D-Tyrosyl RNA: formation, hydrolysis and utilization for protein synthesis

Highly purified tyrosyl RNA synthetases from Escherichia coli and Bacillus subtilis (Calendar &; Berg, 1966a,b,) catalyze the esterification of d-tyrosine, as well as l-tyrosine, to tRNA_tyr. Although the extent of tyrosyl RNA formation is the same with either enantiomer, the V_max value with l-tyrosine is considerably greater than with d-tyrosine; the K_m values for d- and l-tyrosine do not differ from one another by more than a factor of four with either enzyme. Extracts of E. coli, yeast, rabbit reticulocytes and rat liver but not B. subtilis, contain an enzyme which rapidly hydrolyzes the ester linkage of d-tyrosyl RNA but not of l-tyrosyl RNA. This deacylase has been partially purified from E. coli and in addition to the above specificity, we find that d-phenylalanyl RNA is split at about 20% the rate of d-tyrosyl RNA, while l-amino acyl RNA's are not detectably cleaved. d-Tyrosyl adenosine is not hydrolyzed, although a d-tyrosyl oligonucleotide produced by t₁ RNase digestion is readily deacylated. With ribosomes from E. coli, soluble proteins from B. subtilis and UA polymer as messenger, we have shown that d-tyrosine from d-tyrosyl RNA can be incorporated into peptide linkage. The d-tyrosine occurs in internal positions as well as in NH₂- and COOH- terminal positions. This means that the protein synthetic mechanism cannot exclude the incorporation of a d-amino acid if it remains esterified to its cognate tRNA.