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. Author manuscript; available in PMC: 2023 Apr 30.
Published in final edited form as: J Mol Biol. 2022 Jan 4;434(8):167440. doi: 10.1016/j.jmb.2021.167440

Figure 4.

Figure 4.

An ensemble kinetic assay to monitor +1 frameshifting during translocation. (A) A 70SIC harboring fMet-tRNAfMet on the AUG codon in the P site, programmed with the mRNA sequence AUG-CCC-CGU-U, is rapidly mixed with an equimolar mixture of the SufB2-TC, Val-TC, and Arg-TC and formation of the tripeptide fMPV or fMPR is monitored over time. The SufB2-TC is in the transcript-state, while the Val-TC and Arg-TC are in the native-state. In the presence of EF-G-GTP, the fMP-PRE complex is translocated to the P site. (B) If the fMP-PRE complex remains in the 0-frame during translocation, the CGU codon for Arg would enter the A site, allowing synthesis of fMPR. (C) If the fMP-PRE complex undergoes +1 frameshifting during translocation, the GUU codon for Val would enter the A site, allowing synthesis of fMPV. The results show 10% synthesis of fMPR but 90% synthesis of fMPV, indicating that SufB2 distributes between a small sub-population that remains in the 0-frame and a large sub-population that has shifted to the +1-frame during translocation.