Stem–loop structures can effectively substitute for an RNA pseudoknot in -1 ribosomal frameshifting

Yu, Chien-Hung; Noteborn, Mathieu H.; Pleij, Cornelis W. A.; Olsthoorn, René C. L.

The loop composition plays a major role in hairpin stability, RNA/RNA and RNA/protein interaction. These factors may directly influence hairpin-induced ribosomal frameshifting efficiency. To explore the correlation between loop composition and frameshifting efficiency, a number of loop mutations were introduced in the context of a 9 bp stem (Figure 4 ). We note that the UUCG tetraloop with a CG closing base pair (cbp) has higher stability ($2 kcal/mol) than that with a GC cbp (31) . Therefore, we first tested if this different cbp affected frameshifting efficiency. Our results showed that there is no difference in frameshifting efficiency between UUCG and UUCG/cg constructs (Figure 4, bars 6 and 8) . Replacing the UUCG tetraloop by GGGC which, due to its high content of purines, is among the most disfavored tetraloops (32) had only a marginal effect on frameshifting (Figure 4 , compare bars 6 and 7 and Figure 5A, lanes 1 and 4) . Interestingly, increasing the loop size to 9 nt, which is predicted to lower the stability of stem did not affect frameshifting ( Figure 4 , bar 1; Figure 5A , lane 3).