Biology subjects

Burghammer, M.

Publications and source records attributed to Burghammer, M..

8 recordsLinked to original sources

bioRxiv2026

In situ radiation damage tracking during scanning small-angle X-ray scattering

Due to the high penetration power of X-rays, X-ray imaging techniques have been employed to image cellular architecture. Among these techniques, scanning small-angle X-ray scattering (SAXS) is a powerful method for investigating ordered structures within cells. With the recent advent of high-brilliance synchrotron sources, nano-focused X-rays and fast scanning modes, scanning SAXS can now provide structural information from hydrated cells in liquid environments in little time. However, the high photon flux results in an increased dose and pronounced radiation damage to the intracellular structures. Therefore, strategies are required to monitor and quantitatively analyze the extent of damage. Here, we present a multimodal approach that employs an on-axis, high-speed, brightfield microscope to monitor cellular damage during scanning. This microscope is integrated into the scanning SAXS setup at ID13 of ESRF and records images in synchrony with SAXS data acquisition. We are able to observe and quantitatively track changes in cellular morphology \textit{in situ} and thereby obtain complementary information to that provided by scanning SAXS measurements. Our approach opens up the potential for a multifaceted investigation of radiation damage to biological cells and other sample types.

biophysics↗

bioRxiv2026

Correlative X-ray imaging and fluorescence microscopy

Imaging the structural organization inside cells in their native state is essential for understanding how the arrangement and interactions among molecular components give rise to biological function. Fluorescence microscopy is one of the pivotal techniques that provides molecular specificity for imaging in real space, however, the technique is limited to labeled components. X-rays, on the contrary, are sensitive to electron density contrast and therefore to label-free samples, and probe structure in reciprocal space. In particular, scanning small-angle X-ray scattering (SAXS) combines information from real and reciprocal space and enables access to intact cells, owing to the high penetration power of the X-rays. Combining both imaging modalities in a synergistic manner promises powerful tools for cellular imaging, but remains challenging, because of the differing requirements the complementary methods introduce. Here we present a correlative imaging platform that integrates a modular, compact and beamline-compatible fluorescence microscope with scanning SAXS, to enable fast sequential imaging of the identical cellular regions. We developed a dedicated microfluidics flow chamber enabling measurements under hydrated, near-native conditions. We demonstrate the utility of our methodology by investigating two different relevant cellular components, i.e., thick keratin bundles in epithelial cells that contribute to cell mechanics, and force-generating actomyosin in cardiomyocytes. Employing adapted data analysis methods, we find a good agreement between the fluorescence-derived and the SAXS-derived orientation maps. This result demonstrates that the label-free approach with SAXS captures cytoskeletal organization through-out the cell, and can be directly linked to specific molecular information provided by the complementary fluorescence imaging, in a physiologically relevant cellular environment. Our work establishes a general strategy for multimodal imaging of cellular architecture and opens ways to investigate living cells under the influence of drugs and chemical manipulation experiments.

biophysics↗

bioRxiv2025

Fast scanning small angle X-ray scattering of hydrated biological cells

Due to their high penetration depth, X-rays enable us to obtain information from the interior of whole, unsliced cells. Scanning small angle X-ray scattering (SAXS), in particular, reveals real-space images in dark field representation as well as structural information in reciprocal space. However, obtaining information on anisotropy and orientation from cells in an aqueous, close-to-physiological environment remains challenging. Here, we extend the recently introduced fast scanning SAXS mode with short exposure times of few milliseconds to such hydrated samples by combining a newly developed, X-ray compatible microfluidic sample chamber and innovative data analysis that includes an effective noise-filtering method. This strategy enables the systematic analysis of radiation damage by quantifying the SAXS signal. Our results demonstrate that scanning SAXS can be used to obtain intracellular information of fixed-hydrated cells and the approach may in the future be applicable to living cells as well. SynopsisFast scanning SAXS on biological cells in aqueous environment reveals intracellular anisotropy and orientation and allows for systematic assessment of radiation damage caused by the measurements.

