Scientific Autobiography – 1

Ribosome Microcrystals

In 1965 I was employed as a research assistant at the Institute of Histology and Embryology in the Medical Faculty of Bologna University and a few months later, in 1966, I read a paper by Breck Byers that described the results of a cooling treatment on one-day old chick embryos (Byers 1966). More precisely, Byers discovered that an incubation of the eggs at 4 degrees centigrade for 24 hours was inducing in the embryonic cells the formation of ribosome microcrystals. I was immediately attracted by this discovery because it was the proof that ribosomes can form crystals and getting a crystal is the first step for reconstructing a three-dimensional structure.

    The technology for the reconstruction of the structure of proteins from X-rays became available in the 1950s and in the 1970s was extended to viruses, structures that are more complex than proteins because they contain a variety of components and have molecular weights in the order of a million dalton.

    The ribosomes are more complex than viruses because in addition to heavier nucleic acids they contain between 50 and 60 different proteins in prokaryotes, and between 70 and 80 proteins in eukaryotes.

    Organelles like nucleoli, mitochondria or chloroplasts are even more complex than ribosomes, but they cannot form crystals because they do not have the molecular identity that is the sine qua non condition for crystallization. To the best of our knowledge, there is no other object that can form crystals and is more complex than ribosomes, and this is why the discovery of ribosome microcrystals was so fascinating to me: those crystals could reveal the three-dimensional structure of the eukaryotic ribosomes and at the same time they were the most complex crystals in the universe.

    The first problem that I addressed was the formation of microcrystals in older embryos and this turned out to be possible but only to a limited extent. Older embryos contain more cells but their cells are forming fewer crystals and in the end I found that the best yield was obtained with 5-days old chick embryos.

    In 1970 I and my colleagues discovered how to isolate ribosome microcrystals from chick embryos (Barbieri et al. 1970) and Max Perutz invited me to the Molecular Research Council in Cambridge (UK) where I discussed the problem of reconstructing the structure of the ribosome not only with Perutz but also with Francis Crick, Hugh Huxley and Aaron Klug.

    The main obstacle was the fact that the microcrystals obtained in chick embryos were too small for the reconstruction of the ribosome structure, and it became necessary to study the mechanism of their crystallization in order to find out whether or not the size of the microcrystals could be increased. This is the problem that I worked on at the Max-Plack-Institute für Molekulare Genetik in Berlin.

    The formation of a crystal requires that the ribosomes are brought together at very close molecular distances, but this is not what normally takes place in the cell because the ribosome transport system distributes them throughout the cytoplasm. This suggested that the cooling of the eggs was concentrating the ribosomes in small regions because it was damaging their transport system. If this hypothesis was correct, therefore, the microcrystals should be induced not only by cooling but also by other processes that damage the ribosome transport system, and I found that this is indeed the case.

    I obtained the formation of ribosome microcrystals in chick embryos with many other treatments, for example by keeping the eggs under sand or under water. Finally I discovered that the microcrystals can be formed not only in intact cells but also in fragmented cells and in cell homogenates and it is actually in these systems that they reach their greatest dimensions.

    These results were published in the Journal of Supramolecular Structure (Barbieri, 1979) and those microcrystals are still the biggest crystals of eukaryotic ribosomes that have been obtained so far. A few electron microscope pictures of those microcrstals are shown in the following gallery.

A Gallery of Eukaryotic Ribosome Microcrystals

    Barbieri M (1979) A Gallery of Ribosome Microcrystals