Mechanisms of the Hsp70 chaperone system

Abstract

Molecular chaperones of the Hsp70 family have diverse functions in cells. They assist the folding of newly synthesized and stress-denatured proteins, as well as the import of proteins into organelles, and the dissociation of aggregated proteins. The well-conserved Hsp70 chaperones are ATP dependent: binding and hydrolysis of ATP regulates their interactions with unfolded polypeptide substrates, and ATPase cycling is necessary for their function. All cellular functions of Hsp70 chaperones use the same mechanism of ATP-driven polypeptide binding and release. The Hsp40 co-chaperones stimulate ATP hydrolysis by Hsp70 and the type 1 Hsp40 proteins are conserved from Escherichia coli to humans. Various nucleotide exchange factors also promote the Hsp70 ATPase cycle. Recent advances have added to our understanding of the Hsp70 mechanism at a molecular level.

Fig. 1

The Hsp70 and Hsp40 type 1 machinery. (A) Schematic of Hsp70 and Hsp40 domains with original structures shown below. The Hsp70 nucleotide-binding domain (NBD) is divided into 2 lobes made up of subdomains 1a, 1b, 2a, and 2b, with ATP (ADP-PO₄ in the structure) bound in the opening (3HSC in PDB) (Flaherty et al. 1990). A cleft between 1a and 2a may be a regulatory interaction site. The Hsp70 substrate binding domain (SBD) has a base and a helical lid, which hold polypeptide substrate (dark gray) between them, in a groove in the base (1DKX) (Zhu et al. 1996). The Hsp40 J domain is elongated with the Hsp70-interacting HPD motif (coloured) at one end (1HDJ) (Qian et al. 1996). The central region of Hsp40 type 1 proteins has subdomains 1, 2, and 3 arranged in a hooked structure, with the zinc finger motifs at the angle and the main dimerization site at the end of 3. Substrate (dark gray) is bound by subdomain 2 (1NLT) (Li et al. 2003). (B– D) Structures of Hsp70 two-domain constructs: (B) with the NBD and SBD in contact (1YUW) (Jiang et al. 2005); (C) domains separated in the ADP-bound state (2KHO) (Bertelsen et al. 2009); (D) complexed with an Hsp110 co-chaperone (3C7N) (Schuermann et al. 2008). Colours of the domains are as in A., the linker in dark blue, and substrate in dark gray. The orientations of the NBD domains are the same as in A. The NBD of Hsp110 is light blue, the SBD of Hsp110 is indigo. (E) Outline of Hsp70 ATPase cycle. Substrate can be bound by Hsp40 type 1 proteins, which stimulates Hsp70 ATP hydrolysis through their J domains. The J domain of a subunit is connected to the central region by a G/F rich linker (gray bar). Hsp70 in the ATP state has the NBD, SBD and the interdomain linker (black bar) closely packed together, and binds substrate poorly. Hsp70 in the ADP state binds substrate tightly, and Hsp40 dissociates (Hartl and Hayer-Hartl, 2002; Young et al. 2004; Mayer and Bukau, 2005). The domains of Hsp70 in the ADP state may be separate and the linker flexible (black line). Hsp110 has an NBD and SBD structurally related to Hsp70, and binds the Hsp70 NBD to open it and release nucleotide. Hsp70 can then return to the ATP state (Polier et al. 2008; Schuermann et al. 2008). (F) Other modes of interaction of the Hsp70 NBD. The NBD may be in close contact with the interdomain linker and SBD in the ATP state (Liu and Hendrickson, 2007; Swain et al. 2007). The J domain of Hsp40 may interact near the linker binding cleft of the NBD (Jiang et al. 2007). In the ADP state, the SBD may also contact the NBD (Jiang et al. 2005), or rotate separately as shown in B (Swain et al. 2007; Bertelsen et al. 2009). The nucleotide exchange factors (NEF) of the Bag family and HspBP1, like Hsp110, shift NBD subdomain 2b to release nucleotide (Sondermann et al. 2001; Shomura et al. 2005; Andréasson et al. 2008a; Xu et al. 2008).