Characterization of protein-deficient ribosomes as starting point for the design of novel antimicrobials interfering with ribosomal protein assembly

The translation of the genetic code into proteins by ribosomes is one of the fundamental processes of life. Intensive basic research has been and is still devoted to ribosome function, and many ongoing studies focus on prokaryotic ribosomes or components thereof as antimicrobial drug targets. Recent structural and biochemical data of bacterial ribosomes, which consist of three RNA molecules and 51 proteins, support the hypothesis that the first ribosomes were entirely composed of RNA, and that ribosomal proteins, the function of which remains largely unknown, were added later during evolution. Surprisingly, my recent studies revealed that the antibiotic kasugamycin interferes with assembly of the small bacterial ribosomal subunit, leading to the formation of stable ~ 61S ribosomal particles, which consist of an intact 50S subunit but lack a number of proteins of the 30S ribosomal subunit. Despite the fact that mutations in ribosomal protein genes are generally deleterious, these "61S" ribosomal particles selectively translate leaderless mRNAs, which start directly with a 5-terminal AUG and lack kingdom-specific ribosome recruitment signals. These initial studies provide the first in vivo evidence for the functionality of protein-deficient ribosomes, and therefore strongly support the ideas that modern ribosomes are protein-stabilized ribozymes, and that proto- ribosomes were one-subunit particles that translated simple polynucleotides. It is conceivable that assembly of the "61S" particles is stalled at an intermediate step towards evolution of the modern ribosome before the kingdoms have diverged. Therefore, a major aim of this study is the structural and functional characterization of the "61S"-particles, which is expected to shed more light on the evolution and mechanism of the contemporary translational apparatus as well as on the function of some selected ribosomal proteins in this process. On the other hand, the analysis of structural changes in the 16s rRNA caused by kasugamycin is expected to contribute to a further understanding of the binding requirements for ribosomal proteins known to be necessary for translation of canonical mRNA in Gram-negative bacterial pathogens, like ribosomal proteins S1 and S2. In perspective these studies could lead to the design of new drugs, which specifically inhibit binding of these ribosomal proteins, and which may not interfere with the function of eukaryotic ribosomes, as many of the antibiotics currently in use do.