CYCLOPHILIN-40 (CYP-40), MAPPING OF ITS HSP90 BINDING DOMAIN AND EVIDENCE THAT FKBP52 COMPETES WITH CYP-40 FOR HSP90 BINDING

The structurally related immunophilins cyclophilin 40 (CyP-40) and FKB P52 have been identified as components of the unactivated estrogen rec eptor. Both immunophilins have a similar molecular architecture that i ncludes a C-terminal segment with a tetratricopeptide repeat (TPR) dom ain predicted to mediate protein interaction. hsp90 is a common cellul ar target for CyP-40 and FKBP52. Deletion mutants of CyP-40 fused to g lutathione S-transferase were immobilized on glutathione-agarose and t hen used in a rapid hsp90 retention assay to define regions of the CyP -40 C terminus that are important for hsp90 binding. Our evidence sugg ests that the TPR domain is not sufficient for stable association of C yP-40 with hsp90 and requires the participation of flanking acidic and basic residues clustered at the N- and C-terminal ends, respectively. Both microdomains are characterized by alpha-helical structures with segregated hydrophobic and charged residues. Corresponding regions wer e identified in FKBP52. By preincubating myometrial cytosol with lysat es containing bacterially expressed FKBP52, we have shown that FKBP52 competes with CyP-40 for hsp90 binding. Our results raise the possibil ity of a mutually exclusive association of CyP-40 and FKBP52 with hsp9 0. This would lead to separate immunophilin-hsp90-receptor complexes a nd place the estrogen receptor under the control of distinct immunophi lin signaling pathways.