Beyond a Passive Tether: Structural Insights into the Disordered Tail of Hsp90

The molecular chaperone
Hsp90 is an abundant and essential
homodimer
that supports the stability and folding of hundreds of client proteins
in cells. Its C-terminal domain (CTD) mediates dimerization and serves
as a docking site for cochaperones bearing tetratricopeptide repeat
(TPR) domains, which recognize the conserved MEEVD motif located at
the end of an intrinsically disordered CTD tail. Despite its conservation,
the structural role of this tail remains poorly understood. We investigated
the conformational behavior of the yeast Hsp90 (yHsp90) CTD tail and
its response to the binding of the TPR-containing PPIase cochaperone
Cpr6. Using site-directed spin labeling with nitroxide and Gd(III)
labels, we examined full-length yHsp90 (FL), isolated CTD (IsoC),
and tail-truncated variants by double electron–electron resonance
(DEER) and EPR spectroscopy. The CTD tails in IsoC and FL adopted
distinct conformational ensembles, attributed to intramolecular interactions
with the middle domain in FL. Cpr6 binding abolished these differences,
indicating disruption of intramolecular tail contacts. In IsoC, the
tail also stabilized an additional CTD conformation near the dimer
interface that was absent in FL and was lost upon tail truncation,
consistent with tail–CTD interactions observed by NMR. This
population was reduced upon Cpr6 binding, shifting the IsoC conformational
distribution toward that of FL. Additionally, this population was
reduced in cellular environments and mimics. Together, our results
demonstrate that the disordered CTD tail is an active structural element
that modulates both its own conformational ensemble and CTD architecture,
highlighting its potential functional relevance in the Hsp90 chaperone
cycle.