Abstract
Hsp70 molecular chaperones facilitate protein disaggregation and proper folding through iterative cycles of polypeptide binding and release that are allosterically coupled to ATP binding and hydrolysis. Hsp70s are ubiquitous and highly conserved across all of life; they bind ATP at an N-terminal nucleotide-binding domain (NBD) and client peptides in the substrate-binding domain (SBD). The NBD and SBD are connected by a highly conserved linker segment that is integrated into the NBD when ATP is bound but is flexible when the NBD is nucleotide-free or bound with ADP. Allosteric coupling is lost when the linker is flexible, and the freed SBD binds peptide clients with high affinity. It was recently discovered that Hsp70-ATP is in an equilibrium between a restraining state (R) with little affinity for peptides and a low ATPase activity, and a stimulating state (S) that binds peptides efficiently, but with rapid kinetics, and has a relatively high ATPase activity. While attempting to characterize the S state, crystal structures of DnaK-ATP were obtained that demonstrate intrinsic Hsp70 plasticity that affects binding interactions with substrate peptides. These structures provide insights into intermediate states along transition pathways in the Hsp70 chaperone cycle.