RNA CODEWORDS AND PROTEIN SYNTHESIS, VI. ON THE NUCLEOTIDE SEQUENCES OF DEGENERATE CODEWORD SETS FOR ISOLEUCINE, TYROSINE, ASPARAGINE, AND LYSINE

Nucleotide sequences of RNA codons have been investigated recently by directing the binding of C'4-AA-sRNA to ribosomes with trinucleotides of defined base sequence. The template activities of 12 trinucleotides have been described and nucleotide sequences have been suggested for RNA codons corresponding to phenylalanine, valine, leucine, cysteine, proline, serine, and lysine.l-4 In this report, the template activities of seven additional trinucleotides are described. ApUpU and ApUpC serve as RNA codons for isoleucine, UpApU and UpApC for tyrosine, ApApU and ApApC for asparagine, and ApApG and ApApA for lysine (cf. ref. 1). These findings are discussed in terms of the recognition of specific bases at different positions within a trinucleotide. Materials and Methods.-Components of reactions: E. coli W 3100 ribosomes and sRNA were prepared by modifications of methods described previously.5-7 Each C14-aminoacyl-sRNA was prepared in the presence of 19 C"2-amino acids. The assay for ribosomal bound C14-aminoacyl-sRNA has been described.' Each 50-1 reaction mixture contained 0.10 M1 Tris-acetate, pH 7.2; 0.05 M KCI; 0.03 M magnesium acetate; 1.5-2.0 A260 units of washed E. coli W 3100 ribosomes; and the amount of C14-AA-sRNA indicated in 'Table 1. Other data concerning each C14-AA-sRNA preparation are also given in Table 1. Uniformly labeled C14-amino acids were purchased from New England Nuclear Corp. or Nuclear-Chicago Corp. Radioactivity was determined in a liquid scintillation counter (Packard Inst. Co.) with a C14-counting efficiency of 55-65% as described previously.l Trinucleotides: A derivative of bovine pancreatic ribonuclease was used to synthesize UpApC.8 All other trinucleotides were prepared with a highly purified preparation of polynucleotide phosphorylase.9 The purity of each trinucleotide preparation was assessed as described previously.-4-" 9 The UpApC preparation contained approximately 8% cytidine-2'(3')-phosphate. Other trinucleotide preparatioils appeared to be homogeneous. Analyses of the ApApA preparation have been reported.l The base ratio, base sequence, and chain length of each oligonucleotide were established as shown in Table 2. The methodology employed has been described previously. 4, 9 Results and Discussion.-In Table 3 are shown the effects of eight trinucleotides upon the binding to ribosomes of 17 C-4-AA-sRNA preparations, each acylated with a different C14-amino acid (radioactive Cys-, Met-, and Tryp-sRNA were not used). Each trinucleotide markedly stimulated the binding to ribosomes of only one C14-AA-sRNA preparation. ApUpU and ApUpC stimulated C'4-Ileu-sRNA binding to ribosomes; UpApU and UpApC stimulated C14-Tyr-sRNA binding; ApApC and ApApU stimulated C14-Asp-NH2-sRNA binding; and ApApA and