PDBsum entry 3eu4

Oxidoreductase

PDB id: 3eu4

Contents

Protein chain

185 a.a. *

Metals

_CA

Waters ×38

* Residue conservation analysis

PDB id:

3eu4

Name:

Oxidoreductase

Title:

Crystal structure of bdbd from bacillus subtilis (oxidised)

Structure:

Bdbd. Chain: a. Synonym: bacillus disulfide bond protein d. Engineered: yes

Source:

Bacillus subtilis. Organism_taxid: 1423. Gene: bdbd, bsu33480, yvgv. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.30Å R-factor: 0.196 R-free: 0.248

Authors:

A.Crow,M.C.Moller,L.Hederstedt,N.Le Brun

Key ref:

A.Crow et al. (2009). Crystal structure and biophysical properties of Bacillus subtilis BdbD. An oxidizing thiol:disulfide oxidoreductase containing a novel metal site. J Biol Chem, 284, 23719-23733. PubMed id: 19535335 DOI: 10.1074/jbc.M109.005785

Date:

09-Oct-08 Release date: 16-Jun-09

Protein chain

O32218  (BDBD_BACSU) -  Disulfide bond formation protein D from Bacillus subtilis (strain 168)

Seq: 222 a.a.

Struc: 185 a.a.

DOI no: 10.1074/jbc.M109.005785

J Biol Chem 284:23719-23733 (2009)

PubMed id: 19535335

Crystal structure and biophysical properties of Bacillus subtilis BdbD. An oxidizing thiol:disulfide oxidoreductase containing a novel metal site.

A.Crow, A.Lewin, O.Hecht, M.Carlsson Möller, G.R.Moore, L.Hederstedt, N.E.Le Brun.

ABSTRACT

BdbD is a thiol:disulfide oxidoreductase (TDOR) from Bacillus subtilis that functions to introduce disulfide bonds in substrate proteins/peptides on the outside of the cytoplasmic membrane and, as such, plays a key role in disulfide bond management. Here we demonstrate that the protein is membrane-associated in B. subtilis and present the crystal structure of the soluble part of the protein lacking its membrane anchor. This reveals that BdbD is similar in structure to Escherichia coli DsbA, with a thioredoxin-like domain with an inserted helical domain. A major difference, however, is the presence in BdbD of a metal site, fully occupied by Ca(2+), at an inter-domain position some 14 A away from the CXXC active site. The midpoint reduction potential of soluble BdbD was determined as -75 mV versus normal hydrogen electrode, and the active site N-terminal cysteine thiol was shown to have a low pK(a), consistent with BdbD being an oxidizing TDOR. Equilibrium unfolding studies revealed that the oxidizing power of the protein is based on the instability introduced by the disulfide bond in the oxidized form. The crystal structure of Ca(2+)-depleted BdbD showed that the protein remained folded, with only minor conformational changes. However, the reduced form of Ca(2+)-depleted BdbD was significantly less stable than reduced Ca(2+)-containing protein, and the midpoint reduction potential was shifted by approximately -20 mV, suggesting that Ca(2+) functions to boost the oxidizing power of the protein. Finally, we demonstrate that electron exchange does not occur between BdbD and B. subtilis ResA, a low potential extra-cytoplasmic TDOR.

Selected figure(s)

Figure 3.
Active site region of BdbD.A, detailed view of the N terminus of helix α1 of sBdbD showing the Cys-Pro-Ser-Cys active site of sBdbD and the closely lying cis-proline (Pro^193), which is invariant in all thioredoxin-like proteins. B and C, electron density (contoured at 1.0σ) of the active site region of sBdbD in oxidized and reduced states, respectively.

Figure 4.
Ca^2+-binding site of BdbD.A, detailed view (contoured at 1.2 σ) of the Ca^2+-binding site of sBdbD, showing monodentate ligands Gln^49 and Glu^115, bidentate Asp^180, and three crystallographically ordered water molecules. B, an ideal capped octahedral site for comparison with that observed in BdbD. Positions of the idealized capped octahedron correspond to the following: I, Gln^49O-ϵ; II, Glu^115O-ϵ; III–V, waters; VI and VII, Asp^180O-δ1 and Asp^180O-δ2. Ca^2+-ligand distances are indicated.

The above figures are reprinted from an Open Access publication published by the ASBMB: J Biol Chem (2009, 284, 23719-23733) copyright 2009.

Figures were selected by an automated process.