Energetic robustness to large scale structural fluctuations in a photosynthetic supercomplex.

Photosynthetic organisms transport and convert solar energy with near-unity quantum efficiency using large protein supercomplexes held in flexible membranes. The individual proteins position chlorophylls to tight tolerances considered critical for fast and efficient energy transfer. The variability in protein organization within the supercomplexes, and how efficiency is maintained despite variability, had been unresolved. Here, we report on structural heterogeneity in the 2-MDa cyanobacterial PSI-IsiA photosynthetic supercomplex observed using Cryo-EM, revealing large-scale variances in the positions of IsiA relative to PSI. Single-molecule measurements found efficient IsiA-to-PSI energy transfer across all conformations, along with signatures of transiently decoupled IsiA. Structure based calculations showed that rapid IsiA-to-PSI energy transfer is always maintained, and even increases by three-fold in rare conformations via IsiA-specific chls. We postulate that antennae design mitigates structural fluctuations, providing a mechanism for robust energy transfer in the flexible membrane.

Elucidating interprotein energy transfer dynamics within the antenna network from purple bacteria.

In photosynthesis, absorbed light energy transfers through a network of antenna proteins with near-unity quantum efficiency to reach the reaction center, which initiates the downstream biochemical reactions. While the energy transfer dynamics within individual antenna proteins have been extensively studied over the past decades, the dynamics between the proteins are poorly understood due to the heterogeneous organization of the network. Previously reported timescales averaged over such heterogeneity, obscuring individual interprotein energy transfer steps. Here, we isolated and interrogated interprotein energy transfer by embedding two variants of the primary antenna protein from purple bacteria, light-harvesting complex 2 (LH2), together into a near-native membrane disc, known as a nanodisc. We integrated ultrafast transient absorption spectroscopy, quantum dynamics simulations, and cryogenic electron microscopy to determine interprotein energy transfer timescales. By varying the diameter of the nanodiscs, we replicated a range of distances between the proteins. The closest distance possible between neighboring LH2, which is the most common in native membranes, is 25 Šand resulted in a timescale of 5.7 ps. Larger distances of 28 to 31 Šresulted in timescales of 10 to 14 ps. Corresponding simulations showed that the fast energy transfer steps between closely spaced LH2 increase transport distances by ∼15%. Overall, our results introduce a framework for well-controlled studies of interprotein energy transfer dynamics and suggest that protein pairs serve as the primary pathway for the efficient transport of solar energy.

Frequency-Domain Spectroscopic Study of the Photosystem I Supercomplexes, Isolated IsiA Monomers, and the Intact IsiA Ring.

The PSI3-IsiA18 supercomplex is one of the largest and most complicated assemblies in photosynthesis. The IsiA ring, composed of 18 IsiA monomers (IsiA18) surrounding the PSI trimer (PSI3), forms under iron-deficient conditions in cyanobacteria and acts as a peripheral antenna. Based on the supercomplex structure recently determined via cryo-EM imaging, we model various optical spectra of the IsiA monomers and IsiA18 ring. Comparison of the absorption and emission spectra of the isolated IsiA monomers and the full ring reveals that about 2.7 chlorophylls (Chls) are lost in the isolated IsiA monomers. The best fits for isolated monomers spectra are obtained assuming the absence of Chl 508 and Chl 517 and 70% loss of Chl 511. The best model describing all three hexamers and the entire ring suggests that the lowest energy pigments are Chls 511, 514, and 517. Based on the modeling results presented in this work, we conclude that there are most likely three entry points for EET from the IsiA6 hexamer to the PSI core monomer, with two of these entry points likely being located next to each other (i.e., nine entry points from IsiA18 to the PSI3 trimer). Finally, we show that excitation energy transfer inside individual monomers is fast (<2 ps at T = 5 K) and at least 20 times faster than intermonomer energy transfer.

The structure of photosystem I from a high-light-tolerant cyanobacteria.

Photosynthetic organisms have adapted to survive a myriad of extreme environments from the earth's deserts to its poles, yet the proteins that carry out the light reactions of photosynthesis are highly conserved from the cyanobacteria to modern day crops. To investigate adaptations of the photosynthetic machinery in cyanobacteria to excessive light stress, we isolated a new strain of cyanobacteria, Cyanobacterium aponinum 0216, from the extreme light environment of the Sonoran Desert. Here we report the biochemical characterization and the 2.7 Å resolution structure of trimeric photosystem I from this high-light-tolerant cyanobacterium. The structure shows a new conformation of the PsaL C-terminus that supports trimer formation of cyanobacterial photosystem I. The spectroscopic analysis of this photosystem I revealed a decrease in far-red absorption, which is attributed to a decrease in the number of long- wavelength chlorophylls. Using these findings, we constructed two chimeric PSIs in Synechocystis sp. PCC 6803 demonstrating how unique structural features in photosynthetic complexes can change spectroscopic properties, allowing organisms to thrive under different environmental stresses.

