Purple phototrophic bacteria produce two kinds of light-harvesting complexes that function to capture and transmit solar energy: the core antenna (LH1) and the peripheral antenna (LH2). The apoproteins of these antennas, encoded respectively by the genes pufBA and pucBA within and outside the photosynthetic gene cluster, respectively, exhibit conserved amino acid sequences and structural topologies suggesting they were derived from a shared ancestor. Here we present the structures of two photosynthetic complexes from Roseospirillum (Rss.) parvum 930I: an LH1-RC complex and a variant of the LH1 complex also encoded by pufBA that we designate as LH1'. The LH1-RC complex forms a closed elliptical structure consisting of 16 pairs of αβ-polypeptides that surrounds the RC. By contrast, the LH1' complex is a closed ring structure composed of 14 pairs of αβ-polypeptides, and it shows significant similarities to LH2 complexes both spectrally and structurally. Although LH2-like, the LH1' complex is larger than any known LH2 complexes, and genomic analyses of Rss. parvum revealed the absence of pucBA, genes that encode classical LH2 complexes. Characterization of the unique Rss. parvum photocomplexes not only underscores the diversity of such structures but also sheds new light on the evolution of light-harvesting complexes from phototrophic bacteria.

The plastid-encoded RNA polymerase serves as the principal transcription machinery within chloroplasts, transcribing over 80% of all primary plastid transcripts. This polymerase consists of a prokaryotic-like core enzyme known as the plastid-encoded RNA polymerase core, and is supplemented by newly evolved associated proteins known as PAPs. However, the architecture of the plastid-encoded RNA polymerase and the possible functions of PAPs remain unknown. Here, we present the cryo-electron microscopy structure of a 19-subunit plastid-encoded RNA polymerase complex derived from spinach (Spinacia oleracea). The structure shows that the plastid-encoded RNA polymerase core resembles bacterial RNA polymerase. Twelve PAPs and two additional proteins (FLN2 and pTAC18) bind at the periphery of the plastid-encoded RNA polymerase core, forming extensive interactions that may facilitate complex assembly and stability. PAPs may also protect the complex against oxidative damage and has potential functions in transcriptional regulation. This research offers a structural basis for future investigations into the functions and regulatory mechanisms governing the transcription of plastid genes. The plastid-encoded RNA polymerase serves as the principal transcription machinery within chloroplasts. Here, the authors present the cryo-electron microscopy structure of a 19-subunit plastid-encoded RNA polymerase complex derived from spinach.

The intrinsic energy spread of electron beams needs to be measured to characterize and optimize high-brightness electron beam sources such as those driving x-ray free-electron lasers (FELs). We demonstrate the use of the optical klystron effect as a precise and high-resolution method to measure the electron beam energy spread. The optical klystron setup consists of undulator modules and magnetic chicanes placed between them. The energy spread is found by measuring the radiation power produced in the undulators as a function of the chicanes’ strengths. High resolution and simplicity are the advantages of this approach, in contrast to the standard method, which measures the longitudinal phase space of the electron beam with a transverse deflector. The demonstration was performed at Athos, the soft x-ray FEL beamline of SwissFEL, for which we measured energy spreads below 1 MeV at a central beam energy of 3.4 GeV. We have verified the consistency of the method for different parameters (radiation wavelengths, undulator polarization configurations, and electron bunch durations) and we have benchmarked it against the standard measurement approach using a transverse deflector. Our results confirm the optical klystron as a valid approach to measure the electron beam energy spread. The method can be especially useful to reconstruct low energy spread values, where the conventional approach may be resolution limited, such as in ultra high-brightness radiofrequency photoinjectors or plasma sources, or when transverse deflectors are not available.

