Book of Abstracts: Albany 2007

category image Albany 2007
Conversation 15
June 19-23 2007

Ribosomes Under Stress: Structural Variability of the 100S Particles Studied by Cryoelectron Tomography

Since the description of ribonucleoprotein particles in E. coli, it was clear that the 70S ribosome can form dimers (1). The dimerized form, the 100S ribosome, has been associated with stress conditions, such as starvation in stationary phase. The 100S ribosomes are different from polysome dimers in the sense that they are not translating but are in a hibernation-like state until the cell needs higher levels of protein synthesis. Protein factors that are thought to be involved in the formation/dissociation of dimers are RMF, YhbH, YfiA, and SRA (2). The binding sites for some of these components have been determined biochemically but the structure of the 100S particle has not been solved. Cryoelectron tomography (CET) is uniquely suited to obtain three-dimensional reconstructions of pleiomorphic structures such as intact cells or supramolecular assemblies in their native state and their natural environment (3). CET and template matching has been successfully used to detect 70S ribosomes in a crowded cellular environment (4). Here, we have applied these methods to semi purified samples of 100S ribosomes to describe the 3D arrangement of its components. In the 100S particles, we confirmed that the two 70S particles are facing each other through their 30S subunits. Even though the general relative orientation of the two 70S ribosomes in a dimer appears to be antiparallel, we found a broad angular distribution of orientations. The structural variability of this assembly might be the result of a dynamic process or a flexible link. We have aligned subtomograms containing 70S ribosomes in dimers and averaged them to obtain a density map that reveals additional densities to the 70S at the entrance and exit of the mRNA path, at the tRNA binding site in the 30S subunits, as well as a major conformational change of one domain of the 50S subunit close to the protein L4. We expect 3D models for complete 100S ribosomes in particular orientations with classification of larger data sets. Furthermore, we present initial attempts to find the same type of arrangement of the 100S ribosomes in cryo sections of E. coli.

References and Footnotes
  1. Tissieres, A. and Watson, J. D. Nature 182, 778-780 (1958).
  2. Ueta, M. et al., Genes Cells 10, 1103-12 (2005).
  3. Lucic, V., Foerster, F., and Baumeister, W. Annu Rev Biochem 74, 833-865 (2005).
  4. Ortiz, J. O., Foerster, F., Kuerner, J., Linaroudis, A. A., and Baumeister, W. J Struct Biol 156, 334-341 (2006).

Julio O. Ortiz1, *
Stephanie Etchells2
Andrew Leis1
F. Ulrich Hartl2
Wolfgang Baumeister1

1Max-Planck Institute of Biochemistry
Department of Structural Biology
Am Klopferspitz 18
D-82152 Martinsried, Germany
2Max-Planck Institute of Biochemistry
Department of Cellular Biochemistry
Am Klopferspitz 18
D-82152 Martinsried, Germany

*Phone: +49 (89) 8578 - 2032
Fax: +49 (89) 8578 - 2641
Email: ortiz@biochem.mpg.de