Cryo-electron microscopy structure of an archaeal ribonuclease P holoenzyme

Futang Wan,Jian Wu,Shaohua Shi,Pengfei Lan,Juan Chen,Ming Tan,Ming Lei,Qianmin Wang,Jing Tan

doi:10.1038/s41467-019-10496-3

Abstract

Ribonuclease P (RNase P) is an essential ribozyme responsible for tRNA 5′ maturation. Here we report the cryo-EM structures of Methanocaldococcus jannaschii (Mja) RNase P holoenzyme alone and in complex with a tRNA substrate at resolutions of 4.6 Å and 4.3 Å, respectively. The structures reveal that the subunits of MjaRNase P are strung together to organize the holoenzyme in a dimeric conformation required for efficient catalysis. The structures also show that archaeal RNase P is a functional chimera of bacterial and eukaryal RNase Ps that possesses bacterial-like two RNA-based anchors and a eukaryal-like protein-aided stabilization mechanism. The 3′-RCCA sequence of tRNA, which is a key recognition element for bacterial RNase P, is dispensable for tRNA recognition by MjaRNase P. The overall organization of MjaRNase P, particularly within the active site, is similar to those of bacterial and eukaryal RNase Ps, suggesting a universal catalytic mechanism for all RNase Ps.

Highlights

Ribonuclease P (RNase P) is an essential ribozyme responsible for tRNA 5′ maturation
When purified L7Ae was added into the transcription reaction, RNase P RNA (RPR) behaved properly as a monodispersed molecule, suggesting that L7Ae presumably functions as a chaperone for the correct folding and stability of RPR (Supplementary Fig. 1b)
The in vitro transcribed RPR in the presence of L7Ae was sequentially mixed with purified Rpp29Rpp[21] and (Pop5-Pop30)[2] subcomplexes and analyzed by size exclusion chromatography (SEC) (Fig. 1a)

Summary

Introduction

Ribonuclease P (RNase P) is an essential ribozyme responsible for tRNA 5′ maturation. Archaeal RNase P is an evolutionary intermediate with chimeric features of both bacterial and eukaryal nuclear enzymes, and serves as an excellent system to provide insights into the structural and functional alterations that accompanied the gradual transformation of an ancient catalytic RNA to a protein-rich RNP3,20. To gain insights into the structure and function of archaeal RNase P holoenzyme and its evolutionary relationships with bacterial and eukaryal enzymes, we reconstitute the RNase P holoenzyme from Methanocaldococcus jannaschii (Mtype) and determine its cryo-EM structure alone and in complex with a tRNA substrate. The structures fill a void for the structural insights into the RNase P evolution and provide mechanistic understanding of the catalysis of archaeal RNase P

Methods

Results

Conclusion