Monomeric YoeB toxin retains RNase activity but adopts an obligate dimeric form for thermal stability

• Type of Material: Article
• Authors: Ian J Pavelich, Emory University; Tatsuya Maehigashi, Emory University; Eric D Hoffer, Emory University; Ajchareeya Ruangprasert, Emory University; Stacey J Miles, Emory University; Christine Dunham, Emory University
• Language: English
• Date: 2019-11-04
• Publisher: Oxford University Press
• Publication Version: Final Published Version
• Copyright Statement: © The Author(s) 2019.
• License: Creative Commons Attribution-NonCommercial 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1093/nar/gkz760
• Title of Journal or Parent Work: Nucleic Acids Research
• Volume: 47
• Issue: 19
• Start Page: 10400
• End Page: 10413
• Grant/Funding Information: Burroughs Wellcome Fund Investigator in the Pathogenesis of Infectious Disease award [1015487]; National Institutes of Health [R01GM093278]; NE-CAT beamlines, which are funded by the NIGMS from the NIH [P41 GM103403]; Funding for open access charge: Burroughs Wellcome Fund.; The Pilatus 6M detector on 24-ID-C beam line is funded by a NIH-ORIP HEI grant [S10 RR029205]; National Science Foundation CAREER award [MCB 0953714]; Advanced Photon Source, a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory [DE-AC02-06CH11357]
• Keywords: ESCHERICHIA-COLI; RECOGNITION; MAZF; MESSENGER-RNA; Biochemistry & Molecular Biology; MECHANISM; PROTEIN; RIBOSOMAL-RNA; Life Sciences & Biomedicine; Science & Technology; CLEAVAGE; TRANSCRIPTIONAL REPRESSION; INHIBITS TRANSLATION
• Research Categories: Chemistry, Biochemistry; Chemistry, General; Health Sciences, Medicine and Surgery

Abstract

Chromosomally-encoded toxin-antitoxin complexes are ubiquitous in bacteria and regulate growth through the release of the toxin component typically in a stress-dependent manner. Type II ribosome-dependent toxins adopt a RelE-family RNase fold and inhibit translation by degrading mRNAs while bound to the ribosome. Here, we present biochemical and structural studies of the Escherichia coli YoeB toxin interacting with both a UAA stop and an AAU sense codon in pre- and post-mRNA cleavage states to provide insights into possible mRNA substrate selection. Both mRNAs undergo minimal changes during the cleavage event in contrast to type II ribosome-dependent RelE toxin. Further, the 16S rRNA decoding site nucleotides that monitor the mRNA in the aminoacyl(A) site adopt different orientations depending upon which toxin is present. Although YoeB is a RelE family member, it is the sole ribosome-dependent toxin that is dimeric. We show that engineered monomeric YoeB is active against mRNAs bound to both the small and large subunit. However, the stability of monomeric YoeB is reduced ∼20°C, consistent with potential YoeB activation during heat shock in E. coli as previously demonstrated. These data provide a molecular basis for the ability of YoeB to function in response to thermal stress.