Cpr6 and Cpr7, Two Closely Related Hsp90-associated Immunophilins from Saccharomyces cerevisiae, Differ in Their Functional Properties

Abstract

Hsp90 is an abundant cytosolic molecular chaperone. It controls the folding of target proteins including steroid hormone receptors and kinases in complex with several partner proteins. Prominent members of this protein family are large peptidyl prolyl cis/trans isomerases (PPIases), which catalyze the cis/trans isomerization of prolyl peptide bonds in proteins and possess chaperone activity. In Saccharomyces cerevisiae, two closely related large Hsp90-associated PPIases, Cpr6 and Cpr7, exist. We show here that these homologous proteins bind with comparable affinity to Hsp90 but exhibit significant structural and functional differences. Cpr6 is more stable than Cpr7 against thermal denaturation and displays an up to 100-fold higher PPIase activity. In contrast, the chaperone activity of Cpr6 is much lower than that of Cpr7. Based on these results we suggest that the two immunophilins perform overlapping but not identical tasks in the Hsp90 chaperone cycle. PPIases1 are enzymes that are able to catalyze the cis-trans isomerization of Xaa-Pro peptide bonds in short synthetic peptides as well as in proteins (1). The isomerization of these bonds is a slow process (2) and often a rate-determining step in protein folding (3-6). PPIases have been found in all organisms and subcellular compartments studied so far (1,7,8). Until now three unrelated families of PPIases could be identified: parvulins, cyclophilins, and FKBPs (1,5,9). Because of their inhibition by the immunosuppressive drugs cyclosporin A (CsA) or FK506/rapamycin, the cyclophilins and FKBPs are also termed immunophilins (10 -13). Several large immunophilins with a molecular mass of about 40 -54 kDa are components of the Hsp90 chaperone complex. In higher eukaryotes, FKBP51, FKBP52, and Cyp40 act together with Hsp90; in yeast these are Cpr6 and Cpr7, two cyclophilins. The Hsp90 complex seems to regulate the conformation of its target proteins. These substrates include steroid hormone receptors. Here, the Hsp90 chaperone cycle is well established (reviewed in Refs. 14 -16).In the case of the progesterone receptor, the receptor is bound to Hsp70 in an early complex (15,17). Then an intermediate complex is formed in which Hsp90, Hsp70, Hip, Hop, and perhaps Hsp40 participate. The last step of this activation cycle is a complex, consisting of the progesterone receptor, Hsp90, p23, and one of the large immunophilins. The specific function of the immunophilins in the chaperone cycle is far from clear. However, it is well established that progesterone receptor has to be bound in this complex to allow binding of the hormone, dimerization, and subsequently, interaction with DNA. In the absence of hormone, the receptor is released from the complex, and the cycle starts again. The large immunophilins interact with Hsp90 via tetratricopeptide repeats (18 -21). The partner site for the tetratricopeptide-repeat motifs is located in a 12-kDa carboxyl-terminal domain of Hsp90 (22, 23). The tetratricopeptide-repeat proteins compete with each other for this binding site (18,1...

Chaperoning the Chaperone: A Role for the Co-chaperone Cpr7 in Modulating Hsp90 Function in Saccharomyces cerevisiae

“…Endogenous Cpr6 bound WT Hsp82 only in the presence of AMP-PNP, and both Flag-Cpr6 and FlagCpr7 bound Hsp82D211-264 in the presence or absence of AMP-PNP ( Figure 4C). This is consistent with a prior report that Cpr6 and Cpr7 have similar interactions with purified Hsp82 (Mayr et al 2000).…”

Cns1 Is an Activator of the Ssa1 ATPase Activity

“…For Cpr6 and Cpr7, PPIase activity has been demonstrated previously (26,35,46). Since Cns1 is a suppressor of the cellular defects observed upon deletion of the TPR-containing peptidyl-prolyl isomerase Cpr7, the formal possibility existed that Cns1 also may exhibit prolyl isomerase activity.…”

“…Citrate synthase (CS) and Cpr6 were purified as described (34,35). Ydj1 and the constructs Hsp90-⌬16 and Hsp90-⌬MEEVD were purified as described previously (14,36).…”

Saccharomyces cerevisiae KNU5377 Stress Response during High-Temperature Ethanol Fermentation

“…Hsp104 acts with Hsp26 in protein refolding after stress-induced unfolding (Haslbeck et al, 2005). Hsp90 depends on its association with a variety of cochaperones [Hsp40 (or Hsp42) and cyclophilins], and cofactors (Mayr et al, 2000;Zhao et al, 2005). Hsp90 complex formation is mediated by an adaptor protein, Sti, and an essential ER chaperone, Grp (Grp94) (Chu et al, 2006), which functions as an interaction domain for Hsp90 (Wegele et al, 2003).…”