Structural Basis of Disulfide Bond Formation in the Bacterial Periplasm and Mammalian ER


Cellular quality control systems rely on the maintenance of protein and redox homoeostasis, in which the formation, reduction and isomerisation of protein disulfide bonds play important roles. The biological kingdoms have evolved catalytic systems that greatly promote oxidative protein folding. Remarkable progress in structural biology in this field has provided molecular views on how protein disulfide bonds are generated de novo and transferred to downstream folding substrates. Very recently, a novel concept has emerged, proposing that multiple oxidative pathways work redundantly, distinctly and in a complementary manner in the endoplasmic reticulum of mammalian cells to efficiently produce large quantities, and a wide variety, of secretory proteins. Thus, the oxidative protein folding network is even more complicated in higher eukaryotes than previously thought.

Key Concepts:

  • Almost all organisms, from bacteria to humans, possess catalytic systems that promote protein‐disulfide bond formation.

  • Disulfide bond formation networks have diversified dramatically, as evidenced by the appearance of a large number of disulfide oxidoreductases in higher eukaryotes.

  • High‐resolution structures have been solved for an increasing number of disulfide oxidoreductases, thereby revealing the structural and mechanistic basis of cellular disulfide bond formation systems.

  • Disulfide bonds play an important role in folding, assembly and stabilisation of various proteins.

  • Disulfide bond formation networks are closely related to the protein and redox homeostasis in the cell.

Keywords: disulfide bond; crystal structure; DsbA ; DsbB ; PDI ; Ero1; Prx4; GPx; VKOR ; QSOX

Figure 1.

Crystal structures and mechanisms of action of E. coli DsbA and DsbB. (a) Ribbon diagram of E. coli DsbA (PDB ID: 1A2M). Cyan and magenta indicate the α‐helix and β‐strands, respectively. The Cys30–Cys33 redox‐active site is represented by sticks. (b) Ribbon diagram of E. coli DsbB (PDB ID: 2ZUQ). The Cys41, Cys44 and Cys104–Cys130 redox‐active sites and bound ubiquinone are represented by sticks. TM, transmembrane helix. (c) Chemical scheme of cooperative de novo disulfide bond generation by DsbB and quinone in E. coli . (d) Molecular mechanisms that prevent backward electron transfer from DsbB to DsbA. TM, transmembrane helix.

Figure 2.

Domain organisation and crystal structures of human PDI and other representative PDI‐family proteins. (a) Domain organisation of PDI, ERp57, ERp72, ERp44, ERp46, P5 and ERdj5 based on their amino acid sequences. Redox‐active and redox‐inactive thioredoxin domains are indicated by white and grey boxes, respectively. Active‐site sequences are indicated in the redox‐active domains. (b) Ribbon diagrams of oxidised (blue; PDB ID: 4EL1) and reduced (magenta; PDB ID: 4EKZ) forms of human PDI. The two structures are superimposed in the panel on the right, such that the root mean square distance of Cα atoms in their abb′ segments is minimised. The region involved in the redox‐dependent rearrangement of a′ is shown in inset. (c) Ribbon diagrams of ERp57 (PDB ID: 3F8U) (gold), ERp72 a0–a (PDB ID: 3IDV) and b–b′ (PDB ID: 3EC3) segments (green), ERp44 (PDB ID: 2R2J) (grey) and ERdj5 (PDB ID: 3APO) (teal). The side chains of Lys214, Lys274 and Arg282 in ERp57 are represented by sticks.

Figure 3.

Crystal structures of human Ero1α and Prx4. (a) Ribbon diagram of the active form of human Ero1α (PDB ID: 3AHQ). The regulatory loop, which could not be modelled due to the lack of information about electron density, is represented by a dotted line. Four regulatory cysteines (Cys94, Cys99, Cys104 and Cys131) converge in this loop. The FAD molecule is represented by sticks. The four helices that constitute the FAD‐embracing bundle are shown in green. This cofactor‐embracing architecture is conserved in bacterial DsbB and VKOR (see Figure b and Figure a). (b) Superimposition of oxidised (yellow, PDB ID: 3TJB) and reduced (cyan, PDB ID: 3TJF) human Prx4 (C51A) decamer structures. (c) Comparison of oxidised (left) and reduced (right) Prx4 dimer structures. Catalytic cysteines are indicated by yellow spheres. The C‐terminal region of oxidised Prx4, which could not be modelled due to a lack of information about electron density, is indicated by dotted line. Exposure of the active‐site disulfide in the oxidised form allows access (red circle) of the thioredoxin domains (Trxs) of PDIs; access is blocked (red ×) in reduced form.

Figure 4.

Overall structures of human GPx8, bacterial VKOR and trypanosomal QSOX. (a) Ribbon diagram of bacterial homologue of VKOR. The crystal structure of Synechococcus sp. VKOR (PDB ID: 3KPQ) is shown. The quinone molecule is represented by sticks. The quinone‐embracing four‐helix bundle is shown in green. Whereas bacterial VKOR is naturally fused to a thioredoxin‐like domain, human VKOR lacks this domain. (b) Crystal structure of the luminal domain of GPx8 (PDB ID: 3KIJ). The structure shows an extended thioredoxin fold. The active‐site cysteine (Cys79) is indicated by yellow spheres. (c) Crystal structure of Trypanosoma brucei QSOX. The FAD molecule is represented by sticks. (Left) Crystal structure of wild‐type QSOX (PDB ID: 3QCP). Trypanosomal QSOX consists of Trx1 (green), a long helix (dark grey), Erv‐like domain (grey) and Erv domain (salmon). The Trx1 active site is 42 Å from the Erv active site (indicated by dashed line). (Right) Crystal structure of QSOX with the C72S/C353S mutation (PDB ID: 3QD9). In this mutant, the active sites in Trx1 and Erv were mutated to CXXC. The structure mimics an intermediate of the intramolecular disulfide transfer step. Compared with wild‐type QSOX, Trx1 is rotated, allowing two redox‐active sites to form an intramolecular disulfide bond. (Mammalian QSOX has a redox‐inactive Trx2 instead of a long helix.) Trx, thioredoxin domain.



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Further Reading

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Kojima, Rieko, Okumura, Masaki, and Inaba, Kenji(Sep 2013) Structural Basis of Disulfide Bond Formation in the Bacterial Periplasm and Mammalian ER . In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1002/9780470015902.a0024169]