How Thioredoxin can Reduce a Buried Disulphide Bond

Messens, Joris; Molle, Inge Van; Vanhaesebrouck, P.; Limbourg, Maya; Belle, Karolien Van; Wahni, Khadija; Martins, José; Loris, Remy; Wyns, Lode

doi:10.1016/j.jmb.2004.04.016

Cited by 38 publications

(50 citation statements)

References 40 publications

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“…Therefore, the significant conformational exchanges of the Cys 82 -Cys 89 segment in the reduced form are required for the formation of the disulfide bridge between Cys 82 and Cys 89 . On the other hand, the structural geometry and the sub-nanosecond flexibility of the Cys 82 -Cys 89 segment in oxidized ArsC are favorable for the interaction with thioredoxin, which regenerates the reduced catalytically active form of ArsC for the next cycle of arsenate reduction (21). Moreover, Ala 85 undergoes millisecond time scale conformational exchange in addition to sub-nanosecond flexibility in the oxidized form, which may facilitate the subsequent reduction of the enzyme by thioredoxin.…”

Section: Discussionmentioning

confidence: 99%

“…Several families of ArsC have been identified and characterized (10 -14). Among these, Staphylococcus aureus ArsC has been extensively studied (8,(15)(16)(17)(18)(19)(20)(21). These studies revealed that three redox active cysteine residues (Cys 10 , Cys 82 , and Cys 89 ) are critical for arsenate reduction.…”

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confidence: 99%

“…A disulfide bridge (Cys 82 -Cys 89 ) is formed after a single catalytic reaction cycle, converting the enzyme into the inactive form. ArsC is subsequently regenerated by thioredoxin, which converts the enzyme into the reduced form (21). The arsC gene of the Gram-positive bacteria Bacillus subtilis is located in the SKIN element in the chromosome and shares 64% identity with S. aureus ArsC (7,22).…”

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confidence: 99%

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Solution Structures and Backbone Dynamics of Arsenate Reductase from Bacillus subtilis

Guo

Peng

et al. 2005

Journal of Biological Chemistry

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Arsenate reductase encoded by the chromosomal arsC gene in Bacillus subtilis catalyzes the intracellular reduction of arsenate to arsenite, which is then extruded from cells through an efficient and specific transport system. Herein, we present the solution structures and backbone dynamics of both the reduced and oxidized forms of arsenate reductase from B. subtilis. The overall structures of both forms are similar to those of bovine low molecular weight protein-tyrosine phosphatase and arsenate reductase from Staphylococcus aureus. However, several features of the tertiary structure and mobility are notably different between the reduced and oxidized forms of B. subtilis arsenate reductase, particularly in the P-loop region and the segment Cys 82 -Cys 89 . The backbone dynamics results demonstrated that the reduced form of arsenate reductase undergoes millisecond conformational changes in the functional P-loop and Cys 82 -Cys 89 , which may facilitate the formation of covalent intermediates and subsequent reduction of arsenate. In the oxidized form, Cys 82 -Cys 89 shows motional flexibility on both picosecond-to-nanosecond and possibly millisecond time scales, which may facilitate the reduction of the oxidized enzyme by thioredoxin to regenerate the active enzyme. Overall, the internal dynamics and static structures of the enzyme provide insights into the molecular mechanism of arsenate reduction, especially the reversible conformational switch and changes in internal motions associated with the catalytic reaction.

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Solution Structures and Backbone Dynamics of Arsenate Reductase from Bacillus subtilis

Guo

Peng

et al. 2005

Journal of Biological Chemistry

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“…We suggest one way to control oxidation of thiol-containing substrates is to use an intercysteine relay mechanism wherein the second cysteine pair drives certain thiol transfer events. Many different thiol oxidoreductases, including Ero1p, use an intraprotein disulfide transfer event to affect substrate oxidation or reduction, signifying the value of this type of mechanism (Niimura and Massey, 1996;Katzen and Beckwith, 2000;Gross et al, 2002;Kadokura and Beckwith, 2002;Raje and Thorpe, 2003;Argyrou and Blanchard, 2004;Messens et al, 2004). An improved understanding of the specific recognition and oxidation of Pdi1p by Ero1p will likely rely upon defining the association between Pdi1p and the flexible loop domain of Ero1p containing Cys100 and Cys105 at a biochemical and structural level.…”

