Excitation-Emission Correlation Dissects Energy Transfer Pathway Diversity of Photosystem I

Light-driven photosynthetic reactions occur in large pigment-protein complexes. The robust excitation energy transfer (EET) established in these complexes is a key to maintaining the high-efficiency electron transport of light reactions in photosynthesis. The structural variability of the protein dynamically alters the EET pathways, a mechanism that is considered to be potentially beneficial for photoprotection. However, the diversity of EET pathways and their modulation by conformational changes of the protein have been unresolved. Here, using cryogenic single-molecule excitation-emission spectroscopy (SMEES), we study EET in photosystem I (PSI) monomers and trimers, by freezing the conformations and resulting EET pathways, and investigating each of them in a different single molecule. Monomeric PSI binds 96 chlorophylls (Chls) in which a few low-excitation-energy Chls act as terminal emitter domains (TEDs) of fluorescence at low temperatures. SMEES data reflects the EET pathways from upstream Chls to the TEDs and to the oxidized reaction center (RC⁺) in PSI by characterizing excitation-emission correlations. The peak heterogeneity of the excitation spectrum characterizes the complex energy landscape and the resulting EET pathways. A correlation between excitation and emission wavelengths indicates a spatio-energetic correlation of excitation energies of pigments connected asymmetrically to different TEDs. For PSI in aqueous environment such a correlation is not found, proving the diversity and robustness of EET. In contrast, embedding PSI in poly(vinyl alcohol) (PVA) reveals correlations. According to structure-based simulations, the observed correlations can be understood as a red-shift of bulk exciton states located near the low-energy TED Low2, relative to those in the vicinity of the high-energy TED Low1, indicating that PVA strongly alters the EET networks by arresting the dynamics of the antenna system such that certain realizations of site energies are selected. The simulations predict that, at 300 K, the thermal energy is sufficient to overcome the restrictions in EET caused by the PVA environment. In addition, we find that the characteristics of the quenching of excitation energy by RC⁺ is similar at 300 K and at 77 K, providing further evidence for the photoprotective role of this process. The present study proves the SMEES technique to be a versatile tool for deciphering EET in complex systems. These findings have deepened our understanding of the EET network within PSI in particular and pigment-protein complexes in general, providing new insights into the structure-function interplay.