Structural effects of high laser power densities on an early bacteriorhodopsin photocycle intermediate.

Time-resolved serial crystallography at X-ray Free Electron Lasers offers the opportunity to observe ultrafast photochemical reactions at the atomic level. The technique has yielded exciting molecular insights into various biological processes including light sensing and photochemical energy conversion. However, to achieve sufficient levels of activation within an optically dense crystal, high laser power densities are often used, which has led to an ongoing debate to which extent photodamage may compromise interpretation of the results.

Demonstration of full polarization control of soft X-ray pulses with Apple X undulators at SwissFEL using recoil ion momentum spectroscopy.

The ability to freely control the polarization of X-rays enables measurement techniques relying on circular or linear dichroism, which have become indispensable tools for characterizing the properties of chiral molecules or magnetic structures. Therefore, the demand for polarization control in X-ray free-electron lasers is increasing to enable polarization-sensitive dynamical studies on ultrafast time scales. The soft X-ray branch Athos of SwissFEL was designed with the aim of providing freely adjustable and arbitrary polarization by building its undulator solely from modules of the novel Apple X type.

Influence of pump laser fluence on ultrafast myoglobin structural dynamics.

High-intensity femtosecond pulses from an X-ray free-electron laser enable pump-probe experiments for the investigation of electronic and nuclear changes during light-induced reactions. On timescales ranging from femtoseconds to milliseconds and for a variety of biological systems, time-resolved serial femtosecond crystallography (TR-SFX) has provided detailed structural data for light-induced isomerization, breakage or formation of chemical bonds and electron transfer. However, all ultrafast TR-SFX studies to date have employed such high pump laser energies that nominally several photons were absorbed per chromophore.

Correlation of refractive index based and THz streaking arrival time tools for a hard X-ray free-electron laser.

To fully exploit ultra-short X-ray pulse durations routinely available at X-ray free-electron lasers to follow out-of-equilibrium dynamics, inherent arrival time fluctuations of the X-ray pulse with an external perturbing laser pulse need to be measured. In this work, two methods of arrival time measurement were compared to measure the arrival time jitter of hard X-ray pulses. The methods were photoelectron streaking by a THz field and a transient refractive index change of a semiconductor.

Visualizing the DNA repair process by a photolyase at atomic resolution.

Photolyases, a ubiquitous class of flavoproteins, use blue light to repair DNA photolesions. In this work, we determined the structural mechanism of the photolyase-catalyzed repair of a cyclobutane pyrimidine dimer (CPD) lesion using time-resolved serial femtosecond crystallography (TR-SFX). We obtained 18 snapshots that show time-dependent changes in four reaction loci.

Disentangling the evolution of electrons and holes in photoexcited ZnO nanoparticles.

The evolution of charge carriers in photoexcited room temperature ZnO nanoparticles in solution is investigated using ultrafast ultraviolet photoluminescence spectroscopy, ultrafast Zn K-edge absorption spectroscopy, and molecular dynamics (MD) simulations. The photoluminescence is excited at 4.66 eV, well above the band edge, and shows that electron cooling in the conduction band and exciton formation occur in <500 fs, in excellent agreement with theoretical predictions.

An X-ray free-electron laser with a highly configurable undulator and integrated chicanes for tailored pulse properties.

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.

Ultrafast structural changes direct the first molecular events of vision.

Vision is initiated by the rhodopsin family of light-sensitive G protein-coupled receptors (GPCRs). A photon is absorbed by the 11-cis retinal chromophore of rhodopsin, which isomerizes within 200 femtoseconds to the all-trans conformation, thereby initiating the cellular signal transduction processes that ultimately lead to vision. However, the intramolecular mechanism by which the photoactivated retinal induces the activation events inside rhodopsin remains experimentally unclear.

Ultrafast Energy Transfer from Photoexcited Tryptophan to the Haem in Cytochrome c.

We report femtosecond Fe K-edge absorption (XAS) and nonresonant X-ray emission (XES) spectra of ferric cytochrome C (Cyt c) upon excitation of the haem (>300 nm) or mixed excitation of the haem and tryptophan (<300 nm). The XAS and XES transients obtained in both excitation energy ranges show no evidence for electron transfer processes between photoexcited tryptophan (Trp) and the haem, but rather an ultrafast energy transfer, in agreement with previous ultrafast optical fluorescence and transient absorption studies. The reported ( , 115 (46), 13723-13730) decay times of Trp fluorescence in ferrous (∼350 fs) and ferric (∼700 fs) Cyt c are among the shortest ever reported for Trp in a protein.

