Overlapping sites of tetratricopeptide repeat protein binding and chaperone activity in heat shock protein 90

The sequential binding of different tetratricopeptide repeat (TPR) proteins to heat shock protein 90 (hsp90) is essential to its chaperone function in vivo. We have previously shown that three basic residues in the TPR domain of PP5 are required for binding to the acidic C-terminal domain of hsp90. We have now tested which acidic residues in this C-terminal domain are requ ired for binding to three different TPR proteins as follows: PP5, FKBP52, a nd Hop. Mutation of Glu-729, Glu-730, and Asp-732 at the C terminus of hsp9 0 interfered with binding of all three TPR proteins. Mutation of Glu-720, A sp-722, Asp-723, and Asp-724 inhibited binding of FKBP52 and PP5 but not of Hop. Mutation of Glu-651 and Asp-653 did not affect binding of FKBP52 or P P5 but inhibited both Hop binding and hsp90 chaperone activity. We also fou nd that a conserved Lys residue required for PP5 binding to hsp90 was criti cal for the binding of FKBP52 but not for the binding of Hop to hsp90. Thes e results suggest distinct but overlapping binding sites on hsp90 for diffe rent TPR proteins and indicate that the binding site for Hop, which is asso ciated with hsp90 in intermediate stages of protein folding, overlaps with a site of chaperone activity.