Molecular chaperone Hsp70/Hsp90 prepares the mitochondrial outer membrane translocon receptor Tom71 for preprotein loading - PubMed (original) (raw)

Molecular chaperone Hsp70/Hsp90 prepares the mitochondrial outer membrane translocon receptor Tom71 for preprotein loading

Jingzhi Li et al. J Biol Chem. 2009.

Abstract

The preproteins targeted to the mitochondria are transported through the translocase of the outer membrane complex. Tom70/Tom71 is a major surface receptor of the translocase of the outer membrane complex for mitochondrial preproteins. The preproteins are escorted to Tom70/Tom71 by molecular chaperones Hsp70 and Hsp90. Here we present the high resolution crystal structures of Tom71 and the protein complexes between Tom71 and the Hsp70/Hsp90 C terminus. The crystal structures indicate that Tom70/Tom71 may exhibit two distinct states. In the closed state, the N-terminal domain of Tom70/Tom71 partially blocks the preprotein-binding pocket. In the open state, the N-terminal domain moves away, and the preprotein-binding pocket is fully exposed. The complex formation between the C-terminal EEVD motif of Hsp70/Hsp90 and Tom71 could lock Tom71 in the open state where the preprotein-binding pocket of Tom71 is ready to receive preproteins. The interactions between Hsp70/Hsp90 and Tom71 N-terminal domain generate conformational changes that may increase the volume of the preprotein-binding pocket. The complex formation of Hsp70/Hsp90 and Tom71 also generates significant domain rearrangement within Tom71, which may position the preprotein-binding pocket closer to Hsp70/Hsp90 to facilitate the preprotein transfer from the molecular chaperone to Tom71. Therefore, molecular chaperone Hsp70/Hsp90 may function to prepare the mitochondrial outer membrane receptor Tom71 for preprotein loading.

PubMed Disclaimer

Figures

FIGURE 1.

FIGURE 1.

The yeast Tom71 structure. a, ribbon drawing of the yeast Tom71 monomer structure (open state) in side-by-side stereo mode (31). The N- and C-terminal domains of Tom71 are shown in light blue and green, respectively. The N (N-ter) and C termini (C-ter) are labeled. TPR1–TPR3 and TPR9–TPR11 are labeled. The missing residues in the electron density map are shown in dotted lines. b, ribbon drawing of the yeast Tom71 monomer structure in side-by-side stereo mode. The Tom71 structure in this figure is rotated ∼90° along the horizontal axis from that in a. The N and C termini of the structure are labeled. TPR1–TPR11 are labeled. c, ribbon drawing of the yeast Tom70 monomer structure (closed state) in side-by-side stereo mode. The coloring of the N- and C-terminal domains of yeast Tom70 is the same as a. d, the sequence conservation drawing for Tom71. Tom71 in this figure is in similar orientation as that in a. The sequence conservation score obtained by sequence alignment (see Fig. 2 for details) is mapped to the Tom71 molecular surface by Pymol. The red color denotes the conserved regions. The Hsp70/Hsp90-binding site and the preprotein-binding pocket located on the Tom71 surface are indicated by dotted circles. To indicate the size of the preprotein-binding pocket of Tom71, the distance (27 Å) between Pro234 and Phe496 is shown by an arrow. e, the hydrophobicity drawing of Tom71 monomer by Pymol. Gold color denotes hydrophobic regions. Tom71 in this figure is rotated ∼90° along the horizontal axis from that in d and is in similar orientation as that in b. The distance (27 Å) between Pro234 and Phe496 is shown by an arrow. The blue box covers the preprotein-binding pocket, which will be amplified in f. f, this figure shows the magnified version of the area within the blue box in the e. Some conserved hydrophobic residues involved in forming the preprotein-binding pocket are labeled.

FIGURE 2.

FIGURE 2.

Sequence alignment of the Tom71/Tom70 family members. Program ClustalW was utilized to align the Tom71 sequences from S. cerevisiae (Sc Tom71) with that from C. albicans (Ca Tom71) and Tom70 from S. cerevisiae (Sc Tom70), H. sapiens (Hs Tom70), and D. melanogaster (Dm Tom70). The amino acid residues of yeast Tom71 are numbered above the alignment. The eleven TPR motifs (TPR1–TPR11) within Tom71 are labeled. Helices A0–A27 are labeled. The conserved residues involved in binding the Hsp70/Hsp90 C-terminal EEVD motifs are labeled with blue bars. The conserved residues involved in mediating the conformational changes generated by Hsp70/Hsp90 binding are marked with pink bars. The conserved hydrophobic residues forming the Tom70p preprotein-binding pocket among the family are marked with green bars.

FIGURE 3.

FIGURE 3.

