c in aqueous environments, including certain lipids, carotenes, and quinones. A key distinguishing feature of bacteriochlorophyll c aggregates, both in vitro and in chlorosomes, is a large (>60 nm) red shift of their Qy absorption band compared with that of the monomers. In this study, we investigate the self-assembly of bacteriochlorophyll c with the xanthophyll astaxanthin, which leads to the formation of a new type of complexes. Our results indicate that, due to its specific structure, astaxanthin molecules competes with bacteriochlorophylls for the bonds involved in the aggregation, thus preventing the formation of any significant red shift compared with pure bacteriochlorophyll c in aqueous buffer. A strong interaction between both the types of pigments in the developed assemblies, is manifested by a rather efficient (~40%) excitation energy transfer from astaxanthin to bacteriochlorophyll c, as revealed by fluorescence excitation spectroscopy. Results of transient absorption spectroscopy show that the energy transfer is very fast (<500 fs) and proceeds through the S2 state of astaxanthin.

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This study was supported by the Czech Ministry of Education, Youth and Sports (projects MSM0021620835, MSM6007665808, AV0Z50510513), Czech Science Foundation (projects 206/09/0375, 202/09/H041, 202/09/1330), and Spanish Ministry of Science and Innovation (AVCR-CSIC joint project 2008CZ0004). The authors would like to thank Ivana Hunalova, Frantisek Matousek, and Anita Zupcanova for their help with pigment isolation.

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Correspondence to J. Pšenčík.

Electronic supplementary material

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Figure S1: Correction of the absorption spectra of BChl c-astaxanthin assemblies for light-scattering. Spectra were measured at a position further (solid line) and closer (dotted line) to the detector of the spectrophotometer. The difference between them were rescaled to fit the uncorrected spectra above 850 nm which reflected the scattering part of the spectra (dashed line). The corrected spectrum (dash-dot line) was obtained by subtracting the scattering part from the solid line. (PDF 21 kb)

Figure S2: Transient absorption spectra of BChl c assemblies with astaxanthin (molar ratio 0.35:1 astaxanthin-to-BChl c, blue lines) and BChl c dimers with lecithin (green lines). All spectra were measured at 2.1 ps after selective excitation of BChl c at 710 nm. The inset shows an enlargement of the 450-600 nm region. The inverted absorption spectrum of astaxanthin in the assemblies, reproduced from Fig. 3 and scaled to fit, is shown for the sake of comparison (black line, inset). The difference spectrum (red line) reveals a shift of the Qy band of BChl c and a small negative signal in the 450-600 nm region. This small signal can be caused entirely by a change in the BChl–BChl excitonic interaction between the two samples, or by an additional excitonic interaction between BChl c and astaxanthin resulting in bleaching of the astaxanthin upon BChl c excitation. The spectra were smoothed by a 10 point (± 2.5 nm) adjacent averaging. (PDF 28 kb)

Appendix: Transient data analysis

Appendix: Transient data analysis

We assume simple kinetic model for excited state population N i

$$ \dot{N}_{i} (t) = \sum\limits_{i} {\left( { - k_{ij} N_{i} (t) + k_{ji} N_{j} (t)} \right)} $$

where k ij is the rate constant for energy/population transfer from one state i to the another state j and dot represents derivative with respect to time. By solving set of Eq. 1 (one for each state), and considering that each state contributes to transient signal proportionally to its population, we get the following model for describing transient absorption data

$$ \Updelta A(t) = \sum\limits_{j} {\left( {\sum\limits_{i} {K_{ji} \varepsilon_{i} N_{i} } (0)} \right)\exp ( - k_{j} t)} $$

where indices i and j iterates over all energy states present in the sample, k j is the overall rate constant of the state j (\( k_{j} = \sum\limits_{i} {k_{ij} } \)), ε is a spectral profile associated with the given state, and coefficients K ij describe the energy transfer between states, typically \( K_{ij} = \frac{{k_{ij} }}{{k_{j} - k_{i} }} \) or product of similar terms. We can then perform global fitting analysis to find the overall rate constants and corresponding pre-exponential factors (i.e. the DAS)

$$ \Updelta A(t) = \sum\limits_{j} {DAS_{j} \exp ( - k_{j} t)} $$

Let’s assume in accordance with the multi-exponential model that relaxation pathways from an excited state are the same regardless of its population. This is of course an approximation, because relaxation within the BChl c manifold is likely to be influenced by singlet–singlet annihilation, which the model does not describe. It follows that the transient signal consists of parts proportional to the initial population of states directly excited by pump pulse (the other way of summation in Eq. 2)

$$ \Updelta A(t) = \sum\limits_{i} {\left( {\varepsilon_{i} \sum\limits_{j} {K_{ji} \exp ( - k_{j} t)} } \right)N_{i} (0)} = \sum\limits_{i} {\Updelta a_{i} N_{i} (0)} $$

If we perform at least P measurements with different and known ratio of the initial population of the excited states, where P is the number of the initially excited states (with non-zero N i (0)), then we can determine the parts of the transient data corresponding to the relaxation from each of the initially excited states (Δa i in Eq. 4). Note that these parts should be the same in all measurements, except that their weights N i (0) differ. Here we assume that only the S2 state of astaxanthin and Soret band of BChl c are directly excited, and that the transient data separates into two parts: relaxation following excitation of astaxanthin, and BChl c, respectively. Therefore, we need to perform two transient absorption measurements with a different ratio between initial populations of the S2 state of astaxanthin and the Soret band of BChl c. One theoretically possible way to change the excited state population is to use one excitation wavelength and two samples with different astaxanthin-to-BChl c molar ratios. However, different sample composition in this case results in a different way of assembly, and therefore, in different absorption spectra, breaking the condition that relaxation from excited state is the same. The other way is to use one sample and two different excitation wavelengths (close enough to excite the same electronic transitions and far enough to achieve significant change of excited populations ratio). In this case, we populate different parts from the inhomogeneously broadened electronic transition peaks; however, this should not influence the relaxation from the higher excited state significantly. We measured the transient data after excitation at two different wavelengths, 450 and 490 nm. Therefore, we can rewrite Eq. 4 describing the global decay at two excitation wavelength as

$$ \begin{gathered} \Updelta A_{450} = \Updelta a_{\text{BChl}} N_{450}^{\text{BChl}} (0) + \Updelta a_{\text{Car}} N_{450}^{\text{Car}} (0) \hfill \\ \Updelta A_{490} = \Updelta a_{\text{BChl}} N_{490}^{\text{BChl}} (0) + \Updelta a_{\text{Car}} N_{490}^{\text{Car}} (0) \hfill \\ \end{gathered} $$

where ΔA 450 and ΔA 490 represent transient data after excitation at 450 and 490 nm, respectively.

The initial excited state populations N λ molecule (0) could be determined directly from steady-state absorption spectra corrected for scattering, and energy in the pump pulse. From the two equations, two unknown matrices Δa BChl and Δa Car, which represent BChl c and astaxanthin part of the transient data, could be calculated (Fig. 6).

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Alster, J., Polívka, T., Arellano, J.B. et al. Self-assembly and energy transfer in artificial light-harvesting complexes of bacteriochlorophyll c with astaxanthin. Photosynth Res 111, 193–204 (2012).

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  • Light-harvesting
  • Chlorosomes
  • Self-assembly
  • Bacteriochlorophyll aggregates
  • Astaxanthin