EFFECT OF NUCLEOTIDES, PEPTIDES, AND UNFOLDED PROTEINS ON THE SELF-ASSOCIATION OF THE MOLECULAR CHAPERONE HSC70

In a previous study, we showed that the molecular chaperone HSC70 self -associates in solution in a reversible and likely unlimited fashion, Here, we examine the influence of nucleotides, nucleotide analogs, pep tides, and unfolded proteins on the self-association properties of thi s protein. Whereas in the presence of ADP, HSC70 exists as a slow, con centration- and temperature-dependent monomer-oligomer equilibrium, in the presence of ATP, the protein is essentially monomeric, indicating that ATP shifts this equilibrium toward the monomer by stabilizing th e monomer, Dissociation of oligomers into monomers is also obtained wi th the slowly hydrolyzable ATP analogs, adenosine 5'-O-(thiotriphospha te) and 5'-adenylyl-beta,gamma-imidodiphosphate, or the complex betwee n ADP and the phosphate analog, BeF3, indicating that binding but not hydrolysis of ATP is necessary and sufficient for the stabilization of HSC70 monomer, Furthermore, binding of short peptides or permanently unfolded proteins to thepeptide binding site of HSC70 promotes the dis sociation of oligomers into monomers, suggesting that protein substrat es are able to compete with HSC70 for the same binding site, Because t he release of peptides or unfolded proteins from HSC70 has also been s hown to require ATP binding, these results indicate that dissociation of oligomers is controlled by a mechanism similar to that of release o f protein substrates and suggest that binding of HSC70 to itself occur s via the peptide binding site and mimics binding of HSC70 to protein substrates.