Marine photosynthetic dinoflagellates are a group of successful phytoplankton that can form red tides in the ocean and also symbiosis with corals. These features are closely related to the photosynthetic properties of dinoflagellates. We report here three structures of photosystem I (PSI)-chlorophylls (Chls) a/c- peridinin protein complex (PSI- AcpPCI) from two species of dinoflagellates by single- particle cryoelectron microscopy. The crucial PsaA/B subunits of a red tidal dinoflagellate Amphidinium carterae are remarkably smaller and hence losing over 20 pigment- binding sites, whereas its PsaD/ F/I/J/L/M/R subunits are larger and coordinate some additional pigment sites compared to other eukaryotic photosynthetic organisms, which may compensate for the smaller PsaA/B subunits. Similar modifications are observed in a coral symbiotic dinoflagellate Symbiodinium species, where two additional core proteins and fewer AcpPCIs are identified in the PSI-AcpPCI supercomplex. The antenna proteins AcpPCIs in dinoflagellates developed some loops and pigment sites as a result to accommodate the changed PSI core, therefore the structures of PSI-AcpPCI supercomplex of dinoflagellates reveal an unusual protein assembly pattern. A huge pigment network comprising Chls a and c and various carotenoids is revealed from the structural analysis, which provides the basis for our deeper understanding of the energy transfer and dissipation within the PSI-AcpPCI supercomplex, as well as the evolution of photosynthetic organisms.

ABSTRACTHalorhodospira (Hlr.) halochloris is a triply extremophilic phototrophic purple sulfur bacterium, as it is thermophilic, alkaliphilic, and extremely halophilic. The light‐harvesting‐reaction center (LH1–RC) core complex of this bacterium displays an LH1‐Qy transition at 1,016 nm, which is the lowest‐energy wavelength absorption among all known phototrophs. Here we report the cryo‐EM structure of the LH1–RC at 2.42 Å resolution. The LH1 complex forms a tricyclic ring structure composed of 16 αβγ‐polypeptides and one αβ‐heterodimer around the RC. From the cryo‐EM density map, two previously unrecognized integral membrane proteins, referred to as protein G and protein Q, were identified. Both of these proteins are single transmembrane‐spanning helices located between the LH1 ring and the RC L‐subunit and are absent from the LH1–RC complexes of all other purple bacteria of which the structures have been determined so far. Besides bacteriochlorophyll b molecules (B1020) located on the periplasmic side of the Hlr. halochloris membrane, there are also two arrays of bacteriochlorophyll b molecules (B800 and B820) located on the cytoplasmic side. Only a single copy of a carotenoid (lycopene) was resolved in the Hlr. halochloris LH1–α3β3 and this was positioned within the complex. The potential quinone channel should be the space between the LH1–α3β3 that accommodates the single lycopene but does not contain a γ‐polypeptide, B800 and B820. Our results provide a structural explanation for the unusual Qy red shift and carotenoid absorption in the Hlr. halochloris spectrum and reveal new insights into photosynthetic mechanisms employed by a species that thrives under the harshest conditions of any phototrophic microorganism known.

Purple phototrophic bacteria possess light-harvesting 1 and reaction center (LH1-RC) core complexes that play a key role in converting solar energy to chemical energy. High-resolution structures of LH1-RC and RC complexes have been intensively studied and have yielded critical insight into the architecture and interactions of their proteins, pigments, and cofactors. Nevertheless, a detailed picture of the structure and assembly of LH1-only complexes is lacking due to the intimate association between LH1 and the RC. To study the intrinsic properties and structure of an LH1-only complex, a genetic system was constructed to express the Thermochromatium (Tch.) tepidum LH1 complex heterologously in a modified Rhodospirillum rubrum mutant strain. The heterologously expressed Tch. tepidum LH1 complex was isolated in a pure form free of the RC and exhibited the characteristic absorption properties of Tch. tepidum. Cryo-EM structures of the LH1-only complexes revealed a closed circular ring consisting of either 14 or 15 αβ-subunits, making it the smallest completely closed LH1 complex discovered thus far. Surprisingly, the Tch. tepidum LH1-only complex displayed even higher thermostability than that of the native LH1-RC complex. These results reveal previously unsuspected plasticity of the LH1 complex, provide new insights into the structure and assembly of the LH1-RC complex, and show how molecular genetics can be exploited to study membrane proteins from phototrophic organisms whose genetic manipulation is not yet possible.

