Biology subjects

Mathieu, E.

Publications and source records attributed to Mathieu, E..

5 recordsLinked to original sources

bioRxiv2026

Time-resolved functional rotation crystallography reveals protein dynamics and catalysis

Time-resolved crystallography offers a window into the transient, atomic-scale structural changes that underlie biological function. Here we introduce Time-Resolved Functional Rotation Crystallography (TR-FRX), a method that enables molecular-level studies of ligand binding and enzymatic catalysis using single protein crystals at room temperature. Unlike many state-of-the-art time-resolved methodologies, TR-FRX relies on standard rotation-based X-ray data collection and does not require serial sample delivery strategies. By dispensing nanoliter-scale droplets of ligand or substrate directly onto crystals mounted in conventional holders, TR-FRX captures real-time structural snapshots while maintaining experimental accessibility. As a proof-of-concept, we first monitor the binding of N-acetylglucosamine to hen egg-white lysozyme, demonstrating that TR-FRX can reveal ligand recognition in crystallo on sub-second timescales. We then resolve the bidirectional catalytic mechanism of a prototypical enzyme from the tricarboxylic-acid-cycle, revealing sequential cofactor and substrate binding, catalytic loop dynamics, and substrate stabilization across time points spanning from 137 milliseconds to minutes. Requiring only micrograms of protein and standard beamline infrastructure, TR-FRX provides an accessible methodology for capturing transient enzymatic states and enables 100-ms timescale studies at room temperature on single protein crystals.

biochemistry↗

bioRxiv2025

The histone methyltransferase SDG26 shapes cold stress responses in Arabidopsis through chromatin-based regulation of ABA-dependent and ABA-independent pathways

Plants constantly face adverse environmental conditions, including temperature drops that can severely impair growth and productivity. To cope with such stresses, they have evolved complex mechanisms of transcriptional reprogramming. While various cold-responsive pathways have been described, the contribution of chromatin-level regulation, and in particular histone modifications, remains largely obscure. Here, we identify the histone methyltransferase SET DOMAIN GROUP 26 (SDG26) as a positive regulator of cold stress responses in Arabidopsis thaliana. We show that SDG26 is transcriptionally induced and post-transcriptionally stabilized by cold, and that its loss of function leads to increased freezing tolerance but reduced drought tolerance. At the molecular level, SDG26 promotes expression of cold-responsive genes, including members of the CBF-COR regulon, through direct binding and histone H3 lysine 36 trimethylation (H3K36me3) at their chromatin. Concomitantly, SDG26 modulates abscisic acid (ABA) biosynthesis, catabolism, and transport, thereby promoting ABA accumulation, stomatal closure, and drought tolerance. Collectively, our results reveal that SDG26 integrates ABA-dependent and ABA-independent pathways to fine-tune Arabidopsis responses to abiotic stresses. We further establish SDG26 as a chromatin modifier contributing to stress-responsive H3K36me3 enrichment at specific loci. Together, our work identifies SDG26 as a chromatin-based hub balancing cold acclimation with water conservation, thereby enhancing plant resilience.

plant biology↗

bioRxiv2023

A redox switch allows binding of ferrous and ferric ions in the cyanobacterial iron binding protein FutA from Prochlorococcus

The marine cyanobacterium Prochlorococcus is a main contributor to global photosynthesis, whilst being limited by iron availability. Cyanobacterial genomes typically encode two different types of FutA iron binding proteins: periplasmic FutA2 ABC transporter subunits bind Fe(III), while cytosolic FutA1 binds Fe(II). Owing to their small size and their economized genome Prochlorococcus ecotypes typically possess a single futA gene. How the encoded FutA protein might bind different Fe oxidation states was previously unknown. Here we use structural biology techniques at room temperature to probe the dynamic behavior of FutA. Neutron diffraction confirmed four negatively charged tyrosinates, that together with a neutral water molecule coordinate iron in trigonal bipyramidal geometry. Positioning of the positively charged Arg103 side chain in the second coordination shell yields an overall charge-neutral Fe(III) binding state in structures determined by neutron diffraction and serial femtosecond crystallography. Conventional rotation X-ray crystallography using a home source revealed X-ray induced photoreduction of the iron center with observation of the Fe(II) binding state; here, an additional positioning of the Arg203 side chain in the second coordination shell maintained an overall charge neutral Fe(II) binding site. Dose series using serial synchrotron crystallography and an XFEL X-ray pump-probe approach capture the transition between Fe(III) and Fe(II) states, revealing how Arg203 operates as a switch to accommodate the different iron oxidation states. This switching ability of the Prochlorococcus FutA protein may reflect ecological adaptation by genome streamlining and loss of specialized FutA proteins. Significance StatementOceanic primary production by marine cyanobacteria is a main contributor to carbon and nitrogen fixation. Prochlorococcus is the most abundant photosynthetic organism on Earth, with an annual carbon fixation comparable to the net global primary production from agriculture. Its remarkable ecological success is based on the ability to thrive in low nutrient waters. To manage iron limitation, Prochlorococcus possesses the FutA protein for iron uptake and homeostasis. We reveal a molecular switch in the FutA protein that allows it to accommodate binding of iron in either the Fe(III) or Fe(II) state using structural biology techniques at room temperature and provide a plausible mechanism for iron binding promiscuity.

