Glutaredoxins employ parallel monothiol-dithiol mechanisms to catalyze thiol-disulfide exchanges with protein disulfides

Glutaredoxins (Grxs) are a family of glutathione (GSH)-dependent thiol-disulfide oxidoreductases. They feature GSH-binding sites that directly connect the reversible redox chemistry of protein thiols to the abundant cellular nonprotein thiol pool GSSG/GSH. This work studied the pathways for oxidatio...

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Bibliographic Details
Published in:Chemical science (Cambridge) 2018-02, Vol.9 (5), p.1173-1183
Main Authors: Ukuwela, Ashwinie A, Bush, Ashley I, Wedd, Anthony G, Xiao, Zhiguang
Format: Article
Language:English
Subjects:
Binding sites
Catalysis
Disulfides
E coli
Enzymes
Glutathione
Interception
Oxidation
Proteins
Speciation
Substrates
Thiols
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Summary:Glutaredoxins (Grxs) are a family of glutathione (GSH)-dependent thiol-disulfide oxidoreductases. They feature GSH-binding sites that directly connect the reversible redox chemistry of protein thiols to the abundant cellular nonprotein thiol pool GSSG/GSH. This work studied the pathways for oxidation of protein dithiols P(SH) and reduction of protein disulfides P(SS) catalyzed by HsGrx1 and EcGrx1. The metal-binding domain HMA4n(SH) was chosen as substrate as it contains a solvent-exposed CysCys motif. Quenching of the reactions with excess iodoacetamide followed by protein speciation analysis ESI-MS allowed interception and characterization of both substrate and enzyme intermediates. The enzymes shuttle between three catalytically-competent forms (Grx(SH)(S ), Grx(SH)(SSG) and Grx(SS)) and employ conserved parallel monothiol and dithiol mechanisms. Experiments with dithiol and monothiol versions of both Grx enzymes demonstrate which monothiol (plus GSSG or GSH) or dithiol pathways dominate a specific oxidation or reduction reaction. Grxs are shown to be a class of versatile enzymes with diverse catalytic functions that are driven by specific interactions with GSSG/GSH.
ISSN:2041-6520
2041-6539
DOI:10.1039/c7sc04416j