One of the difficulties encountered was that no usable crystals could be produced from mitochondrial ribosomes in order to determine their structure. Until now, X-ray crystallography, where the molecule is isolated, crystallised and analysed by X-rays, has been the method of choice to examine the structure of large biological molecules at high resolution. The X-rays are deflected by the atoms in the crystal, thereby creating a specific pattern that can be used to calculate the atom positions. However, for such an experiment to succeed, the crystal must be sufficiently big and of high quality. The large subunit of the mitochondrial ribosome is not suitable for this procedure, as its structure is too heterogeneous, and insufficient amounts of material can be extracted for the crystallisation process. "We would have needed hundreds of kilograms of pig liver in order to isolate sufficient quantities of ribosomal material for crystallographic structure analysis; it was logistically impossible to achieve," says Basil Greber, the lead author of the study and a post-doctoral researcher in Nenad Ban's group.
Success thanks to a clever combination
The ETH researchers therefore used the latest generation of high-resolution cryo-electron microscopes, which have only recently become available at the Electron Microscopy Center of ETH Zurich (EMEZ) and from the manufacturer. The researchers captured more than a million images of the large subunit of the mitoribosome and reconstructed its three-dimensional structure by performing complex calculations on a computer cluster.
In order to interpret the calculated structure as precisely as possible and to determi
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