The complex structures of Tom71 and the Hsp70/Hsp90 C-terminal EEVD motif. a, the surface potential drawing of Tom71 complexed with Hsp70 C-terminal peptide PTVEEVD. Tom71 is shown in surface potential drawing generated by Pymol and Apbs. Blue and red denote positively and negatively electrostatic potentials, respectively. The bound Hsp70 peptide is shown in a rod model. In the rod model, carbon atoms are shown in green, oxygen atoms are shown in red, and nitrogen atoms are shown in blue. To indicate the enlargement of the preprotein-binding pocket of Tom71 after Hsp70 binding, the distance (32 Å) between Pro234 and Phe496 of Tom71 is shown. b, the surface potential drawing of Tom71 N-terminal domain interacting with the Hsp70 C-terminal peptide PTVEEVD in a rod model. The residues of Tom71 involved in binding Hsp70 are labeled in green, and the residues of Hsp70 peptide PTVEEVD are labeled in black. c, ribbon drawing of Tom71 N-terminal domain complexed with Hsp70 C terminus in stereo mode. Tom71 is in a silver ribbon drawing, and the Hsp70 C-terminal peptide is in a solid rod model. The residues of Tom71 involved in binding Hsp70 are drawn in dotted rod model and labeled in blue. The residues of Hsp70 peptide are labeled in black. d, the surface potential drawing of Tom71 N-terminal domain interacting with Hsp90 C-terminal peptide MEEVD in a rod model. The residues of Tom71 involved in binding Hsp90 are labeled in green, and the residues of Hsp90 peptide MEEVD are labeled in black.

FIGURE 4.

FIGURE 4.

The conformational changes of Tom71 generated by Hsp70 binding. a, the N-terminal domain of Tom71 is superimposed with that in the Tom71-Hsp70 complex and they are shown by a ribbon drawing. The Tom71 N-terminal domain is in light blue. The Tom71 N-terminal domain within the Tom71-Hsp70 complex is in silver. The bound Hsp70 C-terminal peptide is in red. Helices A5, A6, and A7 are labeled in blue. Some residues of Tom71 involved in generating the conformational changes are labeled in black. Residues Lys196, Arg200, and Leu199 of Tom71 involved in binding Hsp70 are labeled. The residues forming hydrophobic cluster to associate A5, A6, and A7 are labeled. Glu206 and Arg238 linking A5 and A7 by forming a salt bridge are also labeled. b, Cα trace drawings of yeast Tom71 structure and the Tom71-Hsp70 complex structure. The N-terminal domain of Tom71 is superimposed with that in the Tom71-Hsp70 complex structure. The molecules in this figure are in a similar orientation as in a. The uncomplexed Tom71 structure is in purple. In the Tom71-Hsp70 C terminus complex, Tom71 is in green, and the Hsp70 C terminus is in red. The N- and C-terminal domains of Tom71 are labeled. Helix A7 acting as the hinge to connect the N-and C-terminal domains of Tom71 is labeled.

FIGURE 5.

FIGURE 5.

The cartoon drawing for the mechanism how Hsp70/Hsp90 prepares Tom70/Tom71 for preprotein loading. a, Tom70/Tom71 may exhibit two distinct states: the open and closed state. The Tom70/Tom71 molecule is shown in blue. The N- and C-terminal domains are labeled. The mitochondria outer membrane is shown in orange. b, the interactions between the Hsp70/Hsp90 C-terminal EEVD motif will lock the Tom70/Tom71 in the open state. The Hsp70/Hsp90 is shown in gold. The binding between Hsp70/Hsp90 and Tom71 could increase the volume of the preprotein-binding pocket. The complex formation might rotate the Tom71 C-terminal domain ∼20° back toward the closed state and therefore position the preprotein-binding pocket closer to the Hsp70/Hsp90. The Hsp70/Hsp90 EEVD motif is shown as a red arrow. The preprotein is shown as a green triangle. c, Hsp70/Hsp90 will then load the preprotein into the enlarged preprotein-binding pocket of Tom70/Tom71.

Similar articles

Cited by

References

    1. Sickmann A., Reinders J., Wagner Y., Joppich C., Zahedi R., Meyer H. E., Schönfisch B., Perschil I., Chacinska A., Guiard B., Rehling P., Pfanner N., Meisinger C. (2003) Proc. Natl. Acad. Sci. U.S.A. 100,13207–13212 - PMC - PubMed
    1. Gray M. W., Burger G., Lang B. F. (1999) Science 283,1476–1481 - PubMed
    1. Neupert W. (1997) Annu. Rev. Biochem. 66,863–917 - PubMed
    1. Neupert W., Brunner M. (2002) Nat. Rev. Mol. Cell Biol. 3,555–565 - PubMed
    1. Pfanner N. (2000) Curr. Biol. 10,R412–415 - PubMed

Publication types

MeSH terms

Substances

Grants and funding

LinkOut - more resources