3D modeling of ribosomal RNA using cryo-electron microscopy density maps

Episode von Fakultät für Chemie und Pharmazie - Digitale Hochschulschriften der LMU - Teil 04/06 von Ludwig-Maximilians-Universität München · 25. Mai 2011

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.

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