The structure was solved by using multiple-wavelength anomalous diffraction (MAD) from crystallized ribosomes of Thermus thermophilus bacteria. The crystals contained a synthetic mRNA analog and tRNA molecules bound to two sites (the P and E sites). The high flux from the ALS wiggler yielded diffraction data with a resolution of 5.5 Å after density modification algorithms were applied. The group additionally used ribosome complexes with and without tRNA bound to a third site (the A site) to make a Fourier difference map that showed the A-site position at 7 Å resolution.

Ribosomes consist of ribosomal RNA (rRNA) and proteins. The ribosome’s ability to function is known to depend more on RNA than on protein, but until now scientists did not know why. The high-resolution structure shows the answer: the protein­protein and protein­RNA interfaces tend to occur away from functional sites, whereas the RNA­RNA interactions exist near functional centers. In addition, the interactions between the ribosome and tRNA occur mainly through contacts with rRNA.

Central to the function of the ribosome and to the revelations of this latest view of it are the intersubunit bridges. These join the two subunits, holding them together around the string of mRNA that is being decoded and the tRNA molecules whose anticodons pair with codons on the mRNA. The new crystal structure shows all the molecular components of the known contacts between the two subunits, plus two new bridges.

The Machinery of Life

The ribosome promises to answer one of mankind's oldest questions: How is life perpetuated? Stripped down to biological basics, the question can be posed as "How is the genetic code read to make proteins?"

This process occurs at the cellular level, on the surface of the ribosome.The code for a specific protein is carried out of the cell's nucleus on a string of messenger RNA (mRNA). A ribosome moves along the length of the mRNA, translating the code into a growing chain of amino acids that, when complete, will be the protein. Amino acids arrive via transfer RNA (tRNA). Each tRNA carries a specific amino acid to be paired with a corresponding sequence of three bases (a codon) on the mRNA and added to the amino acid chain.

The physical mechanism behind all this activity remains unclear. Recent studies have shown the ribosome to be capable of a surprising amount of motion, like the gyrations of a man-made machine. And as a student of engineering might come to understand the workings of a machine by studying the structure of its gears and pulleys, biologists must study the structure of the ribosome to grasp the nature of its workings.

Interfaces of the 50S (left) and 30S (right) subunits of the ribosome with intersubunit bridges numbered. Magenta, RNA–RNA contacts; yellow, protein–protein and protein–RNA contacts; A, P, and E mark tRNAs at left and tRNA anticodon stem loops at right.

Previous studies have shown that the tRNAs move through the space between subunits, translocating from the A site to the P site to the E site. Now, an important structural clue to the mechanism of this motion has been glimpsed. The new structure shows that these sites are all adjacent to intersubunit bridges. Since motion occurs around these sites, and the bridges are near enough to change shape as it occurs, it is likely that this motion is coupled with movement of the subunits relative to each other. This structural information complements cryoelectron microscopy and neutron scattering studies suggesting intersubunit movement. Such studies have also made a strong case for movement of the head of the 30S subunit, relative to both the rest of that subunit and the rest of the ribosome. This is reinforced in the new structure by the finding that the four domains making up 16S rRNA are nearly structurally independent of each other (and hence can move relative to each other with little change in shape). In addition, the four domains converge near sites of functional interactions with mRNA and tRNA, suggesting that their relative movements could be closely coupled with ribosome function.

Much work remains to be done before we have a complete solution to the mystery of how the ribosome works, but this latest effort provides vital structural information against which to test models of the ribosome’s machinations.

Research conducted by M.M. Yusupov, G.Zh. Yusupova, A. Baucom, K. Lieberman, and H.F. Noller (University of California, Santa Cruz); T.N. Earnest (Berkeley Lab); and J.H.D. Cate (Whitehead Institute).

Research funding: National Institutes of Health, Agouron Institute, W.M. Keck Foundation, Whitehead Institute for Biomedical Research, Searle Scholars Program. Operation of the ALS is supported by the U.S. Department of Energy, Office of Basic Energy Sciences.

Publication about this research: M.M. Yusupov, G.Zh. Yusupova, A. Baucom, K. Lieberman, T.N. Earnest, J.H.D. Cate, and H. F. Noller, "Crystal Structure of the Ribosome at 5.5 Å Resolution," Science 292, 883 (2001).

ALSNews Vol. 184, September 12, 2001