This article presents a demonstration of the improved performance of an X-ray free-electron laser (FEL) using the optical klystron mechanism and helical undulator configuration, in comparison with the common planar undulator configuration without optical klystron. The demonstration was carried out at Athos, the soft X-ray beamline of SwissFEL. Athos has variable-polarization undulators, and small magnetic chicanes placed between every two undulators to fully exploit the optical klystron. It was found that, for wavelengths of 1.24 nm and 3.10 nm, the required length to achieve FEL saturation is reduced by about 35% when using both the optical klystron and helical undulators, with each effect accounting for about half of the improvement. Moreover, it is shown that a helical undulator configuration provides a 20% to 50% higher pulse energy than planar undulators. This work represents an important step towards more compact and high-power FELs, rendering this key technology more efficient, affordable and accessible to the scientific community.

We present the generation of x-ray pulses with average pulse energies up to one millijoule and rms pulse durations down to the femtosecond level. We have produced these intense and short pulses by employing the fresh-slice multistage amplification scheme with a transversely tilted electron beam in a free-electron laser. In this scheme, a short pulse is produced in the first stage and later amplified by fresh parts of the electron bunch in up to a total of four stages of amplification. Our implementation is efficient, since practically the full electron beam contributes to produce the x-ray pulse. Our implementation is also compact, utilizing only 32 m of undulator. The demonstration was done at Athos, the soft x-ray beamline of SwissFEL, which was designed with high flexibility to take full advantage of the multistage amplification scheme. It opens the door for scientific opportunities following ultrafast dynamics using nonlinear x-ray spectroscopy techniques or avoiding electronic damage when capturing structures with a single intense pulse via single-particle imaging.

The light-harvesting (LH) and reaction center (RC) core complex of purple bacterium Roseiflexus castenholzii, B880-B800-RC, are different from those of the typical photosynthetic unit, (B850-B800)(x)-B880-RC. To investigate the excitation flowing dynamics in this unique complex, two-dimensional electronic spectroscopy is employed. The obtained time constants for the exciton relaxation in B880, exciton relaxation in B800, B800 -> B880 energy transfer (EET), and B880 -> closed RC EET are 43 fs, 177 fs, 1.9 ps, and 205 ps, respectively. These time constants result in an overall EET efficiency similar to that of the typical photosynthetic unit. Analysis of the oscillatory signals reveals that while several vibronic coherences are involved in the exciton relaxation process, only one prominent vibronic coherence, with a frequency of 27 cm(-1) and coupled to the B880 electronic transition, may contribute to the B800 -> B880 EET process.

In wild-type phototrophic organisms, carotenoids (Crts) are primarily packed into specific pigment-protein complexes along with (Bacterio)chlorophylls and play important roles in the photosynthesis. Diphenylamine (DPA) inhibits carotenogenesis but not phototrophic growth of anoxygenic phototrophs and eliminates virtually all Crts from photocomplexes. To investigate the effect of Crts on assembly of the reaction center-light-harvesting (RC-LH) complex from the filamen- tous anoxygenic phototroph Roseiflexus (Rfl.) castenholzii, we generated carotenoidless (Crt-less) RC-LH complexes by growing cells in the presence of DPA. Here, we present cryoEM structures of the Rfl. castenholzii native and Crt-less RC-LH complexes with resolutions of 2.86 angstrom and 2.85 angstrom, respectively. From the high-quality map obtained, several important but previously unresolved details in the Rfl. castenholzii RC-LH structure were determined unambiguously including the assignment and likely function of three small polypeptides, and the content and spatial arrangement of Crts with bacteriochlorophyll molecules. The overall structures of Crt-containing and Crt-less complexes are similar. However, structural comparisons showed that only five Crts remain in complexes from DPA-treated cells and that the subunit X (TMx) flanked on the N -terminal helix of the Cyt-subunit is missing. Based on these results, the function of Crts in the assembly of the Rfl. castenholzii RC-LH complex and the molecular mechanism of quinone exchange is discussed. These structural details provide a fresh look at the photosynthetic apparatus of an evolutionary ancient phototroph as well as new insights into the importance of Crts for proper assembly and functioning of the RC-LH complex.

