Journal
RNA
Volume 20, Issue 12, Pages 1944-1954Publisher
COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT
DOI: 10.1261/rna.046953.114
Keywords
ribosome; mRNA; reading frame; anticodon stem-loop; decoding; X-ray crystal structure
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Funding
- Department of Defense through the National Defense Science and Engineering Graduate Fellowship Program
- National Institutes of Health (NIH) Training Grant [T32 GM8367]
- National Institute of General Medical Sciences of the NIH Award [R01GM093278]
- US DOE [DE-AC02-06CH11357]
- National Center for Research Resources [NIH] Award [RR-15301]
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The three-nucleotide mRNA reading frame is tightly regulated during translation to ensure accurate protein expression. Translation errors that lead to aberrant protein production can result from the uncoupled movement of the tRNA in either the 5' or 3' direction on mRNA. Here, we report the biochemical and structural characterization of + 1 frameshift suppressor tRNA(SufJ), a tRNA known to decode four, instead of three, nucleotides. Frameshift suppressor tRNA(SufJ) contains an insertion 5' to its anticodon, expanding the anticodon loop from seven to eight nucleotides. Our results indicate that the expansion of the anticodon loop of either ASL(SufJ) or tRNA(SufJ) does not affect its affinity for the A site of the ribosome. Structural analyses of both ASL(SufJ) and ASL(Thr) bound to the Thermus thermophilus 70S ribosome demonstrate both ASLs decode in the zero frame. Although the anticodon loop residues 34-37 are superimposable with canonical seven-nucleotide ASLs, the single C31.5 insertion between nucleotides 31 and 32 in ASL(SufJ) imposes a conformational change of the anticodon stem, that repositions and tilts the ASL toward the back of the A site. Further modeling analyses reveal that this tilting would cause a distortion in full-length A-site tRNA(SufJ) during tRNA selection and possibly impede gripping of the anticodon stem by 16S rRNA nucleotides in the P site. Together, these data implicate tRNA distortion as a major driver of noncanonical translation events such as frameshifting.
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