A User-Friendly DNA Modeling Software for the Interpretation of Cryo-Electron Microscopy Data.

The structural modeling of a macromolecular machine is like a "Lego" approach that is challenged when blocks, like proteins imported from the Protein Data Bank, are to be assembled with an element adopting a serpentine shape, such as DNA templates. DNA must then be built ex nihilo, but modeling approaches are either not user-friendly or very long and fastidious. In this method chapter we show how to use GraphiteLifeExplorer, a software with a simple graphical user interface that enables the sketching of free forms of DNA, of any length, at the atomic scale, as fast as drawing a line on a sheet of paper. We took as an example the nucleoprotein complex of DNA gyrase, a bacterial topoisomerase whose structure has been determined using cryo-electron microscopy (Cryo-EM). Using GraphiteLifeExplorer, we could model in one go a 155 bp long and twisted DNA duplex that wraps around DNA gyrase in the cryo-EM map, improving the quality and interpretation of the final model compared to the initially published data.journal article2017importe

Focused classification and refinement in high-resolution cryo-EM structural analysis of ribosome complexes

Cryo electron microscopy (cryo-EM) historically has had a strong impact on the structural and mechanistic analysis of protein synthesis by the prokaryotic and eukaryotic ribosomes. Vice versa, studying ribosomes has helped moving forwards many methodological aspects in single particle cryo-EM, at the level of automated data collection and image processing including advanced techniques for particle sorting to address structural and compositional heterogeneity. Here we review some of the latest ribosome structures, where cryo-EM allowed gaining unprecedented insights based on 3D structure sorting with focused classification and refinement methods helping to reach local resolution levels better than 3A. Such high-resolution features now enable the analysis of drug interactions with RNA and protein side-chains including even the visualization of chemical modifications of the ribosomal RNA. These advances represent a major breakthrough in structural biology and show the strong potential of cryo-EM beyond the ribosome field including for structure-based drug design