Author Correction: The structure of a red-shifted photosystem I reveals a red site in the core antenna.

A Correction to this paper has been published: https://doi.org/10.1038/s41467-020-19953-w.

The structure of a red-shifted photosystem I reveals a red site in the core antenna.

Photosystem I coordinates more than 90 chlorophylls in its core antenna while achieving near perfect quantum efficiency. Low energy chlorophylls (also known as red chlorophylls) residing in the antenna are important for energy transfer dynamics and yield, however, their precise location remained elusive. Here, we construct a chimeric Photosystem I complex in Synechocystis PCC 6803 that shows enhanced absorption in the red spectral region. We combine Cryo-EM and spectroscopy to determine the structure-function relationship in this red-shifted Photosystem I complex. Determining the structure of this complex reveals the precise architecture of the low energy site as well as large scale structural heterogeneity which is probably universal to all trimeric Photosystem I complexes. Identifying the structural elements that constitute red sites can expand the absorption spectrum of oxygenic photosynthetic and potentially modulate light harvesting efficiency.

On the Red Antenna States of Photosystem I Mutants from Cyanobacteria Synechocystis PCC 6803.

To identify the molecular composition of the low-energy states in cyanobacterial Photosystem I (PSI) of Synechocystis PCC6803, we focus on high-resolution (low-temperature) absorption, emission, resonant, and nonresonant hole-burned spectra obtained for wild-type (WT) PSI and three PSI mutants. In the Red_a mutant, the B33 chlorophyll (Chl) is added to the B31-B32 dimer; in Red_b, histidine 95 (His95) on PsaB (which coordinates Mg in the B7 Chl within the His95-B7-A31-A32-cluster) is replaced with glutamine (Gln), while in the Red_ab mutant, both mutations are made. We show that the C706 state (B31-B32) changes to the C710 state (B31-B32-B33) in both Red_a and Red_ab mutants, while the C707 state in WT Synechocystis (localized on the His95-B7-A31-A32 cluster) is modified to C716 in both Red_b and Red_ab. Excitation energy transfer from C706 to the C714 trap in the WT PSI and Red_b mutant is hampered as reflected by a weak emission at 712 nm. Large electron-phonon coupling strength (exposed via resonant hole-burned spectra) is consistent with a strong mixing of excited states with intermolecular charge transfer states leading to significantly red-shifted emission spectra. We conclude that excitation energy transfer in PSI is controlled by fine-tuning the electronic states of a small number of highly conserved red states. Finally, we show that mutations modify the protein potential energy landscape as revealed by different shapes and shifts of the blue- and red-shifted antiholes.

The structure of the stress-induced photosystem I-IsiA antenna supercomplex.

Photochemical conversion in oxygenic photosynthesis takes place in two large protein-pigment complexes named photosystem II and photosystem I (PSII and PSI, respectively). Photosystems associate with antennae in vivo to increase the size of photosynthetic units to hundreds or thousands of pigments. Regulation of the interactions between antennae and photosystems allows photosynthetic organisms to adapt to their environment. In low-iron environments, cyanobacteria express IsiA, a PSI antenna, critical to their survival. Here we describe the structure of the PSI-IsiA complex isolated from the mesophilic cyanobacterium Synechocystis sp. PCC 6803. This 2-MDa photosystem-antenna supercomplex structure reveals more than 700 pigments coordinated by 51 subunits, as well as the mechanisms facilitating the self-assembly and association of IsiA with multiple PSI assemblies.

Crystal structures of virus-like photosystem I complexes from the mesophilic cyanobacterium Synechocystis PCC 6803.

Oxygenic photosynthesis supports virtually all life forms on earth. Light energy is converted by two photosystems-photosystem I (PSI) and photosystem II (PSII). Globally, nearly 50% of photosynthesis takes place in the Ocean, where single cell cyanobacteria and algae reside together with their viruses. An operon encoding PSI was identified in cyanobacterial marine viruses. We generated a PSI that mimics the salient features of the viral complex, named PSI(PsaJF). PSI(PsaJF) is promiscuous for its electron donors and can accept electrons from respiratory cytochromes. We solved the structure of PSI(PsaJF) and a monomeric PSI, with subunit composition similar to the viral PSI, providing for the first time a detailed description of the reaction center and antenna system from mesophilic cyanobacteria, including red chlorophylls and cofactors of the electron transport chain. Our finding extends the understanding of PSI structure, function and evolution and suggests a unique function for the viral PSI. DOI: http://dx.doi.org/10.7554/eLife.01496.001.

