Ribosome modulation factor (RMF) is an Escherichia coli protein associated with 100S ribosome dimers, which are formed at stationary growth phase or in slowly growing cells. RMF either purified from stationary-phase ribosomes or synthesized by a chemical method was examined for its functions. By adding either natural or synthetic RMF to 70S ribosomes prepared from both exponentially growing and stationary phase cells, 100S ribosome dimers were generated in a concentration-dependent manner. Protein synthesis in vitro was inhibited concomittantly with the formation of 100S ribosomes. The binding of aminoacyl-tRNA to ribosomes was inhibited in parallel. Taken together we propose that RMF is a stationary phase-specific inhibitor of ribosome functions and 100S dimers are stored forms of ribosomes.