James W. Brown

Associate Professor & Undergraduate Coordinator
Department of Microbiology, NC State University

RNA 2000 (The RNA Society), Madison, WI

Characterization of an Archaeal RNase P Holoenzyme


Department of Microbiology, North Carolina State University, Box 7615, Raleigh, NC 27695

Although archaeal RNase P RNAs are similar in both sequence and structure to those of Bacteria, no archaeal sequences with similarity to any known bacterial RNase P protein subunit have been identified. The buoyant density of RNase P from the methanogenic archaeon Methanobacterium thermoautotrophicum delta H in cesium sulfate was found to be 1.42 g/ml, inconsistent with a single small bacterial-like protein subunit and relatively large RNA. A hypothetical open reading frame (MTH687) was identified in the genome M. thermoautotrophicum delta H that has sequence similarity to Pop5p, one of the nine Saccharomyces cerevisiae RNase P protein subunits. Polyclonal antiserum generated to recombinant MTH687 recognized a protein of the predicted molecular weight in western blots of partially-purified M. thermoautotrophicum delta H RNase P, and immunoprecipitated RNase P activity from the same partially-purified preparation, demonstrating the first identification of an archaeal RNase P protein subunit. Three additional M. thermoautotrophicum delta H open reading frames, MTH11, MTH688 and MTH1618 exhibit sequence similarity to yeast RNase P protein subunits Pop4p, Rpr1p and Rpp2p, respectively. These proteins are currently under investigation. Structural similarity of M. thermoautotrophicum delta H RNase P RNA to the RNase P RNAs of Bacteria is supported by the observations that, unlike eukaryotic RNase P RNAs, the M. thermoautotrophicum delta H RNA alone is catalytic under high ionic conditions in vitro, and that activity of this RNA can be reconstituted under low ionic conditions with the RNase P protein subunit from the bacterium Bacillus subtilis. The RNase P from M. thermoautotrophicum delta H represents a single archaeal enzyme complex with components resembling both bacterial and eukaryotic homologs. These observations suggest that a bacterial-like RNase P RNA and eukaryotic-like RNase P proteins may have made up the primitive RNase P structure. If so, a single, simple bacterial RNase P protein may represent a derived characteristic of this enzyme, rather than a present-day remnant of an RNA world.


nullLast updated by James W Brown