Breakthrough in mitochondrial ribosome structure

Mitochondrial ribosomes (mitoribosomes) are indispensable for living because they synthesize essential proteins, which constitute the catalytic core of the respiratory chain complexes embedded in the inner mitochondrial membrane. The aim of the HFSP postdoctoral fellowship, awarded to Alexey Amunts from Venki Ramakrishnan’s lab, was to reveal the atomic structure of mitoribosome. By the end of the 3rd year of the fellowship, it was revealed that the structure of yeast mitochondrial large ribosomal subunit is composed of 40 proteins and ~3000 nucleotides using cryo-EM to an overall resolution of 3.2 Å. This not only currently represents the highest resolution limit reported by cryo-EM, better than most ribosomal structures determined by X-ray crystallography, but it is also the first time that the atomic structure of a large asymmetric complex has been obtained without crystallization.

HFSP Long-Term Fellow Alexey Amunts and colleagues
authored on Sun, 20 April 2014

Methodologically, this work represents a breakthrough in Structural Biology, as it shows that biological macromolecules can be now directly visualized at the level of atomic details using advanced tools in electron cryo-EM combined with model building developments. The main advantage of the method is that it does not require intensive purification of a sample, nor any other biochemical manipulation. Thus, molecules can be directly visualized in their native cellular environment. The amount of material required for the structural analysis is less than 0.5µg, which is nearly four orders of magnitude smaller than for the currently widely used X-ray crystallography technique. Thus, low abundant and transient complexes that were previously intractable, can now be fully described.

Figure 1: Mitochondrial ribosomes are unique in their composition, because their components: proteins (red) and RNA (yellow), are encoded by two different genomes, nuclear and mitochondrial respectively. Mitoribosomal proteins are imported into mitochondria, where they are assembled onto RNA in a spatially and temporally synchronized manner, to form a functional mitoribosome. Another unique feature of mitoribosomes is that being responsible for production of highly hydrophobic transmembrane polypeptides, they have been evolutionary tuned to permanently tether to the inner mitochondrial membrane. This adaptation allows nascent polypeptides to be delivered through the exit tunnel (blue) straight onto the membrane surface in a co-translational manner.

Biologically, the structure shows that mitochondrial ribosomes have diverged greatly from their cytoplasmic counterparts during 2 billion years of separate evolution. Particularly, functionally important regions such as the nascent polypeptide exit tunnel, intra-ribosome communication network and tRNA binding sites exhibit unique and previously unraveled features. This first atomic portrayal of mitochondrial ribosomes significantly advances our understanding of the mitochondrial protein biosynthesis and will be instrumental for developing mitochondrial in vitro translation systems in the future.

Mitoribosomes represent a principal therapeutic target, as their enhanced activity in tumors constitutes the main pathway required for cancer proliferation. Thus, structure of mitoribosome will serve as a component of the unabated effort to develop new therapeutic interventions for cancer and drugs with reduced autotoxic effects.

  

Figure 2: The level of structural details described in this work is unprecedented in the field of electron cryo-microscopy. Not only that for the first time single nucleotides and ions could be distinguished in the density map (left), but also amino acid side chains could be modeled without any a priori biochemical information (right). This allowed the complete atomic model of yeast mitoribosomal large subunit to be built.

Thus, the methodology described in this work represents the first successful attempt of de novo model building of asymmetrical biological assembly using cryo-EM. The results will stimulate many other laboratories to reveal further functionally important insights for a wide variety of cellular samples.

Reference

Structure of the Yeast Mitochondrial Large Ribosomal Subunit. Alexey Amunts, Alan Brown, Xiao-chen Bai, Jose L. Llácer, Tanweer Hussain, Paul Emsley, Fei Long, Garib Murshudov, Sjors H. W. Scheres, V. Ramakrishnan. Science 343, 1485 (2014 DOI: 10.1126/science.1249410).

Pubmed link