Partial purification and properties of an endoribonuclease isolated from human KB cells

With the use of a precursor to Escherichia coli tRNA(Tyr) as substrate, the authors have detected and partially purified a novel endoribonuclease from the cytoplasm of human KB tissue culture cells. This activity, which has been called RNase NU, cleaves the tRNA precursor at two sites in that part of the molecule which is not included in the mature tRNA sequence and which is normally degraded in vivo. In keeping with this observation, it has been found that, of a variety of substrates tested, only those which are unstable in vivo are attacked by RNase NU. RNase NU can be purified from the 0.2 M NH₄Cl wash of ribosomes followed by ammonium sulfate fractionation and DEAE Sephadex chromatography. RNase NU cleaves RNA to create 3' phosphate terminated oligonucleotides. It has a pH optimum near 8.0, requires either a monovalent cation (NH₄+ is most efficient) or Ca for optimal activity, and is inhibited by 0.1 M phosphate. In the course of purifying RNase NU, the authors detected and studied the intracellular distribution of other ribonuclease activities in human KB cells.