PARTIAL-PURIFICATION AND CHARACTERIZATION OF RNASE-P FROM DICTYOSTELIUM-DISCOIDEUM

Ribonuclease P (RNase P) from Dictyostelium discoideum has been purifi ed 470-fold. D. discoideum RNase P cleaves the precursor to Schizosacc haromyces pombe suppressor tRNA(Ser) at the same site as S. pombe RNas e P, producing the mature 5' end of tRNA(Ser). pH and temperature opti ma for enzyme activity are 7.6 and 37 degrees C, respectively. The enz yme shows optimal activity in the presence of 5 mM MgCl2 and 10 mM NH4 Cl or 5 mM KCl. The apparent K-m for the S. pombe tRNA precursor deriv ed from the supSl tRNA(Ser) gene is 240 nM, and the apparent V-max is 3.6 pmol/min. Inhibition of D. discoideum RNase P by proteinase K and micrococcal nuclease strongly indicates that the activity requires bot h protein and RNA components. In cesium sulfate density gradients, the enzyme has a buoyant density of 1.23 g/ml, indicating a low RNA/prote in ratio for the holoenzyme.