PDBsum entry 1e94

Chaperone

PDB id: 1e94

Contents

Protein chains

174 a.a. *

408 a.a. *

Ligands

ANP ×2

Waters ×286

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Mutational studies on hslu and its docking mode with hslv.

Authors

H.K.Song, C.Hartmann, R.Ramachandran, M.Bochtler, R.Behrendt, L.Moroder, R.Huber.

Ref.

Proc Natl Acad Sci U S A, 2000, 97, 14103-14108. [DOI no: 10.1073/pnas.250491797]

PubMed id

11114186

Abstract

HslVU is an ATP-dependent prokaryotic protease complex. Despite detailed crystal and molecular structure determinations of free HslV and HslU, the mechanism of ATP-dependent peptide and protein hydrolysis remained unclear, mainly because the productive complex of HslV and HslU could not be unambiguously identified from the crystal data. In the crystalline complex, the I domains of HslU interact with HslV. Observations based on electron microscopy data were interpreted in the light of the crystal structure to indicate an alternative mode of association with the intermediate domains away from HslV. By generation and analysis of two dozen HslU mutants, we find that the amidolytic and caseinolytic activities of HslVU are quite robust to mutations on both alternative docking surfaces on HslU. In contrast, HslVU activity against the maltose-binding protein-SulA fusion protein depends on the presence of the I domain and is also sensitive to mutations in the N-terminal and C-terminal domains of HslU. Mutational studies around the hexameric pore of HslU seem to show that it is involved in the recognition/translocation of maltose-binding protein-SulA but not of chromogenic small substrates and casein. ATP-binding site mutations, among other things, confirm the essential role of the "sensor arginine" (R393) and the "arginine finger" (R325) in the ATPase action of HslU and demonstrate an important role for E321. Additionally, we report a better refined structure of the HslVU complex crystallized along with resorufin-labeled casein.

Figure 2.
Fig. 2. Representation of the electrostatic potential surfaces of HslV (Left) and HslU (Center) involved in the EM mode of docking. Negatively charged regions are in red, and positively charged regions are in blue. Sites of mutations in the HslU (Right). Numbers 1 (green) and 3 (pink) mark sites of pentaglycine insertions after residues 264 and 387 as well as changes of surface charges (E266Q; E266Q/E385K), 2 (blue) marks the site of introduction of a bulky side chain (I312W), and 4 (red) marks the site of a charge reversal (E436K/D437K). The hexamer pore is colored in yellow. This figure was drawn by using GRASP (28).

Figure 3.
Fig. 3. Sites of mutations in the hexamer pore. Side-chain atoms (yellow) are shown only in one subunit for clarity. Mutation sites in the hexamer pore are colored in pink. Top view of HslU (Left). Side view of the central pore of HslU hexamer (Right). Two subunits from the ring nearest to the reader are removed to expose the interior. This figure was drawn by using GRASP (28).

Secondary reference #1

Title

The structures of hsiu and the ATP-Dependent protease hsiu-Hsiv.

Authors

M.Bochtler, C.Hartmann, H.K.Song, G.P.Bourenkov, H.D.Bartunik, R.Huber.

Ref.

Nature, 2000, 403, 800-805. [DOI no: 10.1038/35001629]

PubMed id

10693812

Figure 1.
Figure 1: Summary of the three crystal forms (a-c) that were used for structure determination. Subunits in the respective asymmetric units are numbered 1-6.

Figure 2.
Figure 2: Comparison of HsIU and NSF main chains. a, Superposition of the ligand-bound (coloured) and free (white) HslU forms. Chains 1 and 2 of the P321 crystals (see Fig. 1c) are shown. The N domains (shown in green and red) have been superimposed (r.m.s.d. C bond lengths = 0.5 Å for the central -sheet, r.m.s.d. C bond lengths = 1.2 Å for the whole domain). For clarity, the N and I domains of the free form have been omitted. b, Stereo diagram of NSF D2.

The above figures are reproduced from the cited reference with permission from Macmillan Publishers Ltd

Secondary reference #2

Title

Crystal structure of heat shock locus V (hslv) from escherichia coli.

Authors

M.Bochtler, L.Ditzel, M.Groll, R.Huber.

Ref.

Proc Natl Acad Sci U S A, 1997, 94, 6070-6074. [DOI no: 10.1073/pnas.94.12.6070]

PubMed id

9177170

Figure 5.
Fig. 5. Overlay of HslV (red) with the T. acidophilum -subunit (green) with bound calpain inhibitors. The secondary structural elements are labeled.

Figure 6.
Fig. 6. Overlay of one hexameric ring of HslV (red) with one heptameric ring of T. acidophilum -subunits (green).

Secondary reference #3

Title

Hslv-Hslu: a novel ATP-Dependent protease complex in escherichia coli related to the eukaryotic proteasome.

Authors

M.Rohrwild, O.Coux, H.C.Huang, R.P.Moerschell, S.J.Yoo, J.H.Seol, C.H.Chung, A.L.Goldberg.

Ref.

Proc Natl Acad Sci U S A, 1996, 93, 5808-5813.

PubMed id

8650174

Secondary reference #4

Title

Sequence analysis of four new heat-Shock genes constituting the hslts/ibpab and hslvu operons in escherichia coli.

Authors

S.E.Chuang, V.Burland, G.Plunkett, D.L.Daniels, F.R.Blattner.

Ref.

Gene, 1993, 134, 1-6. [DOI no: 10.1016/0378-1119(93)90167-2]

PubMed id

8244018