Local quality assessment of cryo-EM reconstructions and its applications.

Resum

The cryo-Electron Microscopy (cryoem) addressed to the elucidation of macromolecular complexes has suffered a revolution in the last years. Single Particle Analysis SPA and electron tomography are the two main branches responsible of this revolution. In both cases, the result is a 3D structure of the specimen. However, it is critical to determine the degree of reliability of the reconstructed structure, problem which is solved with resolution measurements. Resolution can be determined as a global parameter of the structure, or as a local property that can spatially vary. The main objective of the present thesis is the study of the local resolution and its applications for the validation of the reconstructed 3D structure and for local sharpening. For that, a mathematical method named MonoRes was developed, which measures the local resolution of reconstructed density maps and its algorithm represents the cornerstone of this thesis. Hence, MonoDir was designed for identifying reconstructions problems like the existence of preferred directions, or alignment errors. Note that the current state of the art does not provide an immediate and simultaneous response to both questions, in particular to the second one, which can be due to a systematic error of reconstruction. The proposed method does it by analyzing local-resolution anisotropy using the information of only the reconstructed map. Moreover, the measurement and analysis of the local resolution for map sharpening can be also used to enhance the visualization of the protein structure keeping its structure factor. For that, method LocalDeBlur was developed, which modifies the local amplitudes of the different frequencies according to the local resolution values. Apart from that, other methods were developed using slight variations of MonoRes algorithmic core. One of these methods is MonoTomo, which has been developed as the first method of local resolution in Electron Tomography. The estimation of the local resolution in electron tomograms involves work with spatially dependent noise and very large maps, so the core of MonoRes was extended in that direction, resulting in method MonoTomo, which solves those drawbacks. These other MonoRes core-derived methods extend the concept of resolution in a local manner. However, resolution is not only a local parameter, but it must be considered a tensor and, therefore, it depends on the position (voxel) and the direction. Thus, the concept of local resolution is then enriched with the notion of directionality.