The broad, long-term objective of this research is to know the structure of ribosomes and to use the structure to decipher the molecular basis of the function of the particle in protein synthesis. The research proposed here is dedicated to an aspect of the undertaking-the structure of rRNA and the chemistry of the interaction of the nucleic acid with proteins. The strategy for the research is epitomized in the phase, the ribosome-in- pieces. This is a reductionist approach in which the structure and function of domains are analyzed; it is predicated, and is absolutely dependent, on the ribosomal domain retaining relevant structure and relevant function in isolation. With the binding of EF-G to sarcin-ricin and thiostrepton region oligoribonucleotides we have achieved this goal. We propose to expand on this success by refining the characterization of these two domains and addressing the structure and function of other ribosome domains. In the research we shall combine, wherever and whenever possible, biochemistry, genetics, and structure. The specific aims are: 1. To continue the characterization of the binding of EF-G to oligoribonucleotides that reproduce the sarcin/ricin and the thiostrepton domains by constructing mutations in nucleotides in the RNAs and in amino acids in EF-G. We shall also attempt to determine the ribosome binding site on EF-G by cross-linking the factor to sarcin/ricin and thiostrepton RNAs. 2. To determine the role of sarcin/ricin and thiostrepton RNAs, and of ribosomal proteins, in the activation of the GTPase of EF-G. 3. To characterize the binding of EF-Tu to tRNA oligonucleotides. 4. To continue to construct mutations in vivo in the sarcin/ricin domain in Escherichia coli 23S rRNA and to determine the effects on growth and on the partial reactions of protein synthesis. 5. To determine, in collaboration with Dr. Carl Correll, the three-dimensional (3-D) structure of rRNA domains by x- ray diffraction of crystals.