Drop-off during ribosome hopping

Ribosomes bypass a 50 nucleotide non-coding segment of mRNA between the two open reading frames of bacteriophage T4 gene 60 in order to synthesize a t opoisomerase subunit. While nearly all ribosomes appear to initiate bypassi ng, only 50% resume translation in the second open reading frame. Failure t o bypass is shown here to be independent of the stop codon at the end of th e first open reading frame and to be amplified by mutant variants of tRNA(2 )(Gly) known to diminish bypassing efficiency. Unproductive bypassing may r esult from premature dissociation of peptidyl-tRNAs from ribosomes (drop-of f) or resumption of translation at inappropriate sites. Assessment of the i nfluence of factors known to induce drop-off reveals that ribosome recyclin g factor accounts for a small fraction of unproductive bypassing products, but none of the other known factors appear to play a significant role. Resu mption of translation at inappropriate sites appears to be minimal, which s uggests that spontaneous release of the peptidyl-tRNA may account for the r emaining unproductive bypassing products and may be inherent to the gene 60 bypassing mechanism. (C) 2001 Academic Press.