9] Human ribonuclease P

Abstract

Publisher Summary Ribonuclease P is a ribonucleoprotein nuclease required for the site-specific cleavage of the 5′ leader sequence of precursor tRNAs. In eubacteria, the RNA subunit of RNase P is the catalytic moiety and is capable of processing precursor tRNA in the presence of divalent metal ions. The RNA subunits of eukaryotic RNase P (human and yeast) do not exhibit enzymatic activity In Vitro, an indication that their protein components are required for the catalytic reaction. This chapter describes a purification procedure for human RNase P from HeLa cells that facilitated the characterization of nine of its protein subunits. This procedure also allows the separation of RNase P from the structurally related RNase MRP nuclease. The purification procedure shows that nuclear RNase P from HeLa cells has at least nine protein subunits in association with a single RNA species, H1 RNA. Nuclear RNase P in other eukaryotes, such as S. cerevisiae and Aspergillus nidulans, also consists of multiple protein subunits, some of which are evolutionarily conserved and an RNA subunit. The precise function of these subunits in tRNA processing is not yet understood. Reconstitution of the activity of RNase P in vitro may reveal the role of these subunits in catalysis.