GroEL traps dimeric and monomeric unfolding intermediates of citrate synthase

The prokaryotic molecular chaperone GroE is increasingly expressed under he at shock conditions, GroE protects cells by preventing the irreversible agg regation of thermally unfolding proteins. Here, the interaction of GroE wit h thermally unfolding citrate synthase (CS) was dissected into sever;al ste ps that occur before irreversible aggregation, and the conformational state s of the unfolding protein recognized by GroEL were determined, The kinetic analysis of CS unfolding revealed the formation of inactive dimeric and mo nomeric intermediates. GroEL binds both intermediates without affecting the unfolding pathway. Furthermore, the dimeric intermediates are not protecte d against dissociation in the presence of GroEL. Monomeric CS is stably ass ociated with GroEL, thus preventing further irreversible unfolding steps an d subsequent aggregation. During refolding, monomeric CS is encapsulated in side the cavity of GroEL.GroES complexes. Taken together our results sugges t that for protection of cells against heat stress both the ability of GroE L to interact with a large variety of nonnative conformations of proteins a nd the active, GroES-dependent refolding of highly unfolded species are imp ortant.