Structure of a bacterial homologue of vitamin K epoxide reductase

Li, Weikai; Schulman, Sol; Dutton, Rachel J.; Boyd, Dana; Beckwith, Jon; Rapoport, Tom A.

doi:10.1038/nature08720

Cited by 178 publications

(343 citation statements)

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“…A similar reaction mechanism has been proposed for the mammalian VKOR, with a Trx-like protein in the ER lumen serving as its redox partner (4,5,9). However, conflicting topology models with three or four TM segments have been proposed for human VKOR (6,9,11); the loop cysteines interacting with the Trx-like protein would thus be on either the cytosolic or luminal side of the ER membrane.…”

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confidence: 98%

“…However, conflicting topology models with three or four TM segments have been proposed for human VKOR (6,9,11); the loop cysteines interacting with the Trx-like protein would thus be on either the cytosolic or luminal side of the ER membrane. In addition, both the cytosolic thioredoxin protein and the luminal PDI protein have been shown to support the VKOR reaction in vitro (4,5,12,13).…”

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confidence: 99%

“…1A). A recent crystal structure of the VKOR homolog from Synechococcus sp., in which the enzyme and a Trx-like domain are contained in a single polypeptide chain, revealed many details of this reaction (9). In a first step (Fig.…”

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confidence: 99%

“…These include several classes of bacteria in which these enzymes are involved in disulfide bridge formation of newly synthesized secretory proteins (7)(8)(9)(10). The reduced cysteines of exported proteins are initially oxidized by a Trx-like protein in the periplasm, leading to the reduction of the disulfide bridge in the active site CXXC motif of the Trx-like protein.…”

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Vitamin K epoxide reductase prefers ER membrane-anchored thioredoxin-like redox partners

Schulman

Wang

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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Vitamin K epoxide reductase (VKOR) sustains blood coagulation by reducing vitamin K epoxide to the hydroquinone, an essential cofactor for the γ-glutamyl carboxylation of many clotting factors. The physiological redox partner of VKOR remains uncertain, but is likely a thioredoxin-like protein. Here, we demonstrate that human VKOR has the same membrane topology as the enzyme from Synechococcus sp., whose crystal structure was recently determined. Our results suggest that, during the redox reaction, Cys43 in a luminal loop of human VKOR forms a transient disulfide bond with a thioredoxin (Trx)-like protein located in the lumen of the endoplasmic reticulum (ER). We screened for redox partners of VKOR among the large number of mammalian Trx-like ER proteins by testing a panel of these candidates for their ability to form this specific disulfide bond with human VKOR. Our results show that VKOR interacts strongly with TMX, an ER membrane-anchored Trx-like protein with a unique CPAC active site. Weaker interactions were observed with TMX4, a close relative of TMX, and ERp18, the smallest Trx-like protein of the ER. We performed a similar screen with Ero1-α, an ER-luminal protein that oxidizes the Trx-like protein disulfide isomerase. We found that Ero1-α interacts with most of the tested Trx-like proteins, although only poorly with the membrane-anchored members of the family. Taken together, our results demonstrate that human VKOR employs the same electron transfer pathway as its bacterial homologs and that VKORs generally prefer membrane-bound Trx-like redox partners.disulfide bond formation | warfarin | quinone | blood coagulation | electron transfer

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Vitamin K epoxide reductase prefers ER membrane-anchored thioredoxin-like redox partners

Schulman

Wang

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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“…Alternatively, attack by the C B from the accepting C A XXC B motif restores the initial conditions. It is possible to make the mixed disulfide resistant to resolution and capture it for observation by eliminating the C B cysteines, [6][7][8]10,11 but at the expense of losing key players in the reaction.…”

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confidence: 99%

Enzyme structure captures four cysteines aligned for disulfide relay

et al. 2014

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Thioredoxin superfamily proteins introduce disulfide bonds into substrates, catalyze the removal of disulfides, and operate in electron relays. These functions rely on one or more dithiol/ disulfide exchange reactions. The flavoenzyme quiescin sulfhydryl oxidase (QSOX), a catalyst of disulfide bond formation with an interdomain electron transfer step in its catalytic cycle, provides a unique opportunity for exploring the structural environment of enzymatic dithiol/disulfide exchange. Wild-type Rattus norvegicus QSOX1 (RnQSOX1) was crystallized in a conformation that juxtaposes the two redox-active di-cysteine motifs in the enzyme, presenting the entire electron-transfer pathway and proton-transfer participants in their native configurations. As such a state cannot generally be enriched and stabilized for analysis, RnQSOX1 gives unprecedented insight into the functional group environments of the four cysteines involved in dithiol/disulfide exchange and provides the framework for analysis of the energetics of electron transfer in the presence of the bound flavin adenine dinucleotide cofactor. Hybrid quantum mechanics/molecular mechanics (QM/MM) free energy simulations based on the X-ray crystal structure suggest that formation of the interdomain disulfide intermediate is highly favorable and secures the flexible enzyme in a state from which further electron transfer via the flavin can occur.

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Oxidative Protein Folding

Gennaris

Collet

2012

Encyclopedia of Life Sciences

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Oxidative protein folding is the process by which a protein recovers both its native structure and its native disulphide bonds. Disulphide bonds are vital for the correct folding of many secreted proteins, such as insulin, albumin, tissue plasminogen activator and antibodies. The formation of a disulphide bond between two cysteine residues is a rate‐limiting step of the folding process. Therefore, living cells contain proteins that catalyse this reaction (DsbA, protein disulphide isomerase (PDI), Mia40). Pathways that form disulphide bonds have now been unraveled in the bacterial periplasm, the endoplasmic reticulum and the mitochondrial intermembrane space. These pathways have in common to form a relay that channels the electrons released upon cysteine oxidation to a final electron acceptor. Key Concepts: Disulphide bond formation is required for the correct folding of many secreted proteins. Disulphide bond formation is a catalysed process in vivo . Pathways of disulphide bond formation are found both in prokaryotes and in eukaryotes. Pathways of disulphide bond formation form a relay that channels the electrons away from the substrate protein to a final electron acceptor. Pathways of disulphide bond formation involve a direct disulphide donor and a disulphide generator. Formation of native disulphides in proteins with multiple cysteine residues involves a disulphide isomerase. A disulphide formation pathway and a disulphide isomerisation pathway co‐exist in bacteria such as Escherichia coli . Protein disulphide isomerase (PDI) catalyses both disulphide bond formation and disulphide bond isomerisation in the endoplasmic reticulum.

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Structure of a bacterial homologue of vitamin K epoxide reductase

Cited by 178 publications

References 43 publications

Vitamin K epoxide reductase prefers ER membrane-anchored thioredoxin-like redox partners

Vitamin K epoxide reductase prefers ER membrane-anchored thioredoxin-like redox partners

Enzyme structure captures four cysteines aligned for disulfide relay

Oxidative Protein Folding

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