Missions

The main objective of the project is the discovery of active molecules for the treatment of Friedreich's ataxia (FA), a neurodegenerative and cardiac disease. AF is the most common recessive ataxia, it is caused by a defect in the expression of frataxin, a mitochondrial protein involved in the assembly of iron-sulfur clusters. Iron-sulfur (Fe-S) clusters are protein cofactors providing catalytic activities to a multitude of enzymes involved in essential biological processes such as ATP production, Krebs cycle, protein synthesis, immunity, signaling and maintenance of genome integrity. Their synthesis is carried out in the mitochondria by a multi-protein complex via a sequential process during which frataxin acts as an accelerator. The lack of expression of frataxin therefore leads to a deficit in the synthesis of Fe-S clusters, which leads to a multitude of cellular defects having pleiotropic effects on the organism (ataxia, cardiac hypertrophy, diabetes, etc.). The objective of the project is to target the primary defect of the disease by discovery of molecules replacing the frataxin protein (at least partially). For this, the laboratory has developed a high-throughput screening system using the human Fe-S cluster assembly machinery reconstituted in vitro with recombinant proteins. We have developed virtual in silico screening in order to preselect molecules from different chemical libraries according to their probability of interaction with the Fe-S cluster biosynthetic machinery at the frataxin binding site. The mission of the post-doctoral fellow will be to carry out analyzes by virtual screening and high-throughput screening tests on the preselected molecules. The active molecules will then be validated by various specific enzymatic tests in order to eliminate false positives. Then structure-function analyzes by NMR, EPR and cryo-EM will be carried out in order to study the mode of action of the molecules of interest in order to improve their activities by targeted chemical modifications. In parallel, in vivo tests will be carried out in Drosophila FA models.

Activités

The post-doctoral fellow will be in charge of studies by virtual screening, production of recombinant proteins, reconstitution of complexes and high-throughput screening in multi-well plates. These analyzes will be carried out under anaerobic conditions in a dedicated chamber to preserve the oxidation state of the iron. The post-doctoral fellow will be in charge of interactions with the chemical libraries and the affiliated laboratories providing the different molecules. He/she will carry out enzymatic tests (assembly of Fe-S clusters, sulfur transfer) in order to validate the activity of the molecules of interest identified by high-throughput screening. The post-doctoral fellow will work in close collaboration with the University of Paris-Cité to carry out in vivo tests in Drosophila models of FA. He/she will be in charge of spectroscopic and structural studies (NMR, EPR, cryo-EM, X-ray crystallography) on the reconstituted system, in collaboration with project partner laboratories in Gif-Sur-Yvette, Grenoble and Marseille, in order to determine the mode of action of the candidate molecules and to be able to define the modifications to be implemented to improve their activity.

Compétences

Strong technical skills in the biochemistry of recombinant proteins are expected (production, purification and handling). Technical or analytical skills in NMR, EPR, cryo-EM and crystallography will be an asset. A past experience in the study of metalloproteins and redox system would be a plus. A very great rigor in the execution of the experiments will be required, necessary to carry out the screening tests as well as a very strong motivation for this type of project.

Contexte de travail

The I2BC is a leading institute in biology, biochemistry and biophysics. It is located in the southern region of Paris on the campus of the Paris-Saclay University and brings together cell biologists, biochemists and biophysicists (between 600 and 700 people). Our team has been studying the redox biology and assembly mechanisms of Fe-S clusters for many years and is specialized in in vitro reconstitution approaches of complex systems as well as structural and spectroscopic analyses.

Contraintes et risques

No constraint or particular risk