After peptide release by a class-1 release factor, the ribosomal subunits must be recycled back to initiation. We have demonstrated that the distance between a strong Shine-Dalgarno (SD) sequence and a codon in the P site is crucial for the binding stability of the deacylated tRNA in the P site of the posttermination ribosome and the in-frame maintenance of its mRNA. We show that the elongation factor EF-G and the ribosomal recycling factor RRF split the ribosome into subunits in the absence of initiation factor 3 (IF3) by a mechanism that requires both GTP and GTP hydrolysis. Taking into account that EF-G in the GTP form and RRF bind with positive cooperativity to the free 50S subunit but with negative cooperativity to the 70S ribosome, we suggest a mechanism for ribosome recycling that specifies distinct roles for EF-G, RRF, and IF3.