Phycobilisomes Research Articles

Absence of the third linker domain of ApcE subunit in phycobilisome from Synechocystis 6803 reduces rods-to-core excitation energy transfer.

Phycobilisome (PBS) is a pigment-protein complex utilized by red algae and cyanobacteria in photosynthesis for light harvesting. A cyanobacterium Synechocystis sp. PCC 6803 contains PBS with a tricylindrical core built of allophycocyanin (APC) disks where six phycocyanin (PC) rods are attached. The top core cylinder is seemingly involved in attaching four PC rods and binding orange carotenoid protein (OCP) to quench excess of excitation energy. In this study, we have deleted the third linker domain (LD3) of ApcE subunit of PBS which assembles four APC discs into the top core cylinder. The mutation resulted in PBS with bicylindrical core, structurally comparable to the naturally existing PBS from Synechococcus 7942. Lack of LD3 and the top APC cylinder reduces the excitation energy transfer between PC and APC in the mutant. Moreover, these PBSs are more prone to light induced-photodamage and do not bind to the photoactivated orange carotenoid protein (OCP), a known PBS excitation quencher. These findings highlight the complex and elegant interplay between PBS architecture and functional efficiency, suggesting that in PBSs with naturally tri-cylindrical cores, the top cylinder has essential roles in recruiting the rods and proper binding of OCP and recruitment of the four PC rods.

N-terminal domain homologs of the orange carotenoid protein increase quenching of cyanobacterial phycobilisomes.

Stress exerted by excess captured light energy in cyanobacteria is prevented by the photoprotective activity of the orange carotenoid protein (OCP). Under high light, the OCP converts from an orange, inactive form (OCPO) into the red form (OCPR) that binds to and quenches the phycobilisome (PBS). Structurally, the OCP consists of two domains: the N-terminal effector domain and a C-terminal regulatory domain. Structural analysis of the OCP-PBS complex showed that the N-terminal domains of an OCP dimer interact with the PBS core. These N-terminal OCP domains have single domain protein paralogs known as Helical Carotenoid Proteins (HCPs). Using phycobilisome quenching assays, we show that the HCP4 and HCP5 homologs efficiently quench PBS fluorescence in vitro, surpassing the quenching ability of the OCP. This is consistent with computational quantum mechanics/molecular mechanics results. Interestingly, when using a maximum quenching concentration of OCP with phycobilisomes, HCP5 addition further increases phycobilisome quenching. Our results provide mechanistic insight into the quenching capacity and roles of HCP4 and HCP5 in cyanobacteria, suggesting that they are more than simply functionally redundant to the OCP.

Energetic Landscape and Terminal Emitters of Phycobilisome Cores from Quantum Chemical Modeling.

Phycobilisomes (PBs) are giant antenna supercomplexes of cyanobacteria that use phycobilin pigments to capture sunlight and transfer the collected energy to membrane-bound photosystems. In the PB core, phycobilins are bound to particular allophycocyanin (APC) proteins. Some phycobilins are thought to be terminal emitters (TEs) with red-shifted fluorescence. However, the precise identification of TEs is still under debate. In this work, we employ multiscale quantum-mechanical calculations to disentangle the excitation energy landscape of PB cores. Using the recent atomistic PB structures from Synechoccoccus PCC 7002 and Synechocystis PCC 6803, we compute the spectral properties of different APC trimers and assign the low-energy pigments. We show that the excitation energy of APC phycobilins is determined by geometric and electrostatic factors and is tuned by the specific protein-protein interactions within the core. Our findings challenge the simple picture of a few red-shifted bilins in the PB core and instead suggest that the red-shifts are established by the entire TE-containing APC trimers. Our work provides a theoretical microscopic basis for the interpretation of energy migration and time-resolved spectroscopy in phycobilisomes.

