[29] Surface topography of ribosomal RNA

Ribosomal ribonucleic acids (rRNA) form the very compact central core of ribosomal subunits, whereas both ribosomal proteins and rRNA segments have been found on their surface. Since there is scarcely any doubt now that rRNA is directly involved in ribosome functioning, investigation of these exposed rRNA regions is of great interest. Hence they are likely to be involved in the interactions of ribosome functional centers with tRNA, mRNA, and initiation factors, as well as in subunit association. This chapter concentrates on the methods which have been developed in the laboratory for identification of RNA regions located on the surface of rRNA both in the isolated state and in ribosomal subunits. The protocol for analytical cleavage of 16S rRNA at m7G can easily be converted to large-scale use by proportionally increasing the amounts of all the reagents, keeping the concentrations unchanged. In the course of RNA cleavage, the methyl-RNA is degraded to rather short fragments which do not interfere with the 16S rRNA fragments either in the context of analytical electrophoresis, or preparative sucrose gradient fractionation.