Structural insights into RNA unwinding and degradation by RNase R

Abstract

RNase R is a conserved exoribonuclease in the RNase II family that primarily participates in RNA decay in all kingdoms of life. RNase R degrades duplex RNA with a 3′ overhang, suggesting that it has RNA unwinding activity in addition to its 3′-to-5′ exoribonuclease activity. However, how RNase R coordinates RNA binding with unwinding to degrade RNA remains elusive. Here, we report the crystal structure of a truncated form of Escherichia coli RNase R (residues 87–725) at a resolution of 1.85 Å. Structural comparisons with other RNase II family proteins reveal two open RNA-binding channels in RNase R and suggest a tri-helix ‘wedge’ region in the RNB domain that may induce RNA unwinding. We constructed two tri-helix wedge mutants and they indeed lost their RNA unwinding but not RNA binding or degrading activities. Our results suggest that the duplex RNA with an overhang is bound in the two RNA-binding channels in RNase R. The 3′ overhang is threaded into the active site and the duplex RNA is unwound upon reaching the wedge region during RNA degradation. Thus, RNase R is a proficient enzyme, capable of concurrently binding, unwinding and degrading structured RNA in a highly processive manner during RNA decay.

A Novel Cold-Adapted and Salt-Tolerant RNase R from Antarctic Sea-Ice Bacterium Psychrobacter sp. ANT206

“…Based on sequence alignments with the related RNase Rs, the residues (Thr 646, Phe 654, Leu 665 and His 667) were identified as the catalytic sites in the sequence of PsRNR (Figure 1). However, RNase R from E. coli DEC6A was seen to have two catalytic sites (Asp 272 and Asp 281) [15]. Besides, it was reported that two catalytic sites from E. coli were identified in Tyr 324 and Asp 280 [16].…”

Bacterial ribonucleases and their roles in RNA metabolism

“…Based on these findings, the intrinsic helicase activity of RNase R is essential for its proper functioning in vivo . The crystal structure of a truncated form of E. coli RNase R has been determined recently and reveals that the protein has two RNA-binding channels and that it contains a ‘wedge’ region that appears to induce RNA unwinding (Chu et al 2017). How the ‘wedge’ relates to the intrinsic RNA helicase activity remains to be determined.…”

Organellar and Secretory Ribonucleases: Major Players in Plant RNA Homeostasis

“…It was noticed that the impaired maturation of the rrn5S transcript in the mutant is associated with the accumulation of an antisense RNA (asRNA), and this defective maturation was recapitulated in transplastomic plants artificially overexpressing the asRNA (Hotto et al, 2010;Sharwood et al, 2011b). Based on the ability of the E. coli enzyme to unwind RNA duplexes in vitro, it can be speculated that RNR1 might degrade the asRNA in vivo Sharwood et al, 2011b;Chu et al, 2017). A more thorough investigation of the rnr1 RNA patterns also demonstrated the essential role of the enzyme in mRNA 39 maturation, in cooperation with PNPase.…”