In situ structural determination of cyanobacterial phycobilisome–PSII supercomplex by STAgSPA strategy

Photosynthesis converting solar energy to chemical energy is one of the most important chemical reactions on earth. In cyanobacteria, light energy is captured by antenna system phycobilisomes (PBSs) and transferred to photosynthetic reaction centers of photosystem II (PSII) and photosystem I (PSI). While most of the protein complexes involved in photosynthesis have been characterized by in vitro structural analyses, how these protein complexes function together in vivo is not well understood. Here we implemented STAgSPA, an in situ structural analysis strategy, to solve the native structure of PBS–PSII supercomplex from the cyanobacteria Arthrospira sp. FACHB439 at resolution of ~3.5 Å. The structure reveals coupling details among adjacent PBSs and PSII dimers, and the collaborative energy transfer mechanism mediated by multiple super-PBS in cyanobacteria. Our results provide insights into the diversity of photosynthesis-related systems between prokaryotic cyanobacteria and eukaryotic red algae but are also a methodological demonstration for high-resolution structural analysis in cellular or tissue samples.

Functional Modification of Cyanobacterial Phycobiliprotein and Phycobilisomes through Bilin Metabolism Control.

Phycobilisomes (PBSs) are light-harvesting antenna complexes in cyanobacteria that adapt to diverse light environments through the use of phycobiliproteins within the PBS structures. Freshwater cyanobacteria, such as Synechococcus elongatus PCC 7942, thrive under red light because of the presence of phycocyanin (PC) and its chromophore, phycocyanobilin (PCB), in the PBS. Cyanobacteria in shorter-wavelength light environments such as green light, employ phycoerythrin paired with phycoerythrobilin (PEB) along with PC in the PBS. Synthetic biology studies have shown that PEB production can be achieved by expression of the heterologous PEB synthases 15,16-dihydrobiliverdin:ferredoxin oxidoreductase (PebA) and PEB:ferredoxin oxidoreductase (PebB), leading to PEB accumulation and cellular browning. This approach is genetically unstable, and the properties of the resulting PEB-bound PBS complexes remain uncharacterized. In this study, we engineered a novel strain of Synechococcus 7942 PEB1 with finely tuned control of PEB biosynthesis. PEB1 exhibited a reversible change in the color of the culture from green to brown and pink based on PebA and PebB induction levels. High induction led to complete PCB-to-PEB substitution, causing the disassembly of the PBS rod complex. In contrast, low induction levels of PebA and PebB resulted in the formation of a stable chimeric PBS complex with partial PCB-to-PEB substitution. This acclimation enabled efficient light harvesting in the green spectrum and energy transfer to the photosynthetic reaction center. These findings, which improve our understanding of PBS and highlight the structural importance of the bilin composition, provide a foundation for future studies on PBS adaptation in bioengineering, synthetic biology, and renewable energy.

Composition Heterogeneity of Metal Ions Bound at the Oxygen-Evolving Center of Photosystem II in Living Cells.

Recent resolution advancement of in situ cryo-electron tomography (cryo-ET) and cryo-electron microscopy (cryo-EM) enables us to visualize large enzymes-in-action in atomic detail in their native environments inside living cells, such as photosystem II (PSII) and the ribosome. A variety of crystallographic and cryo-EM structures of PSII have been published for the purified PSII dimeric core complexes by itself, in supercomplexes with photosystem I (PSI) and light-harvesting complexes (LHC), and in megacomplexes with phycobilisome (PBS). In the latter case, two or five copies of asymmetric dimeric PSII molecules are present in highly asymmetric environments that differ from other 2-fold symmetric structures. Previous systematic analysis of X-ray free-electron laser (XFEL) crystal structures of PSII has shown different degrees of composition heterogeneity of metal ion cofactor bound at the oxygen-evolving center (OEC), including between two monomers of the same PSII dimer. This study analyzed the metal ions bound at four OECs in two asymmetric dimeric PSII molecules within in situ cryo-ET structures reported for an asymmetric PBS-PSII-PSI-LHC megacomplex determined in a living organism without purification and shows that composition heterogeneity with reduced metal ion occupancies at the OEC of PSII is a general phenomenon. This finding could have profound implications for spectroscopic interpretations of unpurified PSII samples.