Radiofrequency (RF) transverse deflection structures (TDSs) are fundamental time-resolved diagnostics in x-ray free-electron lasers. Two x-band TDSs with variable polarization of the deflecting force were recently installed after the undulators of Athos, the soft x-ray beamline of SwissFEL. This contribution summarizes the experience gained over the last few months during the commissioning of the RF system and the measurements made during operations, focusing on the setup of the entire complex RF system, the calibration and the time-resolved measurement that, combined with an energy spectrometer, provides longitudinal phase-space measurements of extreme importance for the commissioning of the complex FEL schemes implemented in the Athos beamline.

We demonstrate the generation of coherent soft x-ray free-electron laser (FEL) pulses with a duration below 1 fs using nonlinear compression with a low-charge electron beam (10 pC). The approach is simple, and it does not require any special hardware, so it can be readily implemented at any x-ray FEL facility. We present temporal and spectral diagnostics confirming the production of single-spike sub-femtosecond pulses for photon energies of 642 and 1111 eV. Our work will be important for ultrafast FEL applications requiring soft x-rays.

X-ray free-electron lasers (FELs) are state-of-the-art scientific tools capable to study matter on the scale of atomic processes. Since the initial operation of X-ray FELs more than a decade ago, several facilities with upgraded performance have been put in operation. Here we present the first lasing results of Athos, the soft X-ray FEL beamline of SwissFEL at the Paul Scherrer Institute in Switzerland. Athos features an undulator layout based on short APPLE-X modules providing full polarisation control, interleaved with small magnetic chicanes. This versatile configuration allows for many operational modes, giving control over many FEL properties. We show, for example, a 35% reduction of the required undulator length to achieve FEL saturation with respect to standard undulator configurations. We also demonstrate the generation of more powerful pulses than the ones obtained in typical undulators. Athos represents a fundamental step forward in the design of FEL facilities, creating opportunities in FEL-based sciences.

SwissFEL has a unique capability, among the normal conducting linac-based light sources, to simultaneously serve two separate undulator lines (Aramis and Athos) up to the machine repetition rate of 100 Hz using the double bunch operation mode. It increases twice the experiments throughput of the facility with modest additional investment. Two electron bunches spaced 28 ns apart are extracted from the cathode by two laser pulses with individually controlled repetition rates. The bunches are accelerated up to about 3 GeV in the main linac using the same rf macropulse. After separation, one bunch serves the Athos soft x-ray beamline and the other is further accelerated to serve the hard x-ray beamline - Aramis. A fast and high-stability beam kicker separates the two bunches without disturbing the electron beam and consequently the x-ray lasing. The timing and control system sets hybrid machine modes utilizing independent operation of the two undulator lines with individually programmed repetition rates. Beam diagnostics and feedback systems have to operate with two closely spaced bunches where the two beams share the same machine path. The low-level rf system manipulates the rf amplitude and phase within a fraction of the rf macropulse to provide decoupling of the acceleration parameters of the first and the second bunch. This manuscript presents measurements that show that the bunch separation does not degrade FEL lasing stability.

We demonstrate the generation of widely tunable two-color x-ray free-electron laser (FEL) pulses at SwissFEL. In a split-undulator configuration, each color is produced in a different undulator section, and a chicane between the two sections allows for a variable time separation between the two pulses of up to 500 fs. We show an unprecedented photon energy ratio between the two colors of about three (350 and 915 eV), with each individual pulse having a peak power of a few gigawatts and a duration down to the femtosecond level. Moreover, we demonstrate the reduction of the required undulator length via the optical klystron mechanism and the time-resolved diagnostics of the FEL pulses utilizing the same beam setup as for the pulse generation. The unique combination of widely tunable energy and time separation of the two-color pulse pair offers opportunities to study ultrafast x-ray-induced energy transfer and relaxation processes in physics, chemistry, and biology.

