The long-term goal of this project is to elucidate why alterations in isoaccepting species of transfer ribonucleic acids (tRNAs) occur, determine what regulated the alterations, and understand the role of the altered tRNA. We will concentrate on the structure and function of specific tRNAs that show alterations during development in Bacillus subtilis. In this system, the alterations in the tRNAs appear to be due to changes in nucleoside modifications that are highly specific, in particular, modified nucleosides that oocur in the anticodon loop. Both modification and "demodification" of the basic modified residue are involved. The function of the modification for lysine tRNAs can be related to a difference in codon-anticodon interactions. Experiments are planned to (a) determine the primary sequence for two lysine isoacceptors, (b) investigate in detail the modification changes in lysine and tyrosine tRNAs, (c) study the differential utilization of lysine isoaccepting tRNAs by polyribosomes "in vivo" and by an "in vitro" protein-synthesizing system, (d) investigate the modification mechanisms for tyrosine and lysine tRNAs in detail, (e) purify the modification enzyme and protein(s) with unique binding selectivity for a particular isoaccepting species by affinity chromatography, and (f) isolate mutant strains lacking the enzymes responsible for the alterations in modified nucleosides.