Adaptation processes in Halomicronema hongdechloris, an example of the light-induced optimization of the photosynthetic apparatus on hierarchical time scales.

Oxygenic photosynthesis in Halomicronema hongdechloris, one of a series of cyanobacteria producing red-shifted Chl f, is adapted to varying light conditions by a range of diverse processes acting over largely different time scales. Acclimation to far-red light (FRL) above 700 nm over several days is mirrored by reversible changes in the Chl f content. In several cyanobacteria that undergo FRL photoacclimation, Chl d and Chl f are directly involved in excitation energy transfer in the antenna system, form the primary donor in photosystem I (PSI), and are also involved in electron transfer within photosystem II (PSII), most probably at the ChlD1 position, with efficient charge transfer happening with comparable kinetics to reaction centers containing Chl a. In H. hongdechloris, the formation of Chl f under FRL comes along with slow adaptive proteomic shifts like the rebuilding of the D1 complex on the time scale of days. On shorter time scales, much faster adaptation mechanisms exist involving the phycobilisomes (PBSs), which mainly contain allophycocyanin upon adaptation to FRL. Short illumination with white, blue, or red light leads to reactive oxygen species-driven mobilization of the PBSs on the time scale of seconds, in effect recoupling the PBSs with Chl f-containing PSII to re-establish efficient excitation energy transfer within minutes. In summary, H. hongdechloris reorganizes PSII to act as a molecular heat pump lifting excited states from Chl f to Chl a on the picosecond time scale in combination with a light-driven PBS reorganization acting on the time scale of seconds to minutes depending on the actual light conditions. Thus, structure-function relationships in photosynthetic energy and electron transport in H. hongdechloris including long-term adaptation processes cover 10-12 to 106 seconds, i.e., 18 orders of magnitude in time.

Altered excitation energy transfer between phycobilisome and photosystems in the absence of ApcG, a small linker peptide, in Synechocystis sp. PCC 6803, a cyanobacterium

Phycobilisome (PBS) is a large pigment-protein complex in cyanobacteria and red algae responsible for capturing sunlight and transferring its energy to photosystems (PS). Spectroscopic and structural properties of various PBSs have been widely studied, however, the nature of so-called complex-complex interactions between PBS and PSs remains much less explored. In this work, we have investigated the function of a newly identified PBS linker protein, ApcG, some domain of which, together with a loop region (PB-loop in ApcE), is possibly located near the PBS-PS interface. Using Synechocystis sp. PCC 6803, we generated an ApcG deletion mutant and probed its deletion effect on the energetic coupling between PBS and photosystems. Steady-state and time-resolved spectroscopic characterization of the purified ΔApcG-PBS demonstrated that ApcG removal weakly affects the photophysical properties of PBS for which the spectroscopic properties of terminal energy emitters are comparable to those of PBS from wild-type strain. However, analysis of fluorescence decay imaging datasets reveals that ApcG deletion induces disruptions within the allophycocyanin (APC) core, resulting in the emergence (splitting) of two spectrally diverse subgroups with some short-lived APC. Profound spectroscopic changes of the whole ΔApcG mutant cell, however, emerge during state transition, a dynamic process of light scheme adaptation. The mutant cells in State I show a substantial increase in PBS-related fluorescence. On the other hand, global analysis of time-resolved fluorescence demonstrates that in general ApcG deletion does not alter or inhibit state transitions interpreted in terms of the changes of the PSII and PSI fluorescence emission intensity. The results revealed yet–to–be discovered mechanism of ApcG-docking induced excitation energy transfer regulation within PBS or to Photosystems.

The structural basis for light harvesting in organisms producing phycobiliproteins.