A superconducting linear accelerator operating in continuous-wave mode could produce X-ray free electron lasers (XFEL) at megahertz repetition rate, with the capability that delivering wide spectral range coherent radiation to multi end stations. In this Letter, the energy recovery Linac (ERL) mode is proposed to flexibly control the electron beam energy for a continuous-wave superconducting Linac. Theoretical investigations and multi-dimensional numerical simulations are applied to the Linac case of Shanghai high-repetition-rate XFEL and extreme light facility. The results show that, with ERL operation in the last 25 cryo-modules, the strict requirements on RF power system could be significantly relaxed. And if one exhaust the RF power, the maximum electron beam energy can be enhanced from 8.74 GeV to 11.41GeV in ERL mode. The optimization of the ERL operation, the multi-energy electron beam transport and the XFEL performance improvements are presented.

Development of advanced light sources has become more and more important in the frontier scientific research. The development of new advanced light sources has pushed experimental studies from macroscopic world into the atomic and molecular levels, from studies of physical systems at static conditions to ultrafast dynamical processes, and from investigations at simple experimental conditions to more complex and real environment. Free electron laser (FEL) with high brightness, ultrafast laser pulses in the extreme ultraviolet (EUV) wavelength region is an ideal light source for excitation of valence electrons and ionization of molecular systems with very high efficiency. Dalian Coherent Light Source (DCLS) delivers 50-150 nm EUV beams with ultrafast pulse durations of 100 fs or ps to users. DCLS, as the first FEL user facility in China, has wide applications in the frontier researches of physics, chemistry, biology, especially in the field of basic energy sciences. This project is supported by National Natural Science Foundation of China and developed jointly by Dalian Institute of Chemical Physics and Shanghai Institute of Applied Physics, Chinese Academy of Sciences.

The Dalian coherent light source is a free-electron laser facility based on the principle of high-gain harmonic generation, which can generate lasers with full coherence, short pulse and high peak power in 50 similar to 150nm. Advanced laser tools can be provided for exploring molecular ionization and dissociation in nature. The main function of the beamline for Dalian coherent light source is to transmit the laser efficiently to the end experimental station and perform optical diagnosis such as light intensity and spectrum. Dalian coherent light source beam line consists of Beam Define Aperture (BDA), gas monitor detector (GMD), Beam Position Monitor (BPM), Photon Block (PB), gas absorber (GA), profile monitor (PRF),on-line spectrum and several different functional mirrors including a focusing mirror and three deflecting mirrors. In this paper, the basic principles and mechanical structure of each functional module are briefly described.

Dalian Coherent Light Source (DCLS) is a free-electron laser (FEL) user facility working in the extreme ultraviolet (EUV) wavelength region from 50 to 150 nm. It mainly operates on the High Gain Harmonic Generation (HGHG) mode with the seed laser, although it can also run in the Self Amplified Spontaneous Emission (SASE) mode. The brightness and bandwidth of FEL radiation strongly depends on electron bunch quality, such as normalized transverse emittance, electron bunch energy, energy spread, peak current, etc. The high peak current with uniform longitudinal distribution is especially helpful for high peak power and narrow bandwidth of FEL, although it is not easy to achieve, due to the nonlinearity of sinusoidal accelerating radio frequency (RF) field and the 2-order momentum compaction coefficient T566 of bunch compressor. An X-band linearizer will be installed before the bunch compressor in order to correct this nonlinearity properly. In this paper, the beam dynamics design of the X-band linear compression system in DCLS is focused, and the simulation results with Elegant are presented and discussed.

The echo-enabled harmonic generation (EEHG) scheme holds promising prospects for efficiently generating intense coherent radiation at very high harmonics of a conventional ultraviolet seed laser. We report the lasing of the EEHG free-electron laser (FEL) at an extreme ultraviolet (EUV) wavelength with a seeded FEL facility, the Shanghai soft x-ray FEL. For the first time, we have benchmarked the basic theory of EEHG by measuring the bunching factor distributions over one octave down to the EUV region. Our results demonstrated the key advantages of the EEHG FEL, i.e., generation of very high harmonics with a small laser-induced energy spread and insensitivity to beam imperfections, and marks a great step towards fully coherent x rays with the EEHG scheme.