The Hsp70 interdomain linker is a dynamic switch that enables allosteric communication between two structured domains

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• Hsp70 molecular chaperones maintain protein homeostasis by binding misfolded proteins.
• This function depends on allosteric communication between the nucleotide-binding domain (NBD) and substrate-binding domain (SBD), mediated by an interdomain linker.
• The linker's flexibility differs between ATP- and ADP-bound states, impacting Hsp70 function.

Purpose of the Study:

• To investigate the conformational landscape of the Hsp70 interdomain linker in the ADP-bound state.
• To elucidate the role of the interdomain linker in Hsp70 allosteric regulation.
• To identify potential therapeutic targets within the Hsp70 allosteric cycle.

Main Methods:

• Molecular dynamics simulations of the interdomain linker in ADP-bound *Escherichia coli* Hsp70 DnaK.
• Experimental validation of simulation findings.
• Analysis of linker conformations, interdomain distances, and transient interactions.

Main Results:

• The interdomain linker adopts a conformation of three ordered segments connected by hinges, limiting NBD-SBD distances and orientations.
• Transient interactions between the linker's C-terminal region and the SBD were observed.
• A key linker-binding site on the SBD is accessible only in the high-affinity substrate-binding state, positioning the linker as an allosteric switch.

Conclusions:

• The Hsp70 interdomain linker functions as a dynamic allosteric switch, crucial for regulating substrate binding.
• The linker's specific conformational states and transient interactions dictate Hsp70's allosteric cycle.
• The linker-binding site on the SBD represents a promising target for developing small molecule modulators of Hsp70 activity.