Poster abstracts

Poster number 8 submitted by Brandon Crowe

Binding characteristics of archaeal RNase P proteins RPP30 and Pop5.

Brandon L. Crowe (Biochemistry, Ohio State University), Chris Bohlen, Ross Wilson, Venkat Gopalan (Biochemistry, Ohio State University), Mark P. Foster (Biochemistry, Ohio State University)

Abstract:
The enzyme RNase P, which is composed of both RNA and protein subunits, is essential in all forms of life. The RNA moiety of the ribonucleoprotein (RNP) complex has been shown to be catalytic on its own, although the protein subunits are required for its in vivo activity. The number of proteins subunits varies from 1 in Bacteria to 9 or 10 in Eukarya. Here we report studies of a protein-protein interaction between subunits of RNase P from the hyperthermophilic Archaeon Pyrococcus furiosus (Pfu). Pfu has five RNase protein subunits (RPPs): RPP21, RPP29, RPP30, RPP38 and Pop5. Four of these subunits form two binary pairs RPP21-RPP29 and RPP30-Pop5. Gel filtration and dynamic light scattering (DLS) were used the show that the complex forms a heterotetramer in solution. Isothermal titration calorimetry (ITC) was used to show the RPP30 binds to Pop5 tightly with a 1:1 stoichiometry. Finally, nuclear magnetic resonance (NMR) was used to study RPP30 upon binding to Pop5. The positions of many signals in the ¹H-¹⁵N correlation spectrum of RPP30 changed upon the addition of Pop5. These chemical shift perturbations indicate the likely interaction surface for the complex. These results continue to add insight into interactions involved in RNP formation and will inform future efforts to elucidate assembly and structure of the whole RNase P complex.

Keywords: RNase P, protein-protein interaction, NMR