Cyanobacteria, red algae, and cryptophytes produce 2 classes of proteins for light harvesting: water-soluble phycobiliproteins (PBP) and membrane-intrinsic proteins that bind chlorophylls (Chls) and carotenoids. In cyanobacteria, red algae, and glaucophytes, phycobilisomes (PBS) are complexes of brightly colored PBP and linker (assembly) proteins. To date, 6 structural classes of PBS have been described: hemiellipsoidal, block-shaped, hemidiscoidal, bundle-shaped, paddle-shaped, and far-red-light bicylindrical. Two additional antenna complexes containing single types of PBP have also been described. Since 2017, structures have been reported for examples of all of these complexes except bundle-shaped PBS by cryogenic electron microscopy. PBS range in size from about 4.6 to 18 mDa and can include ∼900 polypeptides and bind >2000 chromophores. Cyanobacteria additionally produce membrane-associated proteins of the PsbC/CP43 superfamily of Chl a/b/d-binding proteins, including the iron-stress protein IsiA and other paralogous Chl-binding proteins (CBP) that can form antenna complexes with Photosystem I (PSI) and/or Photosystem II (PSII). Red and cryptophyte algae also produce CBP associated with PSI but which belong to the Chl a/b-binding protein superfamily and which are unrelated to the CBP of cyanobacteria. This review describes recent progress in structure determination for PBS and the Chl proteins of cyanobacteria, red algae, and cryptophytan algae.

Structural and quantum chemical basis for OCP-mediated quenching of phycobilisomes.

Cyanobacteria use large antenna complexes called phycobilisomes (PBSs) for light harvesting. However, intense light triggers non-photochemical quenching, where the orange carotenoid protein (OCP) binds to PBS, dissipating excess energy as heat. The mechanism of efficiently transferring energy from phycocyanobilins in PBS to canthaxanthin in OCP remains insufficiently understood. Using cryo-electron microscopy, we unveiled the OCP-PBS complex structure at 1.6- to 2.1-angstrom resolution, showcasing its inherent flexibility. Using multiscale quantum chemistry, we disclosed the quenching mechanism. Identifying key protein residues, we clarified how canthaxanthin's transition dipole moment in its lowest-energy dark state becomes large enough for efficient energy transfer from phycocyanobilins. Our energy transfer model offers a detailed understanding of the atomic determinants of light harvesting regulation and antenna architecture in cyanobacteria.

Insights into energy quenching mechanisms and carotenoid uptake by orange carotenoid protein homologs: HCP4 and CTDH

Photodamage to the photosynthetic apparatus by excessive light radiation has led to the evolution of a variety of energy dissipation mechanisms. A mechanism that exists in some cyanobacterial species, enables non-photochemical quenching of excitation energy within the phycobilisome (PBS) antenna complex by the Orange Carotenoid Protein (OCP). The OCP contains an active N-terminal domain (NTD) and a regulatory C-terminal domain (CTD). Some cyanobacteria also have genes encoding for homologs to both the CTD (CTDH) and the NTD (referred to as helical carotenoid proteins, HCP). The CTDH facilitates uptake of carotenoids from the thylakoid membranes to be transferred to the HCPs. Holo-HCPs exhibit diverse functionalities such as carotenoid carriers, singlet oxygen quenchers, and in the case of HCP4, constitutive OCP-like energy quenching. Here, we present the first crystal structure of the holo-HCP4 binding canthaxanthin molecule and an improved structure of the apo-CTDH from Anabaena sp. PCC 7120. We propose here models of the binding of the HCP4 to the PBS and the associated energy quenching mechanism. Our results show that the presence of the carotenoid is essential for fluorescence quenching. We also examined interactions within OCP-like species, including HCP4 and CTDH, providing the basis for mechanisms of carotenoid transfer from CTDH to HCPs.

