Genetic approaches to the study of ribosomes

Abstract

Specific mutants of Escherichia coli have been used in order to investigate certain aspects of ribosome function, bio synthesis and decay. These studies show that in the function of the ribosome its various components are very interdependent and cooperative. It was possible to demonstrate that a binding site for erythromycin, which is on the 50S ribosomal subunit, could be affected by alterations in the 30S ribosomal subunit. The alterations are in ribosome proteins as well as in 16S rRNA. On the other hand, a mutational alteration, probably in the 16S rRNA, blocks ribosome maturation but at a relatively late stage. This alteration leads to cell death. The maturation cleavages of rRNAs were investigated in as train lacking RNase III, a specific enzyme which cleaves out p16 and p23 from the growing rRNA transcript. These studies suggest that at least three ribonucleases participate in the primary processing (during transcription), while at least three other ribonucleases participate in the secondary processing, which leads to the final mature rRNA molecules. When cells stop growing exponentially, decay of ribosome stakes place. By analyzing the process in mutants lacking or containing altered ribonucleases, it was possible to elucidate a mechanism for this turnover. The process starts by an endonucleolytic attack on the rRNA in the ribosomal subunits. After such an attack the ribosome disintegrates to fragments of RNA and ribosomal proteins, both of which can be further decayed to soluble material. Two endoribonucleases were identified in cell extracts which could be the enzymes which initiate this important turnover process.