The evolution of photosystem I in light of phage-encoded reaction centres.

Recent structural determinations and metagenomic studies shed light on the evolution of photosystem I (PSI) from the homodimeric reaction centre of primitive bacteria to plant PSI at the top of the evolutionary development. The evolutionary scenario of over 3.5 billion years reveals an increase in the complexity of PSI. This phenomenon of ever-increasing complexity is common to all evolutionary processes that in their advanced stages are highly dependent on fine-tuning of regulatory processes. On the other hand, the recently discovered virus-encoded PSI complexes contain a minimal number of subunits. This may reflect the unique selection scenarios associated with viral replication. It may be beneficial for future engineering of productive processes to utilize 'primitive' complexes that disregard the cellular regulatory processes and to avoid those regulatory constraints when our goal is to divert the process from its original route. In this article, we discuss the evolutionary forces that act on viral reaction centres and the role of the virus-carried photosynthetic genes in the evolution of photosynthesis.

Temperature-sensitive PSII and promiscuous PSI as a possible solution for sustainable photosynthetic hydrogen production.

Sustainable hydrogen production in cyanobacteria becomes feasible as a result of our recent studies of the structure of photosystem I encoding operon in a marine phage. We demonstrated that the fused PsaJF subunit from the phage, substituted for the two separate subunits in Synechocystis, enabled the mutated PSI to accept electrons from additional electron donors such as respiratory cytochromes. In this way, a type of photorespiration was created in which the cell consumes organic material through respiratory processes and PSI serves as a terminal electron acceptor, substituting for cytochrome oxidase. We designed a hydrogen-producing bioreactor in which this type of photorespiration could utilize the organic material of the cell as an electron source for H(2) production. We propose, in parallel, to engineer cyanobacterial and/or algal strains with a temperature-sensitive PSII and enhanced respiration rates to achieve efficient and sustainable hydrogen production. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial.

Photosynthetic electron partitioning between [FeFe]-hydrogenase and ferredoxin:NADP+-oxidoreductase (FNR) enzymes in vitro.

Photosynthetic water splitting, coupled to hydrogenase-catalyzed hydrogen production, is considered a promising clean, renewable source of energy. It is widely accepted that the oxygen sensitivity of hydrogen production, combined with competition between hydrogenases and NADPH-dependent carbon dioxide fixation are the main limitations for its commercialization. Here we provide evidence that, under the anaerobic conditions that support hydrogen production, there is a significant loss of photosynthetic electrons toward NADPH production in vitro. To elucidate the basis for competition, we bioengineered a ferredoxin-hydrogenase fusion and characterized hydrogen production kinetics in the presence of Fd, ferredoxin:NADP(+)-oxidoreductase (FNR), and NADP(+). Replacing the hydrogenase with a ferredoxin-hydrogenase fusion switched the bias of electron transfer from FNR to hydrogenase and resulted in an increased rate of hydrogen photoproduction. These results suggest a new direction for improvement of biohydrogen production and a means to further resolve the mechanisms that control partitioning of photosynthetic electron transport.

Structure determination and improved model of plant photosystem I.

Photosystem I functions as a sunlight energy converter, catalyzing one of the initial steps in driving oxygenic photosynthesis in cyanobacteria, algae, and higher plants. Functionally, Photosystem I captures sunlight and transfers the excitation energy through an intricate and precisely organized antenna system, consisting of a pigment network, to the center of the molecule, where it is used in the transmembrane electron transfer reaction. Our current understanding of the sophisticated mechanisms underlying these processes has profited greatly from elucidation of the crystal structures of the Photosystem I complex. In this report, we describe the developments that ultimately led to enhanced structural information of plant Photosystem I. In addition, we report an improved crystallographic model at 3.3-A resolution, which allows analysis of the structure in more detail. An improved electron density map yielded identification and tracing of subunit PsaK. The location of an additional ten beta-carotenes as well as five chlorophylls and several loop regions, which were previously uninterpretable, are now modeled. This represents the most complete plant Photosystem I structure obtained thus far, revealing the locations of and interactions among 17 protein subunits and 193 non-covalently bound photochemical cofactors. Using the new crystal structure, we examine the network of contacts among the protein subunits from the structural perspective, which provide the basis for elucidating the functional organization of the complex.