ApcE plays an important role in light-induced excitation energy dissipation in the Synechocystis PCC6803 phycobilisomes

Phycobilisomes (PBs) play an important role in cyanobacterial photosynthesis. They capture light and transfer excitation energy to the photosynthetic reaction centres. PBs are also central to some photoprotective and photoregulatory mechanisms that help sustain photosynthesis under non-optimal conditions. Amongst the mechanisms involved in excitation energy dissipation that are activated in response to excessive illumination is a recently discovered light-induced mechanism that is intrinsic to PBs and has been the least studied. Here, we used single-molecule spectroscopy and developed robust data analysis methods to explore the role of a terminal emitter subunit, ApcE, in this intrinsic, light-induced mechanism. We isolated the PBs from WT Synechocystis PCC 6803 as well as from the ApcE-C190S mutant of this strain and compared the dynamics of their fluorescence emission. PBs isolated from the mutant (i.e., ApcE-C190S-PBs), despite not binding some of the red-shifted pigments in the complex, showed similar global emission dynamics to WT-PBs. However, a detailed analysis of dynamics in the core revealed that the ApcE-C190S-PBs are less likely than WT-PBs to enter quenched states under illumination but still fully capable of doing so. This result points to an important but not exclusive role of the ApcE pigments in the light-induced intrinsic excitation energy dissipation mechanism in PBs.

Assembly of CpcL-phycobilisomes.

CpcL-phycobilisomes (CpcL-PBSs) are a reduced type of phycobilisome (PBS) found in several cyanobacteria. They lack the traditional PBS terminal energy emitters, but still show the characteristic red-shifted fluorescence at ~670 nm. We established a method of assembling in vitro a rod-membrane linker protein, CpcL, with phycocyanin, generating complexes with the red-shifted spectral features of CpcL-PBSs. The red-shift arises from the interaction of a conserved key glutamine, Q57 of CpcL in Synechocystis sp. PCC 6803, with a single phycocyanobilin chromophore of trimeric phycocyanin at one of the three β82-sites. This chromophore is the terminal energy acceptor of CpcL-PBSs and donor to the photosystem(s). This mechanism also operates in PBSs from Acaryochloris marina MBIC11017. We then generated multichromic complexes harvesting light over nearly the complete visible range via the replacement of phycocyanobilin chromophores at sites α84 and β153 of phycocyanins by phycoerythrobilin and/or phycourobilin. The results demonstrate the rational design of biliprotein-based light-harvesting elements by engineering CpcL and phycocyanins, which broadens the light-harvesting range and accordingly improves the light-harvesting capacity and may be potentially applied in solar energy harvesting.

Expanding the toolbox for phycobiliprotein assembly: phycoerythrobilin biosynthesis in Synechocystis

AbstractPhycobiliproteins (PBPs) play a vital role in light harvesting by cyanobacteria, which enables efficient utilization of photon energy for oxygenic photosynthesis. The PBPs carry phycobilins, open‐chain tetrapyrrole chromophores derived from heme. The structure and chromophore composition of PBPs is dependent on the organism's ecological niche. In cyanobacteria, these holo‐proteins typically form large, macromolecular antenna complexes called phycobilisomes (PBSs). The PBS of Synechocystis sp. PCC 6803 (hereafter Synechocystis) consists of allophycocyanin (APC) and phycocyanin (PC), which exclusively harbor phycocyanobilin (PCB) as a chromophore. Investigations into heterologous PBP biosynthesis in E. coli have proven limiting with respect to PBP assembly and their functional characterization. Consequently, we wanted to engineer a platform for the investigation of heterologously produced PBPs, focusing on unusual, phycoerythrobilin (PEB)‐containing light‐harvesting proteins called phycoerythrins (PEs) in Synechocystis. As a first step, a gene encoding for the synthesis of the natural cyanobacterial chromophore, PEB, was introduced into Synechocystis. We provide spectroscopic evidence for heterologous PEB formation and show covalent attachment of PEB to the α‐subunit of PC, CpcA, by HPLC and LC–MS/MS analyses. Fluorescence microscopy and PBS isolation demonstrate a cellular dispersal of PBPs with modified phycobilin content. However, these modifications have minor effects on physiological responses, as demonstrated by growth rates, oxygen evolution, nutrient accumulation, and PBP content analyses. As a result, Synechocystis demonstrates the capacity to efficiently manage PEB biosynthesis and therefore reflects a promising platform for both biochemical and physiological investigations of foreign and unusual PEs.

