In a context of population aging, discovery and validation of novel oxidative stress biomarkers for screening of neurodegenerative diseases is a key issue. However, this remains a challenge for low concentrated biomarkers. This arises from the high rate of false positives during the discovery phase, due to the complexity and concentration dynamic range of the samples. To improve the detection specificity toward the oxidized proteins, we propose an innovative experimental setup made of a mass spectrometer coupled with visible laser induced dissociation (LID) to add a stringent optical specificity to the mass selectivity. Since peptides do not naturally absorb in the visible range, this novel methodological approach relies on the proper chemical derivatization of oxidized Cys and carbonyl groups with a chromophore. Only the subset of derivatized peptides will be specifically fragmented in LID, improving the consistency of oxidized protein biomarkers quantification in biological samples.

The DyCTheMS project proposes a new experimental approach to the conformational dynamics and thermodynamics of micro-solvated biomolecules, based on ion mobility spectrometry (IMS) and mass spectrometry (MS). The thermodynamics of structural transitions in those systems will be probed using an original calorimetry technique based on IMS measurements. Complementary temperature-dependent isomerization dynamics data will also be collected from tandem-IMS measurements. An originality of the proposal lies in the attempt to bridge the gap between solution and gas-phase measurements by probing how the conformational landscape of a molecular system is modified upon solvation. In particular we will shed light on the extent of solvation required for a biomolecule to exhibit “native” conformational properties. After a validation phase on well-characterized systems, the above procedures will be used to gain insight in the binding-induced structuration of an intrinsically disordered peptide.

Aquatic ecosystems are persistently exposed to environmental stressors such as chemical micropollutants from natural environment or anthropogenic activities. These chemical contaminations may result in alterations of the internal biochemical homeostasis of the aquatic organisms, noticeable at the omics scale. The major limitation in the mechanistic knowledge of environmental chemical toxicity effects on aquatic organisms is the absence of molecular information notably at the genome wide scale in environmentally relevant species. The “omics” technologies – Transcriptomics, Proteomics, Lipidomics, Metabolomics to name a few – offer great promises to help to elucidate molecular responses to exposures in aquatic organisms during specific and vulnerable life cycle stages. Lipid metabolism is the major fundamental metabolic pathway producing energy in animals. In crustacean, lipids play a pivotal role in vulnerable stages like molting, reproduction, development. Recently, it has been shown that chemical compounds interfering with lipid metabolism, recognized as obesogens like tributyltin, mislead the distribution and the synthesis of lipids in the non-target and model organism Daphnia magna. Moreover, pharmaceutical drugs used for hypercholesterolemia to lower cholesterol and triglycerides concentrations like pravastatin or bezafibrate have been detected in sewages. Another study showed that simvastatin exposure in the amphipod Gammarus locusta disturbed the reproduction and population growth at the ng/L range. To understand and predict the effects of toxic exposures, it is crucial to identify the affected metabolic networks. Statins are among the most broadly used pharmaceuticals worldwide and is therefore of emerging environmental concerns. This medication inhibits selectively the enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA reductase) involved in the synthesis of mevalonate, a precursor of sterols including cholesterol. We hypothesize that lipid metabolism may turn out to be the drug-induced toxicity target in aquatic species. However, we are facing a lack of crucial knowledge about the relationships between the abundance of lipid species, the activation/inhibition of signaling and/or biosynthesis pathways and the observed phenotypes. The project PLAN-TOX aim to gain a mechanistic understanding of toxic effects of hypolidiaemia drugs (statins and fibrates) on the sentinel organism G. fossarum. We are proposing to develop and apply an innovative multi-omics approach in ecotoxicology, including (i) the functional proteome and lipidome mapping in the sentinel organism G. fossarum before and after exposure, (ii) the development of methods for the integrative analysis of multi-omics data. G. fossarum is a widely developed sentinel species in ecotoxicology or environmental monitoring and is one of the ecologically relevant species for which we can evaluate the toxicity on different development stages and reproductive cycles. This project will allow to better understand molecular plasticity of energetic metabolism and to identify proteins and lipids involved in physiological changes/responses related to exposure.

Molecular interactions play a key role in all the branch of analytical sciences. Among these interactions, those involving proteins are particularly fascinating as they comprise a large number of degree of freedom and are present in all biological processes. Indeed, a key feature defining protein function in living cells is their capacity to selectively interact with other molecules in their environment. Combined in diverse cellular assemblies that vary with cell cycle, tissue type and in response to external stimuli, proteins constitute the nanomachines of biology. Understanding how proteins move and interact with neighboring molecules is necessary for solving critical puzzles in molecular biology. Filling the sizeable gaps in our knowledge of protein/protein interactions and protein function (or malfunction) is increasingly crucial for the design of potential new drugs and to better understand biological processes leading to specific diseases. These opportunities can be fully exploited only if technological progress is paired with a major step forward in our understanding of the detailed physical and chemical factors regulating protein mechanisms (encompassing dynamics, folding, interactions, and assembly) and consequently in our ability to model and predict these processes. While protein/small molecules interactions are now routinely modeled by molecular dynamics to extract thermodynamical parameters, the study of protein/protein interactions at the molecular level still poses outstanding challenges both for theory and experiment. No single technique can at present span the whole range of typical time and length scales relevant for a protein biological function and interaction. Nevertheless, qualitative and quantitative analyses of these interactions is possible at microscopic and atomic resolutions using Nuclear Magnetic Resonance (NMR) with complementary biophysical methods like Isothermal Titration Calorimetry (ITC) or Small Angle X-ray/neutron scattering (SAXS/SANS). At this level, NMR is especially useful as it can probe dynamical events on a broad time scale and is highly sensitive to processes relevant to biomacromolecules, such as inter-domain motions, exchange equilibria and catalysis. In our quest to see the invisible, new theoretical and computational approaches need to be developed in order to address the current challenge. It is our belief that this goal can be achieved by reconciling biological and biochemical approaches with a physical and mathematical perspective of the problem. Our project is located at the frontier of analytical sciences, biochemistry as well as computational science and will merge researchers of different fields. We do not propose another additional tool to derive structures guided by experimental data. Conversely, we propose to develop a novel general-purpose integrated analytical tool that combines both experimental and molecular dynamics simulation approaches for a better understanding of protein/protein interactions by modeling in the real time these interactions and binding pathways to get access to thermodynamic parameters. Far away from the dedicated national infrastructures that request a huge amount of power and cooling fluids, we propose to quantitatively study protein/protein interactions on everybody's office with an affordable price equipment and to integrate experimental data to accelerate this process. Protein/protein interactions will be right at your fingertips.

Antibiotic resistance is an increasing global public health threat that concerns all major bacterial pathogens and therapeutic drugs. Antimicrobial peptides are an important component of the first line of defense of most living organisms against invading bacteria and are considered as promising alternative therapeutics to fight pathogens. However, bacteria have also evolved mechanisms to resist antimicrobial peptides, one of the most prominent being the use of dedicated ABC (“ATP-Binding Cassette”) transporters. A unique property of these transporters is that they cooperate with two-component regulatory systems to sense the presence of antimicrobial peptides and, in turn, activate the expression of the transporter genes thereby increasing the levels of cellular resistance. The functioning of these unique sensor/transport systems is, however, poorly understood and we propose here to elucidate the molecular mechanism that governs such a resistant module in Streptococcus pneumoniae, a deadly human pathogen.