Abstract
Several E. coli exoribonucleases able to act at the 3′ terminus of tRNA and tRNA precursors have been identified and characterized to varying degrees. The assay for RNase D measures the conversion of a specific radioactive tRNA substrate to an acid-soluble form. The first substrate is an analog of a tRNA precursor, the presumed substrate for RNase D in vivo. The second substrate is a mixture of tRNA molecules lacking two or three of the 3'-terminal residues. Removal of terminal residues from some tRNA species alters their conformation and renders them susceptible to RNase D action. The incorporation of CMP into intact tRNA occurs at a slow rate and is due to an anomalous reaction carried out by the rabbit liver enzyme. RNase D is a 3′-exoribonuclease with a high degree of specificity for tRNA-like molecules. It can remove extra residues from the 3′ terminus of a tRNA precursor in a random fashion and regenerate amino acid acceptor activity. Its rate of hydrolysis of intact tRNA is less than 5% of its rate on tRNA precursors.