Ultrafast Single-Particle Imaging with Intense X-Ray Pulses.

Ultrafast single-particle imaging with intense x-ray pulses from free-electron laser sources provides a new approach for visualizing structure and dynamics on the nanoscale. After a short introduction to the novel free-electron laser sources and methods, we highlight selected applications and discuss how ultrafast imaging flourishes from method development to early applications in physics and biology to opportunities for chemical sciences.

Lipidic cubic phase serial femtosecond crystallography structure of a photosynthetic reaction centre.

Serial crystallography is a rapidly growing method that can yield structural insights from microcrystals that were previously considered to be too small to be useful in conventional X-ray crystallography. Here, conditions for growing microcrystals of the photosynthetic reaction centre of Blastochloris viridis within a lipidic cubic phase (LCP) crystallization matrix that employ a seeding protocol utilizing detergent-grown crystals with a different crystal packing are described. LCP microcrystals diffracted to 2.

Rovibrational investigation of a new high-lying 0 state of Cu by using two-color resonant four-wave-mixing spectroscopy.

A highly excited electronic state of dicopper is observed and characterized for the first time. The [39.6]0 -XΣ (0 ) system is measured at rotational resolution by using degenerate and two-color resonant four-wave-mixing, as well as laser induced fluorescence spectroscopy.

A perfect X-ray beam splitter and its applications to time-domain interferometry and quantum optics exploiting free-electron lasers.

X-ray free-electron lasers (FELs) deliver ultrabright X-ray pulses, but not the sequences of phase-coherent pulses required for time-domain interferometry and control of quantum states. For conventional split-and-delay schemes to produce such sequences, the challenge stems from extreme stability requirements when splitting Ångstrom wavelength beams, where the tiniest path-length differences introduce phase jitter. We describe an FEL mode based on selective electron-bunch degradation and transverse beam shaping in the accelerator, combined with a self-seeded photon emission scheme.

Dynamics and mechanism of a light-driven chloride pump.

Chloride transport by microbial rhodopsins is an essential process for which molecular details such as the mechanisms that convert light energy to drive ion pumping and ensure the unidirectionality of the transport have remained elusive. We combined time-resolved serial crystallography with time-resolved spectroscopy and multiscale simulations to elucidate the molecular mechanism of a chloride-pumping rhodopsin and the structural dynamics throughout the transport cycle. We traced transient anion-binding sites, obtained evidence for how light energy is used in the pumping mechanism, and identified steric and electrostatic molecular gates ensuring unidirectional transport.

Pink-beam serial femtosecond crystallography for accurate structure-factor determination at an X-ray free-electron laser.

Serial femtosecond crystallography (SFX) at X-ray free-electron lasers (XFELs) enables essentially radiation-damage-free macromolecular structure determination using microcrystals that are too small for synchrotron studies. However, SFX experiments often require large amounts of sample in order to collect highly redundant data where some of the many stochastic errors can be averaged out to determine accurate structure-factor amplitudes. In this work, the capability of the Swiss X-ray free-electron laser (SwissFEL) was used to generate large-bandwidth X-ray pulses [Δλ/λ = 2.

Femtosecond X-ray spectroscopy of haem proteins.

We discuss our recently reported femtosecond (fs) X-ray emission spectroscopy results on the ligand dissociation and recombination in nitrosylmyoglobin (MbNO) in the context of previous studies on ferrous haem proteins. We also present a preliminary account of femtosecond X-ray absorption studies on MbNO, pointing to the presence of more than one species formed upon photolysis.

Advances in long-wavelength native phasing at X-ray free-electron lasers.

Long-wavelength pulses from the Swiss X-ray free-electron laser (XFEL) have been used for protein structure determination by native single-wavelength anomalous diffraction (native-SAD) phasing of serial femtosecond crystallography (SFX) data. In this work, sensitive anomalous data-quality indicators and model proteins were used to quantify improvements in native-SAD at XFELs such as utilization of longer wavelengths, careful experimental geometry optimization, and better post-refinement and partiality correction. Compared with studies using shorter wavelengths at other XFELs and older software versions, up to one order of magnitude reduction in the required number of indexed images for native-SAD was achieved, hence lowering sample consumption and beam-time requirements significantly.

