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Abstract

The ribosome can slide along mRNA without establishing codon-anticodon interactions. This movement can be regulated (programmed) by the elements encoded in the mRNA, as observed in bypassing of non-coding gap in gene 60 of bacteriophage T4, or occur spontaneously, such as during traversal by the 70S ribosome of the 3′UTRs or upon re-initiation on bacterial polycistronic genes. In this study, we investigate the kinetic mechanism underlying the programmed and spontaneous ribosome sliding. We show that the translation rate of gene 60 mRNA decreases as the ribosome approaches the take-off site, especially when the KKYK regulatory sequence in the nascent peptide reaches the constriction site in the ribosome exit tunnel. However, efficiency of bypassing increases when the ribosome traverses the gap quickly. With the non-coding gap exceeding the natural 50 nt, the processivity of sliding remains high up to 56 nt, but drops sharply beyond that due to the loss of mRNA elements support. Sliding efficiency is temperature-dependent; while temperature regulates the number of ribosomes initiating programmed bypassing, traversing the long gaps becomes increasingly unfavorable at lower temperatures. This data offers novel insights into the kinetic determinants of programmed and spontaneous ribosome sliding along the mRNA.