biophysics↗

bioRxiv2025

Cement lines are stiffer and harder than bone but exhibit different mineral-mechanics relationships due to thicker and shorter mineral particles

The remarkable mechanical performance of bone arises from its complex hierarchical structure and from the presence of numerous internal interfaces joining the different components. Formed during bone remodeling, the cement line (CL) is a thin interface surrounding osteons. Although often neglected due to its small size, the CL has been suggested to play an important role in bone fracture toughness. However, there is an ongoing debate about its specific structure and mechanical behavior. Here, we investigate the composition-mechanics relationship at the CL and surrounding bone in human osteons using multiple methods in a correlative manner. We found that the CL is stiffer and harder than adjacent osteonal tissue. However, the CL requires more mineral than bone to attain the same stiffness and hardness. Analyzing the nanoscale properties of the mineral, we found thicker but shorter particles at the CL. Using a mechanical model, we interpreted the lower aspect ratio of the mineral particles as a less effective way to reinforce the collagen matrix. Casting our findings into a computational model, we questioned the possible protective role of the CL, which is not a soft interface as traditionally believed.

bioengineering↗

bioRxiv2024

Molecular Polymorphism of tau aggregates in Pick's disease

Tau protein plays a central role in many neuropathies. The trajectory by which tau spreads through neural networks is disease-specific but the events driving progression are unknown. This is due in part to the challenge of characterizing tau aggregates in situ. We address that challenge using in situ micro-x-ray diffraction ({micro}XRD) and micro-X-ray fluorescence (XRF) to examine tau lesions in the brain of a 79-year-old male with dementia. Neuropathological examination revealed classical forms of tau in the hippocampal formation: extensive Pick bodies in the granular layer; modest numbers of neurofibrillary tangles and dystrophic neurites in the CA4 and hilus. {micro}XRD indicated that Pick bodies are low in fibril content, whereas neurofibrillary lesions within adjacent tissue exhibit far greater density of fibrillar tau. XRF demonstrated elevated levels of zinc, calcium and phosphorous in all tau-containing lesions whereas sulfur deposition was greatest in lesions exhibiting high fibrillar content. Correlation of lesion morphology with anatomical localization, tau fibrillation and differential elemental accumulation suggests tau fibrils generate biochemically distinct microenvironments that influence lesion morphology, tau seed formation and spreading.

neuroscience↗

bioRxiv2021

Spores and extracellular matrix components impart molecular order in Bacillus subtilis biofilms.

Biofilms are multicellular microbial communities that encase themselves in an extracellular matrix (ECM) of secreted biopolymers and attach to surfaces and interfaces. Bacterial biofilms are detrimental in hospital and industrial settings, but they can be beneficial in agricultural contexts. An essential property of biofilms that grants them with increased survival relative to planktonic cells is phenotypic heterogeneity; the division of the biofilm population into functionally distinct subgroups of cells. Phenotypic heterogeneity in biofilms can be traced to the cellular level, however, the molecular structures and elemental distribution across whole biofilms as well as possible linkages between them remain unexplored. Mapping X-ray diffraction (XRD) across intact biofilms in time and space, we revealed the dominant structural features in Bacillus subtilis biofilms, stemming from matrix components, spores and water. By simultaneously following the X-ray fluorescence (XRF) signal of biofilms and isolated matrix components, we discovered that the ECM preferentially binds calcium ions over other metal ions, specifically, zinc, manganese and iron. These ions, remaining free to flow below macroscopic wrinkles that act as water channels, eventually accumulate and lead to sporulation. The possible link between ECM properties, regulation of metal ion distribution and sporulation across whole intact biofilms unravels the importance of molecular-level heterogeneity in shaping biofilm physiology and development. Significance StatementBiofilms are multicellular soft microbial communities that are able to colonize synthetic surfaces as well as living organisms. To survive sudden environmental changes and efficiently share their common resources, cells in a biofilm divide into subgroups with distinct functions, leading to phenotypic heterogeneity. Here, by studying intact biofilms by synchrotron X-ray diffraction and fluorescence, we revealed correlations between biofilm macroscopic architectural heterogeneity and the spatio-temporal distribution of extracellular matrix, spores, water and metal ions. Our findings demonstrate that biofilm heterogeneity is not only affected by local genetic expression and cellular differentiation, but also by passive effects resulting from the physicochemical properties of the molecules secreted by the cells, leading to differential distribution of nutrients that propagates through macroscopic length scales.

