Ribosomal Subunit Exchange: Analysis in Vivo

This chapter describes the subunit exchange in ribosomes that can be studied in vivo by centrifugal analysis of the distribution of isotopic labels among ribosomes and ribosomal subunits following transfer of cells uniformly labeled with heavy isotopes to a medium containing only light isotopes. The technique is based on the fact that at the time of transfer, the cell contains only heavy ribosomes and ribosomal subunits. During growth in light medium exchange of subunits among ribosomes is manifested by the progressive replacement of the originally heavy ribosomes by two species of hybrid density. In Escherichia coli, one of these two hybrid species contains a heavy 50 S and a light 30 S subunit, the other a light 50 S and a heavy 30 S subunit. Thus, formation of ribosomes of hybrid density may be detected by a variety of ultracentrifugal methods. The density assay for ribosomal subunit exchange can be used with all organisms capable of growing on heavy isotopes, and this technique allows a direct demonstration of the cyclic dissociation into subunits and reformation of ribosomes under conditions of normal growth.