The in vitro translation of a terminating signal by a single Escherichia coli ribosome. The fate of the subunits.

Abstract

A complex was isolated containing one 70 S ribosome bound to the RNA of the f2 bacteriophage in the region of the coat gene. Either the f2 RNA and 70 S ribosome or both ribosomal subunits in this complex were differentially labeled with tritiated lysine or phosphorous 32 RNA. Incubation of this complex in a ribosome-free protein-synthesizing system lacking initiating factor and appreciable nuclease activities allowed the synthesis of coat protein and subsequent termination. Using this techique, it was found that termination resulted in the release of ribosome as subunits. Intermediate in this process is a transient 46 S complex composed of the f2 RNA and 30 S ribosomal subunit, presumably the result of the initial release of the 50 S ribosomal subunit. This 46 S complex subsequently breaks down to a free f2 RNA and 30 S ribosomal subunit. The rate of this latter process is faster in the presence of crude initiating factors. These results are discussed in terms of the mechanism of termination and reading of the intercistronic region.