PhD in Structural Biology (H/F)

The thesis work will be carried out in the Laboratory of Applied Biology and Pharmacology (LBPA UMR 8113)CNRS / ENS de Paris-Saclay 4 avenue des sciences 91190 Gif-sur-Yvette (France).
The LBPA is a joint research unit (CNRS - École Normale Supérieure Paris-Saclay) based on the new campus of ENS Paris-Saclay in Gif-sur-Yvette, in the heart of the University of Paris-Saclay. This multi/inter-disciplinary laboratory is specialized in the analysis of interactions between biomolecules and conducts studies in the fields of structural biology, biophysics, microbiology, and anti-viral pharmacology.
The candidate will work in the team "structure, function and bioengineering of non-conventional nucleic acids" under the direct supervision of Dr. Brahim Heddi.
This theme will be conducted in collaboration with a team at the University of Strasbourg (Université de Strasbourg/LBP) in the framework of an ANR project.

G quadruplexes (GQs) are non-canonical nucleic acid structures that consist of at least two layers of G quartets held together by stacking interactions, Hoogsteen bonds, and cation binding. GQ forming sequences are widely distributed in mRNAs and non-coding RNAs, where they regulate gene expression. In vivo, RNA GQs are thought to be largely unfolded, due to the activity of RNA binding proteins (RBP). Therefore, RNA GQs are probably transient structures converted by RBPs between their folded and unfolded states. Fluorescence techniques are well-suited to explore their structure and dynamics as well as their interaction with proteins over a wide time range and at low concentrations, but suffering from the need of introducing external labels. Numerous fluorescent purine surrogates have been developed, but they generally destabilize the GQ structure or are highly quenched. In this context, our objective is to validate and apply tzG, an isomorphic fluorescent analog of G from the isothiazolo[4,3-d]-pyrimidine family developed by our collaborator as a unique tool to site-specifically characterize the conformations, dynamics, and molecular interactions of RNA GQs. We will first characterize the effect of tzG on GQ topology and stability CD, thermal melting experiments, and NMR experiments. For sequences with well-resolved NMR spectra, the 3D structure will be then determined, to obtain fine structural details of the impact of tzG insertion in GQ structures. We will validate tzG as a key tool for RNA GQs by applying it to decipher the GQ unfolding mechanism of DHX36 and CNBP proteins, through a combination of CD and NMR while our collaborator will use fluorescence spectroscopy and stopped-flow technique. We expect to monitor site-specifically the initial and late events of the unfolding/refolding mechanism. The project gathered an international multidisciplinary consortium of recognized experts in their fields, including synthetic chemistry, fluorescence, and simulation using MD/QM approach. Through this study, tzG should become the first tool that can faithfully monitor any substituted G in RNA GQs and thus, lead to breakthroughs in the understanding of GQ properties and RBP mechanisms of action.