Structural studies on the co-chaperone Hop and its complexes with Hsp90

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• The Hsp-organising protein (Hop) is a co-chaperone that bridges Heat Shock Protein 70 (Hsp70) and Hsp90.
• Hop modulates the ATPase activity of Hsp70 and Hsp90, facilitating client protein transfer.
• The full-length structure of Hop and its precise interaction with Hsp90 remain unclear, despite knowledge of its primary binding to the C-terminal MEEVD motif.

Purpose of the Study:

• To elucidate the structural and binding characteristics of full-length Hop in complex with Hsp90.
• To identify all binding sites of Hop on Hsp90 and understand Hop's role in Hsp90 dimerization.
• To determine the quaternary structure of Hop and its nucleotide-dependent binding to Hsp90.

Main Methods:

• Biophysical analysis using truncation mutants of Hop and Hsp90.
• Analysis of Hsp90 mutants affecting its conformation and N-terminal dimerization.
• Small-angle X-ray scattering (SAXS) for shape reconstruction.
• Studies on nucleotide dependence of Hop-Hsp90 binding.

Main Results:

• Hop binds Hsp90 at the C-terminal MEEVD motif and additional sites in the C-terminal and middle domains.
• The TPR2a domain of Hop is critical for Hsp90 binding and dimerization, with potential involvement of TPR2b.
• Full-length Hop adopts a butterfly-like quaternary structure in solution.
• Hop binds the nucleotide-free, open state of Hsp90, and this interaction is weakened by ATP binding.

Conclusions:

• Hop interacts with Hsp90 at multiple sites, forming a clamp-like structure that may regulate Hsp90 activity.
• Hop's binding to Hsp90 likely involves presenting client proteins to Hsp90's middle domain.
• The proposed model suggests Hop prevents ATP hydrolysis by inhibiting domain association within Hsp90 monomers.