Importance of a tRNA anticodon loop modification and a conserved, noncanonical anticodon stem pairing in tRNAProCGG for decoding

• Type of Material: Article
• Authors: Ha An Nguyen, Emory University; Eric D. Hoffer, Emory University; Christine M. Dunham, Emory University
• Language: English
• Date: 2019-02-19
• Publisher: American Society for Biochemistry and Molecular Biology
• Publication Version: Final Published Version
• Copyright Statement: © 2019 Nguyen et al.
• License: Creative Commons Attribution 4.0 International
• Final Published Version (URL): https://doi.org/10.1074/jbc.RA119.007410
• Title of Journal or Parent Work: Journal of Biological Chemistry
• Volume: 294
• Issue: 14
• Start Page: 5281
• End Page: 5291
• Grant/Funding Agency: National Institutes of Health; Department of Energy; National Science Foundation
• Grant/Funding Information: This work was supported by NIGMS, National Institutes of Health, Grant R01 GM093278 and National Science Foundation Grant CHE 1808711 (to C. M. D.).; This research used resources of the APS, a United States Department of Energy Office of Science User Facility operated by Argonne National Laboratory under Contracts DE-AC02-06CH11357 (NE-CAT) and W-31-109-Eng-38 (SER-CAT).
• Keywords: mRNA frame maintenance; ribosome; transfer RNA (tRNA); translation; mRNA; structural biology; RNA modification; decoding
• Subject - Topics: RNA biology; Structural biology; Translation

Abstract

Modification of anticodon nucleotides allows tRNAs to decode multiple codons, expanding the genetic code. Additionally, modifications located in the anticodon loop, but outside the anticodon itself, stabilize tRNA–codon interactions, increasing decoding fidelity. Anticodon loop nucleotide 37 is 3′ to the anticodon and, in tRNAPro CGG, is methylated at the N1 position in its nucleobase (m1G37). The m1G37 modification in tRNAPro CGG stabilizes its interaction with the codon and maintains the mRNA frame. However, it is unclear how m1G37 affects binding at the decoding center to both cognate and +1 slippery codons. Here, we show that the tRNAPro CGG m1G37 modification is important for the association step during binding to a cognate CCG codon. In contrast, m1G37 prevented association with a slippery CCC-U or +1 codon. Similar analyses of frameshift suppressor tRNASufA6, a tRNAPro CGG derivative containing an extra nucleotide in its anticodon loop that undergoes +1 frameshifting, reveal that m1G37 destabilizes interactions with both the cognate CCG and slippery codons. One reason for this destabilization is the disruption of a conserved U32·A38 nucleotide pairing in the anticodon stem through insertion of G37.5. Restoring the tRNASufA6 U32·A37.5 pairing results in a high-affinity association on the slippery CCC-U codon. Further, an X-ray crystal structure of the 70S ribosome bound to tRNASufA6 U32·A37.5 at 3.6 Å resolution shows a reordering of the anticodon loop consistent with the findings from the high-affinity measurements. Our results reveal how the tRNA modification at nucleotide 37 stabilizes interactions with the mRNA codon to preserve the mRNA frame.