The Puromycin Reaction and its Inhibition by Chloramphenicol

Puromycin has been used to release nascent peptides from prelabelled Escherichia coli ribosomes. The puromycin reaction on washed ribosomes is partially inhibited by high concentrations of chloramphenicol and may be partially inhibited by low concentrations of chloramphenicol if this is previously bound to ribosomes not engaged in protein synthesis. If crude extracts are allowed to synthesize protein and then chloramphenicol is added during this synthesis ribosomes obtained from such extracts are highly inhibited in the puromycin reaction although only small amounts of chloramphenicol are actually associated with them. The inhibition cannot be reversed by GTP, supernatant fraction, or crude extract and the inhibition is not overcome by lengthy incubation with puromycin. The inhibition may be completely removed in all cases after washing chloramphenicol‐inhibited ribosomes by centrifugation through 15% sucrose.