A structure of the relict phycobilisome from a thylakoid-free cyanobacterium

Phycobilisomes (PBS) are antenna megacomplexes that transfer energy to photosystems II and I in thylakoids. PBS likely evolved from a basic, inefficient form into the predominant hemidiscoidal shape with radiating peripheral rods. However, it has been challenging to test this hypothesis because ancestral species are generally inaccessible. Here we use spectroscopy and cryo-electron microscopy to reveal a structure of a “paddle-shaped” PBS from a thylakoid-free cyanobacterium that likely retains ancestral traits. This PBS lacks rods and specialized ApcD and ApcF subunits, indicating relict characteristics. Other features include linkers connecting two chains of five phycocyanin hexamers (CpcN) and two core subdomains (ApcH), resulting in a paddle-shaped configuration. Energy transfer calculations demonstrate that chains are less efficient than rods. These features may nevertheless have increased light absorption by elongating PBS before multilayered thylakoids with hemidiscoidal PBS evolved. Our results provide insights into the evolution and diversification of light-harvesting strategies before the origin of thylakoids.

The structural basis for light acclimation in phycobilisome light harvesting systems systems in Porphyridium purpureum

Photosynthetic organisms adapt to changing light conditions by manipulating their light harvesting complexes. Biophysical, biochemical, physiological and genetic aspects of these processes are studied extensively. The structural basis for these studies is lacking. In this study we address this gap in knowledge by focusing on phycobilisomes (PBS), which are large structures found in cyanobacteria and red algae. In this study we focus on the phycobilisomes (PBS), which are large structures found in cyanobacteria and red algae. Specifically, we examine red algae (Porphyridium purpureum) grown under a low light intensity (LL) and a medium light intensity (ML). Using cryo-electron microscopy, we resolve the structure of ML-PBS and compare it to the LL-PBS structure. The ML-PBS is 13.6 MDa, while the LL-PBS is larger (14.7 MDa). The LL-PBS structure have a higher number of closely coupled chromophore pairs, potentially the source of the red shifted fluorescence emission from LL-PBS. Interestingly, these differences do not significantly affect fluorescence kinetics parameters. This indicates that PBS systems can maintain similar fluorescence quantum yields despite an increase in LL-PBS chromophore numbers. These findings provide a structural basis to the processes by which photosynthetic organisms adapt to changing light conditions.

Responding to light signals: a comprehensive update on photomorphogenesis in cyanobacteria.

Cyanobacteria are ancestors of chloroplast and perform oxygen-evolving photosynthesis similar to higher plants and algae. However, an obligatory requirement of photons for their growth results in the exposure of cyanobacteria to varying light conditions. Therefore, the light environment could act as a signal to drive the developmental processes, in addition to photosynthesis, in cyanobacteria. These Gram-negative prokaryotes exhibit characteristic light-dependent developmental processes that maximize their fitness and resource utilization. The development occurring in response to radiance (photomorphogenesis) involves fine-tuning cellular physiology, morphology and metabolism. The best-studied example of cyanobacterial photomorphogenesis is chromatic acclimation (CA), which allows a selected number of cyanobacteria to tailor their light-harvesting antenna called phycobilisome (PBS). The tailoring of PBS under existing wavelengths and abundance of light gives an advantage to cyanobacteria over another photoautotroph. In this work, we will provide a comprehensive update on light-sensing, molecular signaling and signal cascades found in cyanobacteria. We also include recent developments made in other aspects of CA, such as mechanistic insights into changes in the size and shape of cells, filaments and carboxysomes.