microbiology↗

bioRxiv2019

Myelin Basic Protein dynamics from out-of-equilibrium functional state to degraded state in myelin

Living matter is a quasi-stationary out-of-equilibrium system; in this physical condition, structural fluctuations at nano- and meso-scales are needed to understand the physics behind its biological functionality. Myelin has a simple ultrastructure whose fluctuations show correlated disorder in its functional out-of-equilibrium state. However, there is no information on the relationship between this correlated disorder and the dynamics of the intrinsically disordered Myelin Basic Protein (MBP) which is expected to influence the membrane structure and overall functionality. In this work, we have investigated the role of this protein structural dynamics in the myelin ultrastructure fluctuations in and out-of-equilibrium conditions, by using synchrotron Scanning micro X Ray Diffraction and Small Angle X ray Scattering. We have induced the crossover from out-of-equilibrium functional state to in-equilibrium degeneration changing the pH far away from physiological condition. While the observed compression of the cytosolic layer thickness probes the unfolding of the P2 protein and of the cytoplasmic P0 domain (P0cyt), the intrinsic large MBP fluctuations preserve the cytosol structure also in the degraded state. Thus, the transition of myelin ultrastructure from correlated to uncorrelated disordered state, is significantly affected by the unfolding of the P2 and P0 proteins, which in this latter state do not act in synergistic manner with MBP to determine the membrane functionality.\n\nSTATEMENT OF SIGNIFICANCEA better comprehension of myelin degenerative process and the role of protein dynamics in this biological membrane is a topic issue in todays scientific community. The myelin ultrastructural fluctuations exhibit correlated disorder in its functional state, that becomes uncorrelated as it degenerates. In this work we elucidate the interplay of protein structural dynamics and myelin ultrastructure in the transition from its functional state to the degraded state. The results highlight that the intrinsically disordered Myelin Basic Protein (MBP) allows to preserve the myelin structure following both the small correlated fluctuations in physiological state and the large disordered fluctuations in degraded conditions, where the myelin functionality is close to being lost and the MBP remains the single active protein.

biophysics↗

bioRxiv2017

Spatio-temporal elasticity patterns in extracellular matrix during Hydra morphogenesis

Albeit ample evidence has suggested the remodeling of extracellular matrix (ECM) in animals plays crucial roles in development and diseases, little is understood how ECM mechanics correlates with tissue morphogenesis. In this study, we quantitatively determined how spatio-temporal elasticity patterns in ECM change during the asexual reproduction of freshwater polyp Hydra. We first determined the mesoscopic protein arrangement in Hydra ECM (mesoglea) by grazing-incidence small-angle X-ray scattering with nano-beam (nano-GISAXS). Our data unraveled fibrillar type I collagen in Hydra mesoglea (Hcol-I) takes an anisotropic, more strongly distorted hexagonal lattice compared to those in vertebrates that could be attributed to the lower proline content and lack of lysin-crosslinks in Hcol-1 fibers. Then, we "mapped" the spatio-temporal changes in ECM stiffness ex vivo with aid of nano-indentation. We identified three representative elasticity patterns during tissue growth along the oral-aboral body axis of the animals. Our complementary proteome analysis demonstrated that the elasticity patterns of the ECM correlate with a gradient like distribution of proteases. Perturbations of the oral Wnt/{beta}-catenin signaling center further indicated that ECM elasticity patterns are governed by Wnt/{beta}-catenin signaling. The ex vivo biomechanical phenotyping of Hydra mesoglea established in this study will help us gain comprehensive insights into the spatio-temporal coordination of biochemical and biomechanical cues in tissue morphogenesis in vivo.

developmental biology↗