BIOPHYMS · Mass spectrometry for nucleic acid biophysics : dealing with diversity

Mass spectrometry for nucleic acid biophysics : dealing with diversity

The Career Integration Grant BIOPHYMS helped kick-start the new research team of Dr. Valérie Gabelica (originally from Belgium) at the Institut Européen de Chimie et Biologie and the ARNA laboratory in Bordeaux, France. The team’s website can be found at: http://www.iecb.u-bordeaux.fr/teams/GABELICA/ The aim of the BIOPHYMS project was to develop native mass spectrometry and ion mobility spectrometry as biophysical tools to probe small molecule (ligand) interactions with G4 structures. There is now increasing evidence that specific nucleic acid structures modulate gene expression levels both at the transcriptional and at the translational level. In particular, G-quadruplex (G4) nucleic acid structures are attractive targets for anticancer strategies, since several studies showed that their stabilization by ligands caused proliferation arrest, telomere deprotection and changes in gene expression. Understanding the structure-function relationships in G4 DNA and RNA requires innovative biophysical tools to probe the structures adopted by a wide variety of sequences, their macromolecular assemblies, and their interactions with small molecules. The objective is to characterize not only the binding affinity and specificity but also the binding mode of ligands for a variety of DNA and RNA targets. In the first part of the project, we obtained G-quadruplex spectra from potassium-containing solutions (potassium is the physiologically relevant cation). We have now exploited the approach to study the thermodynamics and kinetics of potassium binding to G-quadruplexes, and to study the effects of several ligands on G-quadruplexes. The potassium and ligand count on each oligonucleotide, made possible by mass spectrometry, led us to characterize unprecedented ligand binding modes. This was the object of the PhD thesis of Mr. Adrien Marchand, who graduated on November 29, 2016 and contributed to 6 published articles. Ion mobility spectrometry is crucial for studying the conformational adaptability of the target, and ligand-induced conformational changes. The acquisition of a new-generation drift tube ion mobility spectrometer coupled to a Q-TOF mass spectrometer was made during the first grant period. The drift tube was modified to enable the determination of collision cross section values in helium, a gas in which the calculation of collision cross sections for model structures is possible in an accurate manner. We have established both the experimental and theoretical workflow and tools to measure and calculate CCSs in helium (published in an open access tutorial in the Journal of Mass Spectrometry in May 2015). The team recently devised new ways to process the data. We also contribute to devising new standard operating procedures and reporting guidelines for ion mobility spectrometry data, within the European COST action BM1403. The modified ion mobility instrumentat and know-how are made accessible to the community through the Structural Biophysico-Chemistry platform of the IECB (Institut Européen de Chimie et Biologie) in Bordeaux, where the grantee is located, and several collaborations have emerged. As anticipated, the biophysical approaches developed here for a specific purpose (G4 ligands) are therefore also being applicable to other targets (nucleic acids and others). Seven additional publications emerged from all the collaborations made possible by the Career Integration Grant. The Career Integration Grant BIOPHYMS greatly contributed in the integration of Dr. Valérie Gabelica in France. She has obtained a permanent position as research director of the Inserm in December 2013, affiliated to the Mixed Research Unit ARNA (Inserm U1212, CNRS UMR5320, Université de Bordeaux) as an independent group leader. She is well integrated at the local level, and member of the laboratory council and of the University of Bordeaux, Department of Life Sciences Research Council where she was confided several missions. She has since obtained two other major European grants (ERC, ITN), and at the term of the project her team includes 2 PhD students, 1 engineer and 3 post-docs. One PhD student (Adrien Marchand) has graduated and is now post-doc at ETH Zurich. Five post-docs trained in the group have made their transition to academia or industry. The PI also hosted six foreign PhD students and one independent scientist for short training visits, and eleven undergraduate students for internships. She also participated to dissemination activities to a broader public, with for example an invited talk at a ministerial event, or an interview in a very popular local newspaper (Sud Ouest). She is also fully integrated in the scientific community in France: for example she served three years in the board of the French Society for Mass Spectrometry and organized the national conference in 2016.

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