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Jarasch, Alexander (2011): 3D modeling of ribosomal RNA using cryo-electron microscopy density maps. Dissertation, LMU München: Faculty of Chemistry and Pharmacy
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Abstract

Ribosomes are macromolecular protein-RNA complexes translating mRNA into protein. To date, crystal structures are available for the bacterial 30S and archaeal 50S subunits, as well as the complete bacterial 70S ribosomes. Eukaryotic ribosomes are much more complex in terms of ribosomal RNA and proteins. However, to date high-resolution crystal structures of eukaryotic ribosomes or ribosomal subunits are lacking. In order to build reliable models for the eukaryotic rRNA, we developed an approach for large scale homology and de novo modeling of RNA and subsequent exible tting into high-resolution cryo-EM density maps. Using this approach we built a model of the T. aestivum and the S. cerevisiae ribosome based on available cryo-EM maps at 5.5 Å and 6.1 Å resolution, respectively. The model comprises of 98% of the eukaryotic rRNA including all 21 RNA expansion segments (ES) and structurally six variable regions. Further, we were able to localize 74/80 (92.5%) of the ribosomal proteins. The model reveals unique ES-ES and r-protein-ES interactions, providing new insight into the structure and evolution of the eukaryotic ribosome. Moreover, the model was used for analyzing functional ribosomal complexes, i.e. the characterization of dierent nascent polypeptide chains within the ribosomal tunnel, intermediates of protein translocation as well as mRNA quality control.