Using X-ray diffraction for structure determination is further complicated when the irradiated particles have random and unknown orientations, because it means that the relations between the diffraction images are also unknown. Algorithms exist for recovering the relative orientations between diffraction patterns, but they do not always converge, especially not when faced with scarce or noisy...
MS SPIDOC is an innovative sample delivery system tailored for single-particle imaging at X-ray Free-Electron Lasers (XFELs) and adaptable to most large-scale facility beamlines. It accommodates a wide range of biological samples, from small proteins to megadalton (MDa) complexes. Utilizing nano-electrospray ionization, ionic samples are $m/z$-filtered and structurally separated before...
Non-aromatic fluorescence in biomolecules represents a fascinating photophysical phenomenon that challenges conventional understanding of fluorescent mechanisms.. This presentation outlines our ongoing investigation using a combined approach of time-resolved X-ray absorption spectroscopy (TrXAS) and multiple vibrational spectroscopic techniques to explore the fundamental processes governing...
Normal-to-cancer transition (NTC) is still an ill-understood process, closely associated to cellular biomechanical properties. These are strongly dependent on intracellular water´s structural and dynamical profiles, which play a fundamental role in cellular function. Improved chemotherapeutic strategies are an urgent clinical need, since cancer is still the second leading cause of death...
Studying structural changes associated with protein aggregation is challenging and often requires a combination of experimental techniques to capture insights at the molecular level across different scales, from nanometers to microns. Studying this process becomes even more complex when aggregation occurs in the presence of molecular co-factors, nucleic acids among them, and when the resulting...
X-ray Single Particle Imaging has the aim of imaging biomolecules without the need of crystallization. The invention of X-ray Free Electron Lasers (XFELs) provided the instruments for this imaging process, however the technique is still suffering from low signal to noise ratio. Retrieval of the orientation of the sample in the moment of photon-sample interaction would greatly improve the...
Marine microplastic pollution has emerged as a major global concern, with growing implications for both marine ecosystems and human health [1]. Polystyrene nanoparticles (PS-NPs) can induce significant biological responses, as demonstrated by combined in vivo and in vitro models. In zebrafish, exposure to PS-NPs resulted in marked changes in eye pigmentation patterns that were not associated...
Proteins, constituting the virus structure, cover a wide and diverse range of functions. Spike glycoprotein (S) of SARS-CoV-2 is a notable example. As the largest structural protein of the virus, the S protein plays a crucial role in attaching to the host receptor ACE2 through its receptor-binding domain (RBD) [1]. The functionalities of these membrane proteins, such as cellular targeting and...
Imaging the structure and observing the dynamics of isolated proteins using single-particle X-ray diffractive imaging (SPI) is one of the potential applications of X-ray free-electron lasers (XFELs). Currently, SPI experiments on isolated proteins are limited by three factors: low signal strength, limited data and high background from gas scattering. The last two factors are largely due to the...
Picosecond-long x-ray pulses of moderate intensity and up to MHz CW repetition rate for time-resolved analysis of matter in the linear response regime are proposed for un upgrade operation of Elettra 2.0, now in construction as funded successor of the Elettra storage ring light source in Trieste, Italy. The scheme, based on the adoption of radiofrequency transverse deflecting cavities,...
Single particle imaging of biomolecules using Free electron lasers (FEL) is an imaging technique that has been under development since the dawn of FEL:s more than two decades ago. Due to the heavy ionisation, biomolecules exposed to FEL pulses explode. In a recent publication (Phys. Rev. Lett. 134, 128403 (2025)) we have described how we can harvest information about the molecular structure of...
A way to induce local damage to cancerous tissue is by using radiotherapy-amplifying bioagents doped with high-Z elements. This enables deep core-level ionisation during radiotherapy with X-rays above the K-edge threshold, significantly increasing radiation absorption. Core electron ejection from high-Z elements also triggers a cascade of secondary particles, amplifying damage.
We studied...
Classical all-atom molecular dynamics (MD) simulations are a powerful tool for investigating the structures and interactions of biomolecules in the gas phase. Until recently, these simulations were limited to systems containing only a few thousand atoms, due to the quadratic scaling of computational cost with system size. However, the recent integration of a linearly-scaling algorithm for...
The European XFEL (EuXFEL) enables high resolution and time-resolved structural studies of biological systems. Diffraction, scattering and spectroscopy experiments with a MHz pulse repetition rate, in combination with optical pump-probe or mixing set-ups, give new insights into protein motion and dynamics. To accommodate the MHz repetition rate, sample delivery methods are in demand which...
X-ray emission spectroscopy (XES) complements structural techniques like serial femtosecond crystallography (SFX) by providing insights into the electronic states at specific sites within a sample. At X-ray free-electron lasers, simultaneous SFX and XES measurements using a single pulse have already been performed — such as the determination of the oxidation states in metalloproteins. By using...
TRISS (TRapped Ion Spectrometer Setup) is a novel experimental station at the MAX IV synchrotron facility, designed to investigate fundamental molecular processes relevant to biomolecular and biochemical physics. TRISS uses an electrospray ionization (ESI) source to create ions, which are then fragmented using photons, electrons, or gas. The TRISS setup combines a segmented linear ion trap...
Diffractive single-particle imaging (SPI) using X-ray free-electron
lasers (XFELs) offers a promising approach for determining protein
structures without crystallization (Neutze2000) . For successful
reconstruction thousands of diffraction images of individual proteins
have to be assembled. It has been shown with molecular dynamics
simulations that proteins carrying a dipole moment can be...
Serial macromolecular crystallography (SMX) has emerged as a transformative technique enabling measurements across expansive parameter spaces in structural biology. Here I will talk about the evolution of serial crystallography techniques, particularly focusing on developments for time-resolved, pH-responsive, and temperature-controlled experiments and their application in unraveling dynamic...
Fourier transform infrared (FTIR) (micro-)spectroscopy is a label-free and non-destructive vibrational tool that has been successfully applied to study not only the amyloid structural properties and aggregation mechanisms directly in cells, tissues, or biofluids, but also to gain new insights into the mechanisms of amyloid formation and toxicity[1]. In particular, the use of an infrared...
Ultrafast multidimensional IR spectroscopy (2D-IR) can provide detailed information on local structural dynamics in peptide, proteins or complex systems like models of protein condensates. 2D-IR is a femtosecond laser spectroscopy with its intrinsic time-resolution on par with the fundamental timescale of chemical dynamics, fs to ps. For biological systems the capability to directly probe...
