Search

X-ray free-electron lasers (XFELs) can probe chemical and biological reactions as they unfold with unprecedented spatial and temporal resolution. A principal challenge in this pursuit involves the delivery of samples to the X-ray interaction point in such a way that produces data of the highest possible quality and with maximal efficiency. This is hampered by intrinsic constraints posed by the light source and operation within a beamline environment. For liquid samples, the solution typically involves some form of high-speed liquid jet, capable of keeping up with the rate of X-ray pulses. However, conventional jets are not ideal because of radiation-induced explosions of the jet, as well as their cylindrical geometry combined with the X-ray pointing instability of many beamlines which causes the interaction volume to differ for every pulse. This complicates data analysis and contributes to measurement errors. An alternative geometry is a liquid sheet jet which, with its constant thickness over large areas, eliminates the problems related to X-ray pointing. Since liquid sheets can be made very thin, the radiation-induced explosion is reduced, boosting their stability. These are especially attractive for experiments which benefit from small interaction volumes such as fluctuation X-ray scattering and several types of spectroscopy. Although their use has increased for soft X-ray applications in recent years, there has not yet been wide-scale adoption at XFELs. Here, gas-accelerated liquid sheet jet sample injection is demonstrated at the European XFEL SPB/SFX nano focus beamline. Its performance relative to a conventional liquid jet is evaluated and superior performance across several key factors has been found. This includes a thickness profile ranging from hundreds of nanometres to 60 nm, a fourfold increase in background stability and favorable radiation-induced explosion dynamics at high repetition rates up to 1.13 MHz. Its minute thickness also suggests that ultrafast single-particle solution scattering is a possibility.

Phytochromes are red-light photoreceptors that were first characterized in plants, with homologs in photosynthetic and non-photosynthetic bacteria known as bacteriophytochromes (BphPs). Upon absorption of light, BphPs interconvert between two states denoted Pr and Pfr with distinct absorption spectra in the red and far-red. They have recently been engineered as enzymatic photoswitches for fluorescent-marker applications in non-invasive tissue imaging of mammals. This article presents cryo- and room-temperature crystal structures of the unusual phytochrome from the non-photosynthetic myxobacterium Stigmatella aurantiaca (SaBphP1) and reveals its role in the fruiting-body formation of this photomorphogenic bacterium. SaBphP1 lacks a conserved histidine (His) in the chromophore-binding domain that stabilizes the Pr state in the classical BphPs. Instead it contains a threonine (Thr), a feature that is restricted to several myxobacterial phytochromes and is not evolutionarily understood. SaBphP1 structures of the chromophore binding domain (CBD) and the complete photosensory core module (PCM) in wild-type and Thr-to-His mutant forms reveal details of the molecular mechanism of the Pr/Pfr transition associated with the physiological response of this myxobacterium to red light. Specifically, key structural differences in the CBD and PCM between the wild-type and the Thr-to-His mutant involve essential chromophore contacts with proximal amino acids, and point to how the photosignal is transduced through the rest of the protein, impacting the essential enzymatic activity in the photomorphogenic response of this myxobacterium.