During protein synthesis, mRNA and tRNAs are iteratively translocated by the ribosome. Precisely what molecular event is rate limiting for translocation is not known. Here we show that disruption of the interactions between the A-site codon and the ribosome accelerates translocation, suggesting that the release of the mRNA from the decoding center of the ribosome is the rate-limiting step of translocation. These results provide insight into the molecular mechanism of translocation.