Ribonucleases Active at 3′ Terminus of Transfer RNA

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. © 1990, Elsevier Inc. All rights reserved.