High-Efficiency “–1” and “–2” Ribosomal Frameshiftings Revealed by Force Spectroscopy

Ribosomal frameshifting is a rare but ubiquitous process that is being studied extensively. Meanwhile, frameshifting motifs without any secondary mRNA structures were identified but rarely studied experimentally. We report unambiguous observation of highly efficient “–1” and “–2” frameshiftings on a GA7G slippery mRNA without the downstream secondary structure, using force-induced remnant magnetization spectroscopy combined with unique probing schemes. The result represents the first experimental evidence of multiple frameshifting steps. It is also one of the rare reports of the “–2” frameshifting. Our assay removed the ambiguity of transcriptional slippage involvement in other frameshifting assays. Two significant insights for the frameshifting mechanism were revealed. First, EF-G·GTP is indispensable to frameshifting. Although EFG·GDPCP has been shown to prompt translocation before, we found that it could not induce frameshifting. This implies that the GTP hydrolysis is responsible for the codon–anticod...