Generation and simple characterization of flat, liquid jets.

We present an approach to determine the absolute thickness profile of flat liquid jets, which takes advantage of the information of thin film interference combined with light absorption, both captured in a single microscopic image. The feasibility of the proposed method is demonstrated on our compact experimental setup used to generate micrometer thin, free-flowing liquid jet sheets upon collision of two identical laminar cylindrical jets. Stable operation was achieved over several hours of the flat jet in vacuum (10 mbar), making the system ideally suitable for soft x-ray photon spectroscopy of liquid solutions.

Post-treatment of ozonated wastewater with activated carbon and biofiltration compared to membrane bioreactors: Toxicity removal in vitro and in Potamopyrgus antipodarum.

Wastewater treatment plants are major point sources of (micro)pollutant emissions and advanced wastewater treatment technologies can improve their removal capacity. While abundant data on individual advanced treatment technologies is available, there is limited knowledge regarding the removal performance of ozonation combined with multiple post-treatments and stand-alone membrane bioreactors. This is especially true for the removal of in vitro and in vivo toxicity.

Spin cascade and doming in ferric hemes: Femtosecond X-ray absorption and X-ray emission studies.

The structure-function relationship is at the heart of biology, and major protein deformations are correlated to specific functions. For ferrous heme proteins, doming is associated with the respiratory function in hemoglobin and myoglobins. Cytochrome (Cyt c) has evolved to become an important electron-transfer protein in humans.

Femtosecond-to-millisecond structural changes in a light-driven sodium pump.

Light-driven sodium pumps actively transport small cations across cellular membranes. These pumps are used by microorganisms to convert light into membrane potential and have become useful optogenetic tools with applications in neuroscience. Although the resting state structures of the prototypical sodium pump Krokinobacter eikastus rhodopsin 2 (KR2) have been solved, it is unclear how structural alterations over time allow sodium to be translocated against a concentration gradient.

Optical second harmonic generation in LiBO modulated by intense femtosecond X-ray pulses.

Many of the scientific applications for X-ray free-electron lasers seek to exploit the ultrashort pulse durations of intense X-rays to obtain femtosecond time resolution of various processes in a "pump-probe" scheme. One of the limiting factors for such experiments is the timing jitter between the X-rays and ultrashort pulses from more conventional lasers operating at near-optical wavelengths. In this work, we investigate the potential of using X-ray-induced changes in the optical second harmonic generation efficiency of a nonlinear crystal to retrieve single-shot arrival times of X-ray pulses with respect to optical laser pulses.

Taking a snapshot of the triplet excited state of an OLED organometallic luminophore using X-rays.

OLED technology beyond small or expensive devices requires light-emitters, luminophores, based on earth-abundant elements. Understanding and experimental verification of charge transfer in luminophores are needed for this development. An organometallic multicore Cu complex comprising Cu-C and Cu-P bonds represents an underexplored type of luminophore.

Core-level nonlinear spectroscopy triggered by stochastic X-ray pulses.

Stochastic processes are highly relevant in research fields as different as neuroscience, economy, ecology, chemistry, and fundamental physics. However, due to their intrinsic unpredictability, stochastic mechanisms are very challenging for any kind of investigations and practical applications. Here we report the deliberate use of stochastic X-ray pulses in two-dimensional spectroscopy to the simultaneous mapping of unoccupied and occupied electronic states of atoms in a regime where the opacity and transparency properties of matter are subject to the incident intensity and photon energy.

How the methyl group position influences the ultrafast deactivation in aromatic radicals.

Excited xylyl (methyl-benzyl) radical isomers have been studied by femtosecond time-resolved photoelectron spectroscopy and mass spectrometry. Depending on the substitution we find different deactivation channels after excitation into the D3(2A'') state (310 nm, 4 eV). While the ortho and para isomer exhibit deactivation rates similar to the benzyl radical, meta-xylyl sticks out and depletes twice as fast into the vibrationally hot ground state.