Structure and mechanistic features of the prokaryotic minimal RNase P.

Rebecca Feyh,Roland K Hartmann,Florian Altegoer,Gert Bange,Pietro Ivan Giammarinaro,Georg Hochberg,Nadine B Waeber,Simone Prinz

doi:10.7554/elife.70160

Abstract

Endonucleolytic removal of 5'-leader sequences from tRNA precursor transcripts (pre-tRNAs) by ribonuclease P (RNase P) is essential for protein synthesis. Beyond RNA-based RNase P enzymes, protein-only versions of the enzyme exert this function in various eukarya (there termed PRORPs) and in some bacteria (Aquifex aeolicus and close relatives); both enzyme types belong to distinct subgroups of the PIN domain metallonuclease superfamily. Homologs of Aquifex RNase P (HARPs) are also expressed in some other bacteria and many archaea, where they coexist with RNA-based RNase P and do not represent the main RNase P activity. Here, we solved the structure of the bacterial HARP from Halorhodospira halophila by cryo-electron microscopy, revealing a novel screw-like dodecameric assembly. Biochemical experiments demonstrate that oligomerization is required for RNase P activity of HARPs. We propose that the tRNA substrate binds to an extended spike-helix (SH) domain that protrudes from the screw-like assembly to position the 5'-end in close proximity to the active site of the neighboring dimer. The structure suggests that eukaryotic PRORPs and prokaryotic HARPs recognize the same structural elements of pre-tRNAs (tRNA elbow region and cleavage site). Our analysis thus delivers the structural and mechanistic basis for pre-tRNA processing by the prokaryotic HARP system.

Highlights

Ribonuclease P (RNase P) is the essential endonuclease that catalyzes the 5’-end maturation of tRNAs (Klemm et al, 2016; Rossmanith and Hartmann, 2020; Guerrier-Takada et al, 1983)
There are many examples of proteins forming symmetric homo-dodecameric assemblies (e.g., glutamine synthase [van Rooyen et al, 2011]; helicases [Bazin et al, 2015] or bacterial DNA-binding proteins expressed in the stationary phase (DPS) [Roy et al, 2008]) and recent studies have discussed the theoretical types of possible quaternary structures (Laniado and Yeates, 2020; Ahnert et al, 2015)
The screw-like assembly of HARPs leads to an asymmetric and imperfect novel type of oligomer

Summary

Introduction

Ribonuclease P (RNase P) is the essential endonuclease that catalyzes the 5’-end maturation of tRNAs (Klemm et al, 2016; Rossmanith and Hartmann, 2020; Guerrier-Takada et al, 1983). A bacterial protein-only RNase P, associated with a single polypeptide as small as ~23 kDa, was discovered in the hyperthermophilic bacterium Aquifex aeolicus that lost the genes for the RNA and protein subunits (rnpB and rnpA) of the classical and ancient bacterial RNase P (Nickel et al, 2017). This prokaryotic type of minimal RNase P system was named HARP We here present the structural basis for the RNase P-like pretRNA processing activity of prokaryotic HARPs

Results

Discussion

Materials and methods

Funding Funder