Section: Internal Disulfide Transfer and Specificitymentioning

confidence: 99%

Disulfide Transfer between Two Conserved Cysteine Pairs Imparts Selectivity to Protein Oxidation by Ero1

Sevier

Kaiser

2006

MBoC

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The membrane-associated flavoprotein Ero1p promotes disulfide bond formation in the endoplasmic reticulum (ER) by selectively oxidizing the soluble oxidoreductase protein disulfide isomerase (Pdi1p), which in turn can directly oxidize secretory proteins. Two redox-active disulfide bonds are essential for Ero1p oxidase activity: Cys100-Cys105 and Cys352-Cys355. Genetic and structural data indicate a disulfide bond is transferred from Cys100-Cys105 directly to Pdi1p, whereas a Cys352-Cys355 disulfide bond is used to reoxidize the reduced Cys100-Cys105 pair through an internal thiol-transfer reaction. Electron transfer from Cys352-Cys355 to molecular oxygen, by way of a flavin cofactor, maintains Cys352-Cys355 in an oxidized form. Herein, we identify a mixed disulfide species that confirms the Ero1p intercysteine thiol-transfer relay in vivo and identify Cys105 and Cys352 as the cysteines that mediate thiol-disulfide exchange. Moreover, we describe Ero1p mutants that have the surprising ability to oxidize substrates in the absence of Cys100-Cys105. We show the oxidase activity of these mutants results from structural changes in Ero1p that allow substrates increased access to Cys352-Cys355, which are normally buried beneath the protein surface. The altered activity of these Ero1p mutants toward selected substrates leads us to propose the catalytic mechanism involving transfer between cysteine pairs evolved to impart substrate specificity to Ero1p.

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“…Thioredoxin-1 acting with peroxiredoxin-1 is an antioxidant that scavenges H 2 O 2 (3). Thioredoxins are also able to reduce buried oxidized thiol residues in proteins (4) and regulate the activity of redox-sensitive transcription factors, including p53 (5), nuclear factor-nB (6), the glucocorticoid receptor (7), activator protein-1 (8), hypoxia-inducible factor-1 (HIF-1; ref. 9), Sp1 (10), and Nrf2 (11).…”

Section: Introductionmentioning

confidence: 99%

Molecular pharmacology and antitumor activity of palmarumycin-based inhibitors of thioredoxin reductase

Powis

Wipf

Lynch

et al. 2006

Molecular Cancer Therapeutics

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The cytosolic thioredoxin redox system composed of thioredoxin-1 and the NADPH-dependent thioredoxin reductase-1 reductase is an important regulator of cell growth and survival. Thioredoxin-1 is overexpressed in many human tumors where it is associated with increased cell proliferation, decreased apoptosis, and decreased patient survival. We hypothesized that thioredoxin reductase-1 provides a target to inhibit the activity of overexpressed thioredoxin-1 for the development of novel anticancer agents. We found that the naphthoquinone spiroketal fungal metabolite palmarumycin CP1 is a potent inhibitor of thioredoxin reductase-1, but attempts to exploit the activity of palmarumycin CP1 analogues as antitumor agents in vivo were hampered by their insolubility. We have therefore developed PX-916, a water-soluble prodrug of a palmarumycin CP1 analogue. PX-916 rapidly releases the parent compound at physiologic pH and in plasma but is stable at acid pH, allowing its i.v. administration. PX-916 is a potent inhibitor of purified human thioredoxin reductase-1 and of thioredoxin reductase-1 activity in cells and tumor xenografts when given to mice and inhibits the downstream targets of thioredoxin-1 signaling, hypoxiainducible factor-1A, and vascular endothelial growth factor in tumors. PX-916 showed excellent antitumor activity against several animal tumor models with some cures. Thus, the study shows that water-soluble inhibitors of thioredoxin reductase-1, such as PX-916, can block thioredoxin-1 signaling in tumors producing marked inhibition of tumor growth. [Mol Cancer Ther 2006; 5(3):630 -6]

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How Thioredoxin can Reduce a Buried Disulphide Bond

Cited by 38 publications

References 40 publications

Solution Structures and Backbone Dynamics of Arsenate Reductase from Bacillus subtilis

Solution Structures and Backbone Dynamics of Arsenate Reductase from Bacillus subtilis

Disulfide Transfer between Two Conserved Cysteine Pairs Imparts Selectivity to Protein Oxidation by Ero1

Molecular pharmacology and antitumor activity of palmarumycin-based inhibitors of thioredoxin reductase

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