RNA Structure and RNA-Protein Neighborhoods in the Ribosome

Abstract

One of the central objectives in ribosomology is a detailed determination of the three-dimensional organization of the ribosomal RNA (rRNA) in situ in the subunits. Research in this direction has been spurred by the fact that reliable secondary structure models are now available for both the large and small subunit rRNA molecules. The secondary structures have been derived by phylogenetic comparisons of different rRNA sequences in combination with various experimental approaches, and in the case of Escherichia coli approximately 85% of the proposed secondary structural elements in the 16 S and 23 S rRNA can be regarded as proven (Maly and Brimacombe, 1983; Noller, 1984). Thus, the problem of determining the RNA structures has now become the question of how these secondary structures are packaged into three dimensions in the intact ribosome, and how the ribosomal proteins are fitted into the RNA lattice. Our approach to this problem is based on crosslinking techniques; E. coli ribosomal subunits are used as substrates for reactions in which both intra-RNA crosslinks (e.g., Stiege et al., 1983) or RNA-protein crosslinks (e.g., Wower and Brimacombe, 1983) are generated, and in each case the sites of crosslinking on the RNA are determined, with a view to building up a catalog of points of precise topographical neighborhood both within the RNA as well as between the RNA and the individual ribosomal proteins.