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. 2010 Jan 11;5(1):e8625.
doi: 10.1371/journal.pone.0008625.

Crystal structures of the ATPase domains of four human Hsp70 isoforms: HSPA1L/Hsp70-hom, HSPA2/Hsp70-2, HSPA6/Hsp70B', and HSPA5/BiP/GRP78

Magdalena Wisniewska  1 , Tobias Karlberg, Lari Lehtiö, Ida Johansson, Tetyana Kotenyova, Martin Moche, Herwig Schüler
Affiliations

Crystal structures of the ATPase domains of four human Hsp70 isoforms: HSPA1L/Hsp70-hom, HSPA2/Hsp70-2, HSPA6/Hsp70B', and HSPA5/BiP/GRP78

Magdalena Wisniewska et al. PLoS One. .

Abstract

The 70-kDa heat shock proteins (Hsp70) are chaperones with central roles in processes that involve polypeptide remodeling events. Hsp70 proteins consist of two major functional domains: an N-terminal nucleotide binding domain (NBD) with ATPase activity, and a C-terminal substrate binding domain (SBD). We present the first crystal structures of four human Hsp70 isoforms, those of the NBDs of HSPA1L, HSPA2, HSPA5 and HSPA6. As previously with Hsp70 family members, all four proteins crystallized in a closed cleft conformation, although a slight cleft opening through rotation of subdomain IIB was observed for the HSPA5-ADP complex. The structures presented here support the view that the NBDs of human Hsp70 function by conserved mechanisms and contribute little to isoform specificity, which instead is brought about by the SBDs and by accessory proteins.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Relationship between the ATPase domains of human Hsp70 isoforms.
Sequence alignment of the NBDs of selected Hsp70 proteins. Secondary structure elements are indicated for HSPA1A above and for HSPA6 below the alignment. Sequences shown are human HSPA1A (1HJO; gi:5123454); HSPA1L (3GDQ; gi:124256496); HSPA2 (3I33; gi:13676857); bovine Hsc70 (PDB entry 1YUW; gi:76253709), E.coli DnaK (1DKG; gi:16128008); HSPA5 (3IUC; gi:16507237); HSPA9 (no structure available; gi:24234688); and HSPA6 (3FE1; gi:34419635).
Figure 2
Figure 2. Purification and crystallization of Hsp70 isoforms.
(A) Coomassie-stained SDS-polyacrylamide gel showing the purity of the crystallized proteins. (B-F) Examples of crystals grown under the conditions that yielded the datasets. (B) HSPA1A; (C) HSPA1L; (D) HSPA2; (E) HSPA5; (F) HSPA6.
Figure 3
Figure 3. Electron density around the nucleotide in the NBDs of HSPA2 and HSPA5.
Representative parts of the 2Fobs-Fcalc density maps around the nucleotide binding site (contoured at 1.5σ above the mean) of (A) HSPA2 with MgADP + Pi, and (B) HSPA5 with CaADP. Selected side chain and main chain atoms within hydrogen bonding distance are indicated.
Figure 4
Figure 4. Crystal structures of the human Hsp70 ATPase domains.
(A) The structure of the HSPA5/BiP NBD in complex with CaADP at 2.4 Å resolution. (B) Superposition of the five Hsp70 NBD structures determined in this study. Dark blue, HSPA1A; yellow, HSPA1L; cyan, HSPA2; red, HSPA6; green, HSPA5/BiP. (C) Cartoon representation of HSPA6, colored to illustrate rms differences in Cα-positions between HSPA6 and HSPA1L (left), HSPA6 and HSPA5 (center), and HSPA6 and DnaK (right).
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