The COOH-terminal peptide binding domain is essential for self-association of the molecular chaperone HSC70.

We have previously shown that the molecular chaperone HSC70 self-associates in solution into dimers, trimers, and probably high order oligomers, according to a slow temperature- and concentration-dependent equilibrium that is shifted toward the monomer upon binding of ATP peptides or unfolded proteins. To determine the structural basis of HSC70 self-association, the oligomerization properties of the isolated amino- and carboxyl-terminal domains of this protein have been analyzed by gel electrophoresis, size exclusion chromatography, and analytical ultracentrifugation. Whereas the amino-terminal ATPase domain (residues 1-384) was found to be monomeric in solution even at high concentrations, the carboxyl-terminal peptide binding domain (residues 385-646) exists as a slow temperature- and concentration-dependent equilibrium involving monomers, dimers, and trimers. The association equilibrium constant obtained for this domain alone is on the order of 10(5) M-1, very close to that determined previously for the entire protein, suggesting that self-association of HSC70 is determined solely by its carboxyl-terminal domain. Furthermore, oligomerization of the isolated carboxyl-terminal peptide binding domain is, like that of the entire protein, reversed by peptide binding, indicating that self-association of the protein may be mediated by the peptide binding site and, as such, should play a role in the regulation of HSC70 chaperone function. A general model for self-association of HSP70 is proposed in which the protein is in equilibrium between two states differing by the conformation of their carboxyl-terminal domain and their self-association properties.