A comparative study has investigated the in vitro dissociation and self-assembly of chaperonin 60 14-mers isolated from E. coli (GroEL), yeast mitochondria and pea chloroplasts. In all cases Mg2+ inhibits, and low temperature stimulates, the urea-induced dissociation. ATP or ADP in the presence of Mg2+ enhance the dissociation of the chaperonins. Re-assembly of the 14-mers from their monomers shows different efficiencies between the three proteins. In all cases, however, self-assembly is stimulated by Mg-adenine nucleotides. Surprisingly, effective self-assembly of GroEL is promoted by 20% glycerol in the absence of ATP. The role of Mg-adenine nucleotides in the dissociation and assembly of the chaperonins is discussed.