Phycobilisome protein ApcG interacts with PSII and regulates energy transfer in Synechocystis.

Photosynthetic organisms harvest light using pigment-protein complexes. In cyanobacteria, these are water-soluble antennae known as phycobilisomes (PBSs). The light absorbed by PBS is transferred to the photosystems in the thylakoid membrane to drive photosynthesis. The energy transfer between these complexes implies that protein-protein interactions allow the association of PBS with the photosystems. However, the specific proteins involved in the interaction of PBS with the photosystems are not fully characterized. Here, we show in Synechocystis sp. PCC 6803 that the recently discovered PBS linker protein ApcG (sll1873) interacts specifically with PSII through its N-terminal region. Growth of cyanobacteria is impaired in apcG deletion strains under light-limiting conditions. Furthermore, complementation of these strains using a phospho-mimicking version of ApcG causes reduced growth under normal growth conditions. Interestingly, the interaction of ApcG with PSII is affected when a phospho-mimicking version of ApcG is used, targeting the positively charged residues interacting with the thylakoid membrane, suggesting a regulatory role mediated by phosphorylation of ApcG. Low-temperature fluorescence measurements showed decreased PSI fluorescence in apcG deletion and complementation strains. The PSI fluorescence was the lowest in the phospho-mimicking complementation strain, while the pull-down experiment showed no interaction of ApcG with PSI under any tested condition. Our results highlight the importance of ApcG for selectively directing energy harvested by the PBS and imply that the phosphorylation status of ApcG plays a role in regulating energy transfer from PSII to PSI.

Anti-stokes fluorescence of phycobilisome and its complex with the orange carotenoid protein

Phycobilisomes (PBSs) are giant water-soluble light-harvesting complexes of cyanobacteria and red algae, consisting of hundreds of phycobiliproteins precisely organized to deliver the energy of absorbed light to chlorophyll chromophores of the photosynthetic electron-transport chain. Quenching the excess of excitation energy is necessary for the photoprotection of photosynthetic apparatus. In cyanobacteria, quenching of PBS excitation is provided by the Orange Carotenoid Protein (OCP), which is activated under high light conditions. In this work, we describe parameters of anti-Stokes fluorescence of cyanobacterial PBSs in quenched and unquenched states. We compare the fluorescence readout from entire phycobilisomes and their fragments. The obtained results revealed the heterogeneity of conformations of chromophores in isolated phycobiliproteins, while such heterogeneity was not observed in the entire PBS. Under excitation by low-energy quanta, we did not detect a significant uphill energy transfer from the core to the peripheral rods of PBS, while the one from the terminal emitters to the bulk allophycocyanin chromophores is highly probable. We show that this direction of energy migration does not eliminate fluorescence quenching in the complex with OCP. Thus, long-wave excitation provides new insights into the pathways of energy conversion in the phycobilisome.

Cryo-EM and femtosecond spectroscopic studies provide mechanistic insight into the energy transfer in CpcL-phycobilisomes

Phycobilisomes (PBS) are the major light harvesting complexes of photosynthesis in the cyanobacteria and red algae. CpcL-PBS is a type of small PBS in cyanobacteria that transfers energy directly to photosystem I without the core structure. Here we report the cryo-EM structure of the CpcL-PBS from the cyanobacterium Synechocystis sp. PCC 6803 at 2.6-Å resolution. The structure shows the CpcD domain of ferredoxin: NADP+ oxidoreductase is located at the distal end of CpcL-PBS, responsible for its attachment to PBS. With the evidence of ultrafast transient absorption and fluorescence spectroscopy, the roles of individual bilins in energy transfer are revealed. The bilin 1Iβ822 located near photosystem I has an enhanced planarity and is the red-bilin responsible for the direct energy transfer to photosystem I.