biophysics↗

bioRxiv2022

Broadly applicable and accurate protein design by integrating structure prediction networks and diffusion generative models

There has been considerable recent progress in designing new proteins using deep learning methods1-9. Despite this progress, a general deep learning framework for protein design that enables solution of a wide range of design challenges, including de novo binder design and design of higher order symmetric architectures, has yet to be described. Diffusion models10,11 have had considerable success in image and language generative modeling but limited success when applied to protein modeling, likely due to the complexity of protein backbone geometry and sequence-structure relationships. Here we show that by fine tuning the RoseTTAFold structure prediction network on protein structure denoising tasks, we obtain a generative model of protein backbones that achieves outstanding performance on unconditional and topology-constrained protein monomer design, protein binder design, symmetric oligomer design, enzyme active site scaffolding, and symmetric motif scaffolding for therapeutic and metal-binding protein design. We demonstrate the power and generality of the method, called RoseTTAFold Diffusion (RFdiffusion), by experimentally characterizing the structures and functions of hundreds of new designs. In a manner analogous to networks which produce images from user-specified inputs, RFdiffusion enables the design of diverse, complex, functional proteins from simple molecular specifications.

biochemistry↗

bioRxiv2020

The microbiota plays a critical role in the reactivity of lung immune components to innate ligands

The microbiota contributes to shaping efficient and safe immune defenses in the gut. However, little is known about the role of the microbiota in the education of pulmonary innate immune responses. Here, we tested whether the endogenous microbiota can modulate reactivity of pulmonary tissue to pathogen stimuli by comparing the response of specific pathogen-free (SPF) and germ-free (GF) mice. Using SPF and GF mice intranasally exposed to lipopolysaccharide (LPS), a component of Gram-negative bacteria, we observed earlier and greater inflammation in the pulmonary compartment of GF mice than that of SPF mice. Toll-like receptor 4 (TLR4) was more abundantly expressed in the lungs of GF mice than those of SPF mice at steady state, which could predispose the innate immunity of GF mice to strongly react to environmental stimuli. Lung explants were stimulated with different TLR agonists or infected with the human airways pathogen, respiratory syncytial virus (RSV), resulting in greater inflammation under almost all conditions for the GF explants. Finally, alveolar macrophages (AM) from GF mice presented a higher innate immune response upon RSV infection than those of SPF mice. Overall, these data suggest that the presence of microbiota in SPF mice induced a process of innate immune tolerance in the lungs by a mechanism which remains to be elucidated. Our study represents a step forward to establishing the link between the microbiota and the immune reactivity of the lungs. Plain Language summaryMicrobiota represents an important partner of immunologic system at the interface between immune cells and epithelium. It is well known, notably in the gut, that the microbiota contributes in shaping efficient and safe defenses. However, little is known about the role of the microbiota in the education of pulmonary innate immune responses. In this study, we postulate that endogenous microbiota could dampen an excessive reactivity of pulmonary tissue to external stimuli. Thus, we sought to study the innate immune reaction switched on by viral or bacterial ligands in respiratory tract cells coming from mice with or without microbiota (germ-free condition, GF). Altogether, our results show a higher inflammatory reaction in GF condition. This study represents a step forward to better establish the link between the microbiota and the reactivity of the lung tissue. Not only these data demonstrate that the microbiota educates the pulmonary innate immune system, but also contributes the emerging concept of using respiratory commensal bacteria as potential next-generation probiotics to prevent susceptibility